From 0c0197818bf42de5cef0dfbc010c2726630176db Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 3 Dec 2023 13:32:04 -0500 Subject: [PATCH 01/91] feat(derive): new readlen subcommand --- src/derive.rs | 1 + src/derive/command.rs | 4 ++ src/derive/command/readlen.rs | 85 +++++++++++++++++++++++ src/derive/readlen.rs | 3 + src/derive/readlen/compute.rs | 124 ++++++++++++++++++++++++++++++++++ src/main.rs | 3 + 6 files changed, 220 insertions(+) create mode 100644 src/derive/command/readlen.rs create mode 100644 src/derive/readlen.rs create mode 100644 src/derive/readlen/compute.rs diff --git a/src/derive.rs b/src/derive.rs index 4c16afc..30b50f3 100644 --- a/src/derive.rs +++ b/src/derive.rs @@ -2,3 +2,4 @@ pub mod command; pub mod instrument; +pub mod readlen; diff --git a/src/derive/command.rs b/src/derive/command.rs index c372eb4..869a875 100644 --- a/src/derive/command.rs +++ b/src/derive/command.rs @@ -1,6 +1,7 @@ //! Functionality related to the `ngs derive` subcommand itself. pub mod instrument; +pub mod readlen; use clap::Args; use clap::Subcommand; @@ -22,4 +23,7 @@ pub struct DeriveArgs { pub enum DeriveSubcommand { /// Derives the instrument used to produce the file. Instrument(self::instrument::DeriveInstrumentArgs), + + /// Derives the read length of the file. + Readlen(self::readlen::DeriveReadlenArgs), } diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs new file mode 100644 index 0000000..407b20d --- /dev/null +++ b/src/derive/command/readlen.rs @@ -0,0 +1,85 @@ +//! Functionality relating to the `ngs derive readlen` subcommand itself. + +use std::collections::HashMap; +use std::path::PathBuf; + +use clap::Args; +use tracing::info; + +use crate::derive::readlen::compute; +use crate::utils::formats::bam::ParsedBAMFile; +use crate::utils::formats::utils::IndexCheck; + +/// Utility method to parse the Majority Vote Cutoff passed in on the command line and +/// ensure the cutoff is within the range [0.0, 1.0]. +pub fn cutoff_in_range(cutoff_raw: &str) -> Result { + let cutoff: f64 = cutoff_raw + .parse() + .map_err(|_| format!("{} isn't a float", cutoff_raw))?; + + match (0.0..=1.0).contains(&cutoff) { + true => Ok(cutoff), + false => Err(String::from("Error rate must be between 0.0 and 1.0")), + } +} + +/// Clap arguments for the `ngs derive readlen` subcommand. +#[derive(Args)] +pub struct DeriveReadlenArgs { + // Source BAM. + #[arg(value_name = "BAM")] + src: PathBuf, + + /// Only examine the first n records in the file. + #[arg(short, long, value_name = "USIZE")] + num_records: Option, + + /// Majority vote cutoff value as a fraction between [0.0, 1.0]. + #[arg(short, long, value_name = "F64", default_value = "0.7")] + #[arg(value_parser = cutoff_in_range)] + majority_vote_cutoff: Option, +} + +/// Main function for the `ngs derive readlen` subcommand. +pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { + let mut read_lengths = HashMap::new(); + + info!("Starting derive readlen subcommand."); + + let ParsedBAMFile { + mut reader, header, .. + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + + // (1) Collect read lengths from reads within the + // file. Support for sampling only a portion of the reads is provided. + let mut samples = 0; + let mut sample_max = 0; + + if let Some(s) = args.num_records { + sample_max = s; + } + + for result in reader.records(&header.parsed) { + let record = result?; + let len = record.sequence().len(); + + read_lengths.entry(len).and_modify(|e| *e += 1).or_insert(1); + + if sample_max > 0 { + samples += 1; + if samples > sample_max { + break; + } + } + } + + // (2) Derive the consensus read length based on the read lengths gathered. + let result = compute::predict(read_lengths, args.majority_vote_cutoff.unwrap()).unwrap(); + + // (3) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); + + Ok(()) +} diff --git a/src/derive/readlen.rs b/src/derive/readlen.rs new file mode 100644 index 0000000..d6f220e --- /dev/null +++ b/src/derive/readlen.rs @@ -0,0 +1,3 @@ +//! Supporting functionality for the `ngs derive instrument` subcommand. + +pub mod compute; diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs new file mode 100644 index 0000000..27a524f --- /dev/null +++ b/src/derive/readlen/compute.rs @@ -0,0 +1,124 @@ +//! Module holding the logic for computing the consensus read length. + +use anyhow::bail; +use serde::Serialize; +use std::collections::HashMap; + +/// Struct holding the final results for an `ngs derive readlen` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedReadlenResult { + /// Whether or not the `ngs derive readlen` subcommand succeeded. + pub succeeded: bool, + + /// The concsensus read length, if available. + pub consensus_read_length: Option, + + /// The majority vote percentage of the consensus read length, if available. + pub majority_pct_detected: f64, + + /// Status of the evidence that supports (or does not support) this + /// read length, if available. + pub evidence: Vec<(usize, i32)>, +} + +impl DerivedReadlenResult { + /// Creates a new [`DerivedReadlenResult`]. + pub fn new( + succeeded: bool, + consensus_read_length: Option, + majority_pct_detected: f64, + evidence: Vec<(usize, i32)>, + ) -> Self { + DerivedReadlenResult { + succeeded, + consensus_read_length, + majority_pct_detected, + evidence, + } + } +} + +/// Main method to evaluate the collected read lengths and +/// return a result for the consensus read length. This may fail, and the +/// resulting [`DerivedReadlenResult`] should be evaluated accordingly. +pub fn predict( + read_lengths: HashMap, + majority_vote_cutoff: f64, +) -> Result { + let mut num_records = 0; + let mut max_count = 0; + let mut max_read_length = 0; + + for (read_length, count) in &read_lengths { + num_records += *count; + if *read_length > max_read_length { + max_read_length = *read_length; + max_count = *count; + } + } + + if num_records == 0 { + bail!("No read lengths were detected in the file."); + } + + let consensus_read_length = max_read_length; + let majority_detected = max_count as f64 / num_records as f64; + + // Sort the read lengths by their key for output. + let mut read_lengths: Vec<(usize, i32)> = read_lengths.into_iter().collect(); + read_lengths.sort_by(|a, b| b.0.cmp(&a.0)); + + let mut result = + DerivedReadlenResult::new(false, None, majority_detected * 100.0, read_lengths); + + if majority_detected >= majority_vote_cutoff { + result.succeeded = true; + result.consensus_read_length = Some(consensus_read_length); + result.majority_pct_detected = majority_detected * 100.0; + } + + Ok(result) +} + +#[cfg(test)] +mod tests { + use super::*; + + #[test] + fn test_derive_readlen_from_empty_hashmap() { + let read_lengths = HashMap::new(); + let result = predict(read_lengths, 0.7); + assert!(result.is_err()); + } + + #[test] + fn test_derive_readlen_when_all_readlengths_equal() { + let read_lengths = HashMap::from([(100, 10)]); + let result = predict(read_lengths, 1.0).unwrap(); + assert!(result.succeeded); + assert_eq!(result.consensus_read_length, Some(100)); + assert_eq!(result.majority_pct_detected, 100.0); + assert_eq!(result.evidence, Vec::from([(100, 10)])); + } + + #[test] + fn test_derive_readlen_success_when_not_all_readlengths_equal() { + let read_lengths = HashMap::from([(101, 1000), (100, 5), (99, 5)]); + let result = predict(read_lengths, 0.7).unwrap(); + assert!(result.succeeded); + assert_eq!(result.consensus_read_length, Some(101)); + assert!(result.majority_pct_detected > 99.0); + assert_eq!(result.evidence, Vec::from([(101, 1000), (100, 5), (99, 5)])); + } + + #[test] + fn test_derive_readlen_fail_when_not_all_readlengths_equal() { + let read_lengths = HashMap::from([(101, 5), (100, 1000), (99, 5)]); + let result = predict(read_lengths, 0.7).unwrap(); + assert!(!result.succeeded); + assert_eq!(result.consensus_read_length, None); + assert!(result.majority_pct_detected < 1.0); + assert_eq!(result.evidence, Vec::from([(101, 5), (100, 1000), (99, 5)])); + } +} diff --git a/src/main.rs b/src/main.rs index 1441fef..192889a 100644 --- a/src/main.rs +++ b/src/main.rs @@ -92,6 +92,9 @@ fn main() -> anyhow::Result<()> { derive::command::DeriveSubcommand::Instrument(args) => { derive::command::instrument::derive(args)? } + derive::command::DeriveSubcommand::Readlen(args) => { + derive::command::readlen::derive(args)? + } }, Subcommands::Generate(args) => generate::command::generate(args)?, Subcommands::Index(args) => index::command::index(args)?, From 6bb2c2a993b33d62125806e00b28af03052f4232 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 4 Dec 2023 10:10:23 -0500 Subject: [PATCH 02/91] fix(derive/command/readlen): proper error message --- src/derive/command/readlen.rs | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 407b20d..b04e207 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -19,7 +19,9 @@ pub fn cutoff_in_range(cutoff_raw: &str) -> Result { match (0.0..=1.0).contains(&cutoff) { true => Ok(cutoff), - false => Err(String::from("Error rate must be between 0.0 and 1.0")), + false => Err(String::from( + "Majority Vote Cutoff must be between 0.0 and 1.0", + )), } } From 23d8a6cbad60a04b8be390572fbc471fd275c27b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 4 Dec 2023 11:46:43 -0500 Subject: [PATCH 03/91] [WIP] protoype for endedness and skeleton of encoding --- src/derive.rs | 2 + src/derive/command.rs | 8 ++ src/derive/command/encoding.rs | 24 +++++ src/derive/command/endedness.rs | 173 ++++++++++++++++++++++++++++++++ src/derive/encoding.rs | 3 + src/derive/encoding/compute.rs | 1 + src/derive/endedness.rs | 3 + src/derive/endedness/compute.rs | 111 ++++++++++++++++++++ src/main.rs | 6 ++ 9 files changed, 331 insertions(+) create mode 100644 src/derive/command/encoding.rs create mode 100644 src/derive/command/endedness.rs create mode 100644 src/derive/encoding.rs create mode 100644 src/derive/encoding/compute.rs create mode 100644 src/derive/endedness.rs create mode 100644 src/derive/endedness/compute.rs diff --git a/src/derive.rs b/src/derive.rs index 30b50f3..5b45b41 100644 --- a/src/derive.rs +++ b/src/derive.rs @@ -1,5 +1,7 @@ //! Functionality related to the `ngs derive` subcommand. pub mod command; +pub mod encoding; +pub mod endedness; pub mod instrument; pub mod readlen; diff --git a/src/derive/command.rs b/src/derive/command.rs index 869a875..5304207 100644 --- a/src/derive/command.rs +++ b/src/derive/command.rs @@ -1,5 +1,7 @@ //! Functionality related to the `ngs derive` subcommand itself. +pub mod encoding; +pub mod endedness; pub mod instrument; pub mod readlen; @@ -21,6 +23,12 @@ pub struct DeriveArgs { /// All possible subcommands for `ngs derive`. #[derive(Subcommand)] pub enum DeriveSubcommand { + /// Derives the quality score encoding used to produce the file. + Encoding(self::encoding::DeriveEncodingArgs), + + /// Derives the endedness of the file. + Endedness(self::endedness::DeriveEndednessArgs), + /// Derives the instrument used to produce the file. Instrument(self::instrument::DeriveInstrumentArgs), diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs new file mode 100644 index 0000000..f81b550 --- /dev/null +++ b/src/derive/command/encoding.rs @@ -0,0 +1,24 @@ +//! Functionality relating to the `ngs derive encoding` subcommand itself. + +use std::path::PathBuf; + +use anyhow::Ok; +use clap::Args; +use tracing::info; + +/// Clap arguments for the `ngs derive encoding` subcommand. +#[derive(Args)] +pub struct DeriveEncodingArgs { + // Source NGS file (BAM or FASTQ). + #[arg(value_name = "NGS_FILE")] + src: PathBuf, + + /// Only examine the first n records in the file. + #[arg(short, long, value_name = "USIZE")] + num_records: Option, +} + +/// Main function for the `ngs derive encoding` subcommand. +pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { + Ok(()) +} diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs new file mode 100644 index 0000000..a3367e5 --- /dev/null +++ b/src/derive/command/endedness.rs @@ -0,0 +1,173 @@ +//! Functionality relating to the `ngs derive endedness` subcommand itself. + +use std::collections::HashMap; +use std::path::PathBuf; + +use clap::Args; +use noodles::sam::record::data::field::Tag; +use tracing::info; + +use crate::derive::endedness::compute; +use crate::utils::formats::bam::ParsedBAMFile; +use crate::utils::formats::utils::IndexCheck; + +/// Utility method to parse the Paired Deviance passed in on the command line and +/// ensure the value is within the range [0.0, 0.5]. +pub fn deviance_in_range(deviance_raw: &str) -> Result { + let deviance: f64 = deviance_raw + .parse() + .map_err(|_| format!("{} isn't a float", deviance_raw))?; + + match (0.0..=0.5).contains(&deviance) { + true => Ok(deviance), + false => Err(String::from("Paired Deviance must be between 0.0 and 0.5")), + } +} + +/// Clap arguments for the `ngs derive endedness` subcommand. +#[derive(Args)] +pub struct DeriveEndednessArgs { + // Source BAM. + #[arg(value_name = "BAM")] + src: PathBuf, + + /// Only examine the first n records in the file. + #[arg(short, long, value_name = "USIZE")] + num_records: Option, + + /// Distance from 0.5 split between number of f+l- reads and f-l+ reads + /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate + /// if the whole file is being processed. + #[arg(long, value_name = "F64", default_value = "0.0")] + #[arg(value_parser = deviance_in_range)] + paired_deviance: Option, + + /// Calculate and output Reads-Per-Template. This will produce a more + /// sophisticated estimate for endedness, but uses substantially more memory. + #[arg(long, default_value = "false")] + calc_rpt: bool, + + /// Round RPT to the nearest INT before comparing to expected values. + /// Appropriate if using `-n` > 0. + #[arg(long, default_value = "false")] + round_rpt: bool, +} + +/// Main function for the `ngs derive endedness` subcommand. +pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { + info!("Starting derive endedness subcommand."); + + let mut new_ordering_flags: HashMap = HashMap::new(); + new_ordering_flags.insert("f+l-".to_string(), 0); + new_ordering_flags.insert("f-l+".to_string(), 0); + new_ordering_flags.insert("f+l+".to_string(), 0); + new_ordering_flags.insert("f-l-".to_string(), 0); + + let mut ordering_flags: HashMap> = HashMap::new(); + ordering_flags.insert("overall".to_string(), new_ordering_flags.clone()); + ordering_flags.insert("unknown_read_group".to_string(), new_ordering_flags.clone()); + + new_ordering_flags + .entry("f+l-".to_string()) + .and_modify(|e| *e += 1); + new_ordering_flags + .entry("f-l+".to_string()) + .and_modify(|e| *e += 1); + new_ordering_flags + .entry("f+l+".to_string()) + .and_modify(|e| *e += 1); + new_ordering_flags + .entry("f-l-".to_string()) + .and_modify(|e| *e += 1); + + let ParsedBAMFile { + mut reader, header, .. + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + + // (1) Collect read lengths from reads within the + // file. Support for sampling only a portion of the reads is provided. + let mut samples = 0; + let mut sample_max = 0; + + if let Some(s) = args.num_records { + sample_max = s; + } + + for result in reader.records(&header.parsed) { + let record = result?; + + // Only count primary alignments and unmapped reads. + if (record.flags().is_secondary() || record.flags().is_supplementary()) + && !record.flags().is_unmapped() + { + continue; + } + + let read_group = record + .data() + .get(Tag::ReadGroup) + .and_then(|v| v.as_str()) + .unwrap_or("unknown_read_group"); + + if record.flags().is_first_segment() && !record.flags().is_last_segment() { + ordering_flags.entry("overall".to_string()).and_modify(|e| { + e.entry("f+l-".to_string()).and_modify(|e| *e += 1); + }); + ordering_flags + .entry(read_group.to_string()) + .and_modify(|e| { + e.entry("f+l-".to_string()).and_modify(|e| *e += 1); + }) + .or_insert(new_ordering_flags.clone()); + } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { + ordering_flags.entry("overall".to_string()).and_modify(|e| { + e.entry("f-l+".to_string()).and_modify(|e| *e += 1); + }); + ordering_flags + .entry(read_group.to_string()) + .and_modify(|e| { + e.entry("f-l+".to_string()).and_modify(|e| *e += 1); + }) + .or_insert(new_ordering_flags.clone()); + } else if record.flags().is_first_segment() && record.flags().is_last_segment() { + ordering_flags.entry("overall".to_string()).and_modify(|e| { + e.entry("f+l+".to_string()).and_modify(|e| *e += 1); + }); + ordering_flags + .entry(read_group.to_string()) + .and_modify(|e| { + e.entry("f+l+".to_string()).and_modify(|e| *e += 1); + }) + .or_insert(new_ordering_flags.clone()); + } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { + ordering_flags.entry("overall".to_string()).and_modify(|e| { + e.entry("f-l-".to_string()).and_modify(|e| *e += 1); + }); + ordering_flags + .entry(read_group.to_string()) + .and_modify(|e| { + e.entry("f-l-".to_string()).and_modify(|e| *e += 1); + }) + .or_insert(new_ordering_flags.clone()); + } else { + unreachable!(); + } + + if sample_max > 0 { + samples += 1; + if samples > sample_max { + break; + } + } + } + + // (2) Derive the consensus endedness based on the ordering flags gathered. + let result = compute::predict(ordering_flags, args.paired_deviance.unwrap()).unwrap(); + + // (3) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); + + Ok(()) +} diff --git a/src/derive/encoding.rs b/src/derive/encoding.rs new file mode 100644 index 0000000..a20dc0d --- /dev/null +++ b/src/derive/encoding.rs @@ -0,0 +1,3 @@ +//! Supporting functionality for the `ngs derive encoding` subcommand. + +pub mod compute; diff --git a/src/derive/encoding/compute.rs b/src/derive/encoding/compute.rs new file mode 100644 index 0000000..469063d --- /dev/null +++ b/src/derive/encoding/compute.rs @@ -0,0 +1 @@ +//! Module holding the logic for computing the quality score encoding. diff --git a/src/derive/endedness.rs b/src/derive/endedness.rs new file mode 100644 index 0000000..bf321e0 --- /dev/null +++ b/src/derive/endedness.rs @@ -0,0 +1,3 @@ +//! Supporting functionality for the `ngs derive endedness` subcommand. + +pub mod compute; diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs new file mode 100644 index 0000000..f8f5a0d --- /dev/null +++ b/src/derive/endedness/compute.rs @@ -0,0 +1,111 @@ +//! Module holding the logic for computing the endedness of a BAM. + +use anyhow::bail; +use serde::Serialize; +use std::collections::HashMap; + +/// Struct holding the final results for an `ngs derive endedness` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedEndednessResult { + /// Whether or not the `ngs derive endedness` subcommand succeeded. + pub succeeded: bool, + + /// The endedness, if available. + pub endedness: String, + + /// The f+l- read count. + pub first: usize, + + /// The f-l+ read count. + pub last: usize, + + /// The f+l+ read count. + pub both: usize, + + /// The f-l- read count. + pub neither: usize, +} + +impl DerivedEndednessResult { + /// Creates a new [`DerivedEndednessResult`]. + pub fn new( + succeeded: bool, + endedness: String, + first: usize, + last: usize, + both: usize, + neither: usize, + ) -> Self { + DerivedEndednessResult { + succeeded, + endedness, + first, + last, + both, + neither, + } + } +} + +/// Main method to evaluate the collected ordering flags and +/// return a result for the endedness of the file. This may fail, and the +/// resulting [`DerivedEndednessResult`] should be evaluated accordingly. +pub fn predict( + ordering_flags: HashMap>, + paired_deviance: f64, +) -> Result { + let first = ordering_flags["overall"]["f+l-"]; + let last = ordering_flags["overall"]["f-l+"]; + let both = ordering_flags["overall"]["f+l+"]; + let neither = ordering_flags["overall"]["f-l-"]; + + let mut result = + DerivedEndednessResult::new(false, "Unknown".to_string(), first, last, both, neither); + + // all zeroes + if first == 0 && last == 0 && both == 0 && neither == 0 { + bail!("No reads were detected in the file."); + } + + // only first present + if first > 0 && last == 0 && both == 0 && neither == 0 { + return Ok(result); + } + // only last present + if first == 0 && last > 0 && both == 0 && neither == 0 { + return Ok(result); + } + // only both present + if first == 0 && last == 0 && both > 0 && neither == 0 { + result.succeeded = true; + result.endedness = "Single-End".to_string(); + return Ok(result); + } + // only neither present + if first == 0 && last == 0 && both == 0 && neither > 0 { + return Ok(result); + } + // first/last mixed with both/neither + if (first > 0 || last > 0) && (both > 0 || neither > 0) { + return Ok(result); + } + // any mix of both/neither, regardless of first/last + if both > 0 && neither > 0 { + return Ok(result); + } + + // both and neither are now guarenteed to be 0 + // We only need to check first and last + + let first_frac = first as f64 / (first + last) as f64; + let lower_limit = 0.5 - paired_deviance; + let upper_limit = 0.5 + paired_deviance; + if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { + result.succeeded = true; + result.endedness = "Paired-End".to_string(); + return Ok(result); + } + + Ok(result) +} diff --git a/src/main.rs b/src/main.rs index 192889a..6d0e557 100644 --- a/src/main.rs +++ b/src/main.rs @@ -89,6 +89,12 @@ fn main() -> anyhow::Result<()> { match cli.subcommand { Subcommands::Convert(args) => convert::command::convert(args)?, Subcommands::Derive(args) => match args.subcommand { + derive::command::DeriveSubcommand::Encoding(args) => { + derive::command::encoding::derive(args)? + } + derive::command::DeriveSubcommand::Endedness(args) => { + derive::command::endedness::derive(args)? + } derive::command::DeriveSubcommand::Instrument(args) => { derive::command::instrument::derive(args)? } From 5ed13e6530c1a3535e8bb1e52467a96c16045dcc Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 5 Dec 2023 10:30:43 -0500 Subject: [PATCH 04/91] [WIP]: doesn't compile. Begin implementing RPT calculations --- Cargo.lock | 26 +++++++++++++++++ Cargo.toml | 1 + src/derive/command/endedness.rs | 49 ++++++++++++++++++++++++++++++++- 3 files changed, 75 insertions(+), 1 deletion(-) diff --git a/Cargo.lock b/Cargo.lock index 8cdefb3..f6669ee 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -438,6 +438,12 @@ version = "1.0.0" source = "registry+https://github.com/rust-lang/crates.io-index" checksum = "34aa73646ffb006b8f5147f3dc182bd4bcb190227ce861fc4a4844bf8e3cb2c0" +[[package]] +name = "endian-type" +version = "0.1.2" +source = "registry+https://github.com/rust-lang/crates.io-index" +checksum = "c34f04666d835ff5d62e058c3995147c06f42fe86ff053337632bca83e42702d" + [[package]] name = "erased-serde" version = "0.3.23" @@ -865,6 +871,7 @@ dependencies = [ "num-format", "plotly", "prettytable-rs", + "radix_trie", "rand", "rand_distr", "regex", @@ -876,6 +883,15 @@ dependencies = [ "tracing-subscriber", ] +[[package]] +name = "nibble_vec" +version = "0.1.0" +source = "registry+https://github.com/rust-lang/crates.io-index" +checksum = "77a5d83df9f36fe23f0c3648c6bbb8b0298bb5f1939c8f2704431371f4b84d43" +dependencies = [ + "smallvec", +] + [[package]] name = "no-std-compat" version = "0.4.1" @@ -1288,6 +1304,16 @@ dependencies = [ "proc-macro2", ] +[[package]] +name = "radix_trie" +version = "0.2.1" +source = "registry+https://github.com/rust-lang/crates.io-index" +checksum = "c069c179fcdc6a2fe24d8d18305cf085fdbd4f922c041943e203685d6a1c58fd" +dependencies = [ + "endian-type", + "nibble_vec", +] + [[package]] name = "rand" version = "0.8.5" diff --git a/Cargo.toml b/Cargo.toml index b06c9ee..306ad22 100644 --- a/Cargo.toml +++ b/Cargo.toml @@ -35,6 +35,7 @@ noodles = { version = "0.34.0", features = [ num-format = "0.4.0" plotly = "0.8.1" prettytable-rs = "0.9.0" +radix_trie = "0.2.1" rand = "0.8.5" rand_distr = "0.4.3" regex = "1.5.5" diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index a3367e5..7977e17 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -1,11 +1,14 @@ //! Functionality relating to the `ngs derive endedness` subcommand itself. use std::collections::HashMap; +use std::collections::HashSet; use std::path::PathBuf; use clap::Args; use noodles::sam::record::data::field::Tag; +use radix_trie::Trie; use tracing::info; +use tracing::trace; use crate::derive::endedness::compute; use crate::utils::formats::bam::ParsedBAMFile; @@ -53,6 +56,31 @@ pub struct DeriveEndednessArgs { round_rpt: bool, } +struct ReadGroup { + name: String, + first: usize, + last: usize, + both: usize, + neither: usize, +} + +struct FoundReadGroups { + read_groups: HashSet, +} + +impl FoundReadGroups { + fn new() -> Self { + FoundReadGroups { + read_groups: HashSet::new(), + } + } + + fn insert_and_get_ref(&mut self, read_group: &str) -> &String { + self.read_groups.insert(read_group.to_string()); + self.read_groups.get(read_group).unwrap() + } +} + /// Main function for the `ngs derive endedness` subcommand. pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); @@ -80,6 +108,10 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { .entry("f-l-".to_string()) .and_modify(|e| *e += 1); + // only used if args.calc_rpt is true + let mut found_rgs = FoundReadGroups::new(); + let mut read_names = Trie::>::new(); + let ParsedBAMFile { mut reader, header, .. } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; @@ -109,6 +141,21 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { .and_then(|v| v.as_str()) .unwrap_or("unknown_read_group"); + if args.calc_rpt { + let rg_ref = found_rgs.insert_and_get_ref(read_group); + + match record.read_name() { + Some(rn) => { + read_names.insert(rn.to_string(), vec![rg_ref]); + } + None => { + trace!("Could not parse a QNAME from a read in the file."); + trace!("Skipping this read and proceeding."); + continue; + } + } + } + if record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry("overall".to_string()).and_modify(|e| { e.entry("f+l-".to_string()).and_modify(|e| *e += 1); @@ -169,5 +216,5 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let output = serde_json::to_string_pretty(&result).unwrap(); print!("{}", output); - Ok(()) + anyhow::Ok(()) } From c2fe1d86a90457131f5218c663cf111a1ac67b95 Mon Sep 17 00:00:00 2001 From: Clay McLeod Date: Tue, 5 Dec 2023 10:38:49 -0600 Subject: [PATCH 05/91] revise: applies Clay's edits --- src/derive/command/encoding.rs | 1 - src/derive/command/endedness.rs | 53 +++++++++++---------------------- src/derive/endedness/compute.rs | 11 +++---- src/qc/command.rs | 2 +- 4 files changed, 25 insertions(+), 42 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index f81b550..c5692f5 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -4,7 +4,6 @@ use std::path::PathBuf; use anyhow::Ok; use clap::Args; -use tracing::info; /// Clap arguments for the `ngs derive encoding` subcommand. #[derive(Args)] diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 7977e17..902c26b 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -3,6 +3,7 @@ use std::collections::HashMap; use std::collections::HashSet; use std::path::PathBuf; +use std::rc::Rc; use clap::Args; use noodles::sam::record::data::field::Tag; @@ -63,24 +64,6 @@ struct ReadGroup { both: usize, neither: usize, } - -struct FoundReadGroups { - read_groups: HashSet, -} - -impl FoundReadGroups { - fn new() -> Self { - FoundReadGroups { - read_groups: HashSet::new(), - } - } - - fn insert_and_get_ref(&mut self, read_group: &str) -> &String { - self.read_groups.insert(read_group.to_string()); - self.read_groups.get(read_group).unwrap() - } -} - /// Main function for the `ngs derive endedness` subcommand. pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); @@ -91,9 +74,9 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { new_ordering_flags.insert("f+l+".to_string(), 0); new_ordering_flags.insert("f-l-".to_string(), 0); - let mut ordering_flags: HashMap> = HashMap::new(); - ordering_flags.insert("overall".to_string(), new_ordering_flags.clone()); - ordering_flags.insert("unknown_read_group".to_string(), new_ordering_flags.clone()); + let mut ordering_flags: HashMap, HashMap> = HashMap::new(); + ordering_flags.insert(Rc::new("overall".to_string()), new_ordering_flags.clone()); + ordering_flags.insert(Rc::new("unknown_read_group".to_string()), new_ordering_flags.clone()); new_ordering_flags .entry("f+l-".to_string()) @@ -109,8 +92,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { .and_modify(|e| *e += 1); // only used if args.calc_rpt is true - let mut found_rgs = FoundReadGroups::new(); - let mut read_names = Trie::>::new(); + let mut found_rgs = HashSet::new(); + let mut read_names = Trie::>>::new(); let ParsedBAMFile { mut reader, header, .. @@ -138,15 +121,15 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let read_group = record .data() .get(Tag::ReadGroup) - .and_then(|v| v.as_str()) - .unwrap_or("unknown_read_group"); + .map(|v| Rc::new(v.to_string())) + .unwrap_or(Rc::new(String::from("unknown_read_group"))); if args.calc_rpt { - let rg_ref = found_rgs.insert_and_get_ref(read_group); + found_rgs.insert(Rc::clone(&read_group)); match record.read_name() { Some(rn) => { - read_names.insert(rn.to_string(), vec![rg_ref]); + read_names.insert(rn.to_string(), vec![Rc::clone(&read_group)]); } None => { trace!("Could not parse a QNAME from a read in the file."); @@ -157,41 +140,41 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry("overall".to_string()).and_modify(|e| { + ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { e.entry("f+l-".to_string()).and_modify(|e| *e += 1); }); ordering_flags - .entry(read_group.to_string()) + .entry(read_group) .and_modify(|e| { e.entry("f+l-".to_string()).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry("overall".to_string()).and_modify(|e| { + ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { e.entry("f-l+".to_string()).and_modify(|e| *e += 1); }); ordering_flags - .entry(read_group.to_string()) + .entry(read_group) .and_modify(|e| { e.entry("f-l+".to_string()).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry("overall".to_string()).and_modify(|e| { + ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { e.entry("f+l+".to_string()).and_modify(|e| *e += 1); }); ordering_flags - .entry(read_group.to_string()) + .entry(read_group) .and_modify(|e| { e.entry("f+l+".to_string()).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry("overall".to_string()).and_modify(|e| { + ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { e.entry("f-l-".to_string()).and_modify(|e| *e += 1); }); ordering_flags - .entry(read_group.to_string()) + .entry(read_group) .and_modify(|e| { e.entry("f-l-".to_string()).and_modify(|e| *e += 1); }) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index f8f5a0d..e0a2608 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -3,6 +3,7 @@ use anyhow::bail; use serde::Serialize; use std::collections::HashMap; +use std::rc::Rc; /// Struct holding the final results for an `ngs derive endedness` subcommand /// call. @@ -52,13 +53,13 @@ impl DerivedEndednessResult { /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: HashMap>, + ordering_flags: HashMap, HashMap>, paired_deviance: f64, ) -> Result { - let first = ordering_flags["overall"]["f+l-"]; - let last = ordering_flags["overall"]["f-l+"]; - let both = ordering_flags["overall"]["f+l+"]; - let neither = ordering_flags["overall"]["f-l-"]; + let first = ordering_flags[&String::from("overall")]["f+l-"]; + let last = ordering_flags[&String::from("overall")]["f-l+"]; + let both = ordering_flags[&String::from("overall")]["f+l+"]; + let neither = ordering_flags[&String::from("overall")]["f-l-"]; let mut result = DerivedEndednessResult::new(false, "Unknown".to_string(), first, last, both, neither); diff --git a/src/qc/command.rs b/src/qc/command.rs index eeaa29c..0f95bac 100644 --- a/src/qc/command.rs +++ b/src/qc/command.rs @@ -349,7 +349,7 @@ fn app( info!("Starting second pass for QC stats."); let mut reader = File::open(&src).map(bam::Reader::new)?; let index = - bai::read(&src.with_extension("bam.bai")).with_context(|| "reading BAM index")?; + bai::read(src.with_extension("bam.bai")).with_context(|| "reading BAM index")?; let mut counter = RecordCounter::new(); From b4ce76bc75c14584af6217eb3d5a4d84c770aad7 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 6 Dec 2023 09:51:30 -0500 Subject: [PATCH 06/91] [WIP]: Broken, not compiling. starting to calc RPT. --- Cargo.lock | 1 + Cargo.toml | 1 + src/derive/command/endedness.rs | 87 ++++++++++++++++++--------------- src/derive/endedness/compute.rs | 66 +++++++++++++++++++------ 4 files changed, 101 insertions(+), 54 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index f6669ee..925f544 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -867,6 +867,7 @@ dependencies = [ "indexmap", "indicatif", "itertools", + "lazy_static", "noodles", "num-format", "plotly", diff --git a/Cargo.toml b/Cargo.toml index 306ad22..c9081d8 100644 --- a/Cargo.toml +++ b/Cargo.toml @@ -19,6 +19,7 @@ git-testament = "0.2.1" indexmap = "1.9.1" indicatif = "0.16.2" itertools = "0.10.5" +lazy_static = "1.4.0" noodles = { version = "0.34.0", features = [ "async", "bam", diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 902c26b..8bae07c 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -12,6 +12,7 @@ use tracing::info; use tracing::trace; use crate::derive::endedness::compute; +use crate::derive::endedness::compute::{BOTH, FIRST, LAST, NEITHER, OVERALL, UNKNOWN_READ_GROUP}; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -31,7 +32,7 @@ pub fn deviance_in_range(deviance_raw: &str) -> Result { /// Clap arguments for the `ngs derive endedness` subcommand. #[derive(Args)] pub struct DeriveEndednessArgs { - // Source BAM. + /// Source BAM. #[arg(value_name = "BAM")] src: PathBuf, @@ -64,32 +65,25 @@ struct ReadGroup { both: usize, neither: usize, } + /// Main function for the `ngs derive endedness` subcommand. pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); let mut new_ordering_flags: HashMap = HashMap::new(); - new_ordering_flags.insert("f+l-".to_string(), 0); - new_ordering_flags.insert("f-l+".to_string(), 0); - new_ordering_flags.insert("f+l+".to_string(), 0); - new_ordering_flags.insert("f-l-".to_string(), 0); + new_ordering_flags.insert(*FIRST, 0); + new_ordering_flags.insert(*LAST, 0); + new_ordering_flags.insert(*BOTH, 0); + new_ordering_flags.insert(*NEITHER, 0); let mut ordering_flags: HashMap, HashMap> = HashMap::new(); - ordering_flags.insert(Rc::new("overall".to_string()), new_ordering_flags.clone()); - ordering_flags.insert(Rc::new("unknown_read_group".to_string()), new_ordering_flags.clone()); - - new_ordering_flags - .entry("f+l-".to_string()) - .and_modify(|e| *e += 1); - new_ordering_flags - .entry("f-l+".to_string()) - .and_modify(|e| *e += 1); - new_ordering_flags - .entry("f+l+".to_string()) - .and_modify(|e| *e += 1); - new_ordering_flags - .entry("f-l-".to_string()) - .and_modify(|e| *e += 1); + ordering_flags.insert(Rc::new(*OVERALL), new_ordering_flags.clone()); + ordering_flags.insert(Rc::new(*UNKNOWN_READ_GROUP), new_ordering_flags.clone()); + + new_ordering_flags.entry(*FIRST).and_modify(|e| *e += 1); + new_ordering_flags.entry(*LAST).and_modify(|e| *e += 1); + new_ordering_flags.entry(*BOTH).and_modify(|e| *e += 1); + new_ordering_flags.entry(*NEITHER).and_modify(|e| *e += 1); // only used if args.calc_rpt is true let mut found_rgs = HashSet::new(); @@ -118,18 +112,28 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { continue; } - let read_group = record - .data() - .get(Tag::ReadGroup) - .map(|v| Rc::new(v.to_string())) - .unwrap_or(Rc::new(String::from("unknown_read_group"))); + let read_group = match record.data().get(Tag::ReadGroup) { + Some(rg) => Rc::new(String::from(rg.as_str().unwrap())), + None => Rc::new(*UNKNOWN_READ_GROUP), + }; if args.calc_rpt { found_rgs.insert(Rc::clone(&read_group)); match record.read_name() { Some(rn) => { - read_names.insert(rn.to_string(), vec![Rc::clone(&read_group)]); + let rg_vec = read_names.get_mut(&rn.to_string()); + + match rg_vec { + Some(rg_vec) => { + rg_vec.push(Rc::clone(&read_group)); + } + None => { + let mut rg_vec = Vec::new(); + rg_vec.push(Rc::clone(&read_group)); + read_names.insert(rn.to_string(), rg_vec); + } + } } None => { trace!("Could not parse a QNAME from a read in the file."); @@ -140,43 +144,47 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { - e.entry("f+l-".to_string()).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { + e.entry(*FIRST).and_modify(|e| *e += 1); }); + ordering_flags .entry(read_group) .and_modify(|e| { - e.entry("f+l-".to_string()).and_modify(|e| *e += 1); + e.entry(*FIRST).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { - e.entry("f-l+".to_string()).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { + e.entry(*LAST).and_modify(|e| *e += 1); }); + ordering_flags .entry(read_group) .and_modify(|e| { - e.entry("f-l+".to_string()).and_modify(|e| *e += 1); + e.entry(*LAST).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { - e.entry("f+l+".to_string()).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { + e.entry(*BOTH).and_modify(|e| *e += 1); }); + ordering_flags .entry(read_group) .and_modify(|e| { - e.entry("f+l+".to_string()).and_modify(|e| *e += 1); + e.entry(*BOTH).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new("overall".to_string())).and_modify(|e| { - e.entry("f-l-".to_string()).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { + e.entry(*NEITHER).and_modify(|e| *e += 1); }); + ordering_flags .entry(read_group) .and_modify(|e| { - e.entry("f-l-".to_string()).and_modify(|e| *e += 1); + e.entry(*NEITHER).and_modify(|e| *e += 1); }) .or_insert(new_ordering_flags.clone()); } else { @@ -192,7 +200,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } // (2) Derive the consensus endedness based on the ordering flags gathered. - let result = compute::predict(ordering_flags, args.paired_deviance.unwrap()).unwrap(); + let result = + compute::predict(ordering_flags, read_names, args.paired_deviance.unwrap()).unwrap(); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index e0a2608..9e496b9 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,10 +1,25 @@ //! Module holding the logic for computing the endedness of a BAM. use anyhow::bail; +use lazy_static::lazy_static; +use radix_trie::iter; +use radix_trie::Trie; use serde::Serialize; use std::collections::HashMap; use std::rc::Rc; +lazy_static! { + // Strings used to index into the HashMaps used to store the Read Group ordering flags. + pub static ref OVERALL: String = String::from("overall"); + pub static ref UNKNOWN_READ_GROUP: String = String::from("unknown_read_group"); + + // Strings used to index into the HashMaps used to store the ordering flag counts. + pub static ref FIRST: String = String::from("f+l-"); + pub static ref LAST: String = String::from("f-l+"); + pub static ref BOTH: String = String::from("f+l+"); + pub static ref NEITHER: String = String::from("f-l-"); +} + /// Struct holding the final results for an `ngs derive endedness` subcommand /// call. #[derive(Debug, Serialize)] @@ -49,60 +64,81 @@ impl DerivedEndednessResult { } } +fn calculate_reads_per_template( + read_names: Trie>>, +) -> HashMap, f64> { + let mut total_reads: usize = 0; + let mut templates_templates: usize = 0; + let mut reads_per_template: HashMap, f64> = HashMap::new(); + + for read_groups in iter::Iter::new(read_names) {} + + reads_per_template +} + /// Main method to evaluate the collected ordering flags and /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( ordering_flags: HashMap, HashMap>, + read_names: Trie>>, paired_deviance: f64, ) -> Result { - let first = ordering_flags[&String::from("overall")]["f+l-"]; - let last = ordering_flags[&String::from("overall")]["f-l+"]; - let both = ordering_flags[&String::from("overall")]["f+l+"]; - let neither = ordering_flags[&String::from("overall")]["f-l-"]; + let overall_ordering_flags = ordering_flags.get(&*OVERALL).unwrap(); + + let overall_first = *overall_ordering_flags.get(&*FIRST).unwrap(); + let overall_last = *overall_ordering_flags.get(&*LAST).unwrap(); + let overall_both = *overall_ordering_flags.get(&*BOTH).unwrap(); + let overall_neither = *overall_ordering_flags.get(&*NEITHER).unwrap(); - let mut result = - DerivedEndednessResult::new(false, "Unknown".to_string(), first, last, both, neither); + let mut result = DerivedEndednessResult::new( + false, + "Unknown".to_string(), + overall_first, + overall_last, + overall_both, + overall_neither, + ); // all zeroes - if first == 0 && last == 0 && both == 0 && neither == 0 { + if overall_first == 0 && overall_last == 0 && overall_both == 0 && overall_neither == 0 { bail!("No reads were detected in the file."); } // only first present - if first > 0 && last == 0 && both == 0 && neither == 0 { + if overall_first > 0 && overall_last == 0 && overall_both == 0 && overall_neither == 0 { return Ok(result); } // only last present - if first == 0 && last > 0 && both == 0 && neither == 0 { + if overall_first == 0 && overall_last > 0 && overall_both == 0 && overall_neither == 0 { return Ok(result); } // only both present - if first == 0 && last == 0 && both > 0 && neither == 0 { + if overall_first == 0 && overall_last == 0 && overall_both > 0 && overall_neither == 0 { result.succeeded = true; result.endedness = "Single-End".to_string(); return Ok(result); } // only neither present - if first == 0 && last == 0 && both == 0 && neither > 0 { + if overall_first == 0 && overall_last == 0 && overall_both == 0 && overall_neither > 0 { return Ok(result); } // first/last mixed with both/neither - if (first > 0 || last > 0) && (both > 0 || neither > 0) { + if (overall_first > 0 || overall_last > 0) && (overall_both > 0 || overall_neither > 0) { return Ok(result); } // any mix of both/neither, regardless of first/last - if both > 0 && neither > 0 { + if overall_both > 0 && overall_neither > 0 { return Ok(result); } // both and neither are now guarenteed to be 0 // We only need to check first and last - let first_frac = first as f64 / (first + last) as f64; + let first_frac = overall_first as f64 / (overall_first + overall_last) as f64; let lower_limit = 0.5 - paired_deviance; let upper_limit = 0.5 + paired_deviance; - if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { + if (overall_first == overall_last) || (lower_limit <= first_frac && first_frac <= upper_limit) { result.succeeded = true; result.endedness = "Paired-End".to_string(); return Ok(result); From 9aeebf455e1681b078fc34426116db230efe7ca1 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 6 Dec 2023 17:50:31 -0500 Subject: [PATCH 07/91] [WIP] --- src/derive/command/endedness.rs | 78 ++++----- src/derive/endedness/compute.rs | 289 ++++++++++++++++++++++++++------ 2 files changed, 273 insertions(+), 94 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 8bae07c..a7bdf28 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -12,7 +12,7 @@ use tracing::info; use tracing::trace; use crate::derive::endedness::compute; -use crate::derive::endedness::compute::{BOTH, FIRST, LAST, NEITHER, OVERALL, UNKNOWN_READ_GROUP}; +use crate::derive::endedness::compute::{OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP}; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -58,36 +58,17 @@ pub struct DeriveEndednessArgs { round_rpt: bool, } -struct ReadGroup { - name: String, - first: usize, - last: usize, - both: usize, - neither: usize, -} - /// Main function for the `ngs derive endedness` subcommand. pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); - let mut new_ordering_flags: HashMap = HashMap::new(); - new_ordering_flags.insert(*FIRST, 0); - new_ordering_flags.insert(*LAST, 0); - new_ordering_flags.insert(*BOTH, 0); - new_ordering_flags.insert(*NEITHER, 0); - - let mut ordering_flags: HashMap, HashMap> = HashMap::new(); - ordering_flags.insert(Rc::new(*OVERALL), new_ordering_flags.clone()); - ordering_flags.insert(Rc::new(*UNKNOWN_READ_GROUP), new_ordering_flags.clone()); - - new_ordering_flags.entry(*FIRST).and_modify(|e| *e += 1); - new_ordering_flags.entry(*LAST).and_modify(|e| *e += 1); - new_ordering_flags.entry(*BOTH).and_modify(|e| *e += 1); - new_ordering_flags.entry(*NEITHER).and_modify(|e| *e += 1); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Rc::new(&OVERALL), OrderingFlagsCounts::new()); + ordering_flags.insert(Rc::new(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true let mut found_rgs = HashSet::new(); - let mut read_names = Trie::>>::new(); + let mut read_names = Trie::>>::new(); let ParsedBAMFile { mut reader, header, .. @@ -113,8 +94,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => Rc::new(String::from(rg.as_str().unwrap())), - None => Rc::new(*UNKNOWN_READ_GROUP), + Some(rg) => Rc::new(rg.as_str().unwrap()), + None => Rc::new(UNKNOWN_READ_GROUP.as_str()), }; if args.calc_rpt { @@ -144,49 +125,53 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { - e.entry(*FIRST).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { + e.first += 1; }); ordering_flags .entry(read_group) .and_modify(|e| { - e.entry(*FIRST).and_modify(|e| *e += 1); + e.first += 1; }) - .or_insert(new_ordering_flags.clone()); + .or_insert(OrderingFlagsCounts::new()) + .first += 1; } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { - e.entry(*LAST).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { + e.last += 1; }); ordering_flags .entry(read_group) .and_modify(|e| { - e.entry(*LAST).and_modify(|e| *e += 1); + e.last += 1; }) - .or_insert(new_ordering_flags.clone()); + .or_insert(OrderingFlagsCounts::new()) + .last += 1; } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { - e.entry(*BOTH).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { + e.both += 1; }); ordering_flags .entry(read_group) .and_modify(|e| { - e.entry(*BOTH).and_modify(|e| *e += 1); + e.both += 1; }) - .or_insert(new_ordering_flags.clone()); + .or_insert(OrderingFlagsCounts::new()) + .both += 1; } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(*OVERALL)).and_modify(|e| { - e.entry(*NEITHER).and_modify(|e| *e += 1); + ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { + e.neither += 1; }); ordering_flags .entry(read_group) .and_modify(|e| { - e.entry(*NEITHER).and_modify(|e| *e += 1); + e.neither += 1; }) - .or_insert(new_ordering_flags.clone()); + .or_insert(OrderingFlagsCounts::new()) + .neither += 1; } else { unreachable!(); } @@ -200,8 +185,13 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } // (2) Derive the consensus endedness based on the ordering flags gathered. - let result = - compute::predict(ordering_flags, read_names, args.paired_deviance.unwrap()).unwrap(); + let result = compute::predict( + ordering_flags, + read_names, + args.paired_deviance.unwrap(), + args.round_rpt, + ) + .unwrap(); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 9e496b9..670e278 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -2,32 +2,49 @@ use anyhow::bail; use lazy_static::lazy_static; -use radix_trie::iter; use radix_trie::Trie; +use radix_trie::TrieCommon; use serde::Serialize; use std::collections::HashMap; +use std::collections::HashSet; use std::rc::Rc; +use tracing::warn; lazy_static! { // Strings used to index into the HashMaps used to store the Read Group ordering flags. pub static ref OVERALL: String = String::from("overall"); pub static ref UNKNOWN_READ_GROUP: String = String::from("unknown_read_group"); +} - // Strings used to index into the HashMaps used to store the ordering flag counts. - pub static ref FIRST: String = String::from("f+l-"); - pub static ref LAST: String = String::from("f-l+"); - pub static ref BOTH: String = String::from("f+l+"); - pub static ref NEITHER: String = String::from("f-l-"); +pub struct OrderingFlagsCounts { + pub first: usize, + pub last: usize, + pub both: usize, + pub neither: usize, } -/// Struct holding the final results for an `ngs derive endedness` subcommand -/// call. +impl OrderingFlagsCounts { + pub fn new() -> Self { + OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 0, + } + } +} + +/// Struct holding the per read group results for an `ngs derive endedness` +/// subcommand call. #[derive(Debug, Serialize)] -pub struct DerivedEndednessResult { - /// Whether or not the `ngs derive endedness` subcommand succeeded. +struct ReadGroupDerivedEndednessResult { + /// Name of the read group. + pub read_group: String, + + /// Whether or not an endedness was determined for this read group. pub succeeded: bool, - /// The endedness, if available. + /// The endedness of this read group or "Unknown". pub endedness: String, /// The f+l- read count. @@ -41,6 +58,67 @@ pub struct DerivedEndednessResult { /// The f-l- read count. pub neither: usize, + + /// The reads per template (RPT). + /// Only available if `args.calc_rpt` is true. + pub rpt: Option, +} + +impl ReadGroupDerivedEndednessResult { + /// Creates a new [`ReadGroupDerivedEndednessResult`]. + fn new( + read_group: String, + succeeded: bool, + endedness: String, + first: usize, + last: usize, + both: usize, + neither: usize, + rpt: Option, + ) -> Self { + ReadGroupDerivedEndednessResult { + read_group, + succeeded, + endedness, + first, + last, + both, + neither, + rpt, + } + } +} + +/// Struct holding the final results for an `ngs derive endedness` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedEndednessResult { + /// Whether or not the `ngs derive endedness` subcommand succeeded. + pub succeeded: bool, + + /// The overall endedness of the file or "Unknown". + pub endedness: String, + + /// The overall f+l- read count. + pub first: usize, + + /// The overall f-l+ read count. + pub last: usize, + + /// The overall f+l+ read count. + pub both: usize, + + /// The overall f-l- read count. + pub neither: usize, + + /// The overall reads per template (RPT). + /// Only available if `args.calc_rpt` is true. + pub rpt: Option, + + /// Vector of [`ReadGroupDerivedEndednessResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, } impl DerivedEndednessResult { @@ -52,6 +130,8 @@ impl DerivedEndednessResult { last: usize, both: usize, neither: usize, + rpt: Option, + read_groups: Vec, ) -> Self { DerivedEndednessResult { succeeded, @@ -60,89 +140,198 @@ impl DerivedEndednessResult { last, both, neither, + rpt, + read_groups, } } } -fn calculate_reads_per_template( - read_names: Trie>>, -) -> HashMap, f64> { +fn calculate_reads_per_template(read_names: Trie>>) -> HashMap, f64> { + let mut reads_per_template: HashMap, f64> = HashMap::new(); let mut total_reads: usize = 0; - let mut templates_templates: usize = 0; - let mut reads_per_template: HashMap, f64> = HashMap::new(); + let mut total_templates: usize = 0; + let mut read_group_reads: HashMap, usize> = HashMap::new(); + let mut read_group_templates: HashMap, usize> = HashMap::new(); + + for (read_name, read_groups) in read_names.iter() { + let num_reads = read_groups.len(); + total_reads += num_reads; + total_templates += 1; - for read_groups in iter::Iter::new(read_names) {} + let read_group_set: HashSet> = read_groups.iter().cloned().collect(); + + if read_group_set.len() == 1 { + let read_group = read_group_set.iter().next().unwrap(); + + read_group_reads + .entry(read_group.clone()) + .and_modify(|e| *e += num_reads) + .or_insert(num_reads); + read_group_templates + .entry(read_group.clone()) + .and_modify(|e| *e += 1) + .or_insert(1); + } else { + warn!( + "Read {} has multiple read groups: {:#?}", + read_name, read_groups + ); + for read_group in read_groups { + let read_group = Rc::new(**read_group); + read_group_reads + .entry(read_group) + .and_modify(|e| *e += 1) + .or_insert(1); + } + for read_group in read_group_set { + read_group_templates + .entry(read_group) + .and_modify(|e| *e += 1) + .or_insert(1); + } + } + } + + reads_per_template.insert( + Rc::new(OVERALL.as_str()), + total_reads as f64 / total_templates as f64, + ); + + for (read_group, num_reads) in read_group_reads.iter() { + let num_templates = read_group_templates.get(read_group).unwrap(); + let rpt = *num_reads as f64 / *num_templates as f64; + reads_per_template.insert(Rc::clone(read_group), rpt); + } reads_per_template } -/// Main method to evaluate the collected ordering flags and -/// return a result for the endedness of the file. This may fail, and the -/// resulting [`DerivedEndednessResult`] should be evaluated accordingly. -pub fn predict( - ordering_flags: HashMap, HashMap>, - read_names: Trie>>, +fn predict_endedness( + read_group_name: String, + rg_ordering_flags: &OrderingFlagsCounts, paired_deviance: f64, -) -> Result { - let overall_ordering_flags = ordering_flags.get(&*OVERALL).unwrap(); + reads_per_template: Option<&f64>, + round_rpt: bool, +) -> Result { + let first = rg_ordering_flags.first; + let last = rg_ordering_flags.last; + let both = rg_ordering_flags.both; + let neither = rg_ordering_flags.neither; - let overall_first = *overall_ordering_flags.get(&*FIRST).unwrap(); - let overall_last = *overall_ordering_flags.get(&*LAST).unwrap(); - let overall_both = *overall_ordering_flags.get(&*BOTH).unwrap(); - let overall_neither = *overall_ordering_flags.get(&*NEITHER).unwrap(); - - let mut result = DerivedEndednessResult::new( + let mut result = ReadGroupDerivedEndednessResult::new( + read_group_name, false, "Unknown".to_string(), - overall_first, - overall_last, - overall_both, - overall_neither, + first, + last, + both, + neither, + reads_per_template.copied(), ); // all zeroes - if overall_first == 0 && overall_last == 0 && overall_both == 0 && overall_neither == 0 { + if first == 0 && last == 0 && both == 0 && neither == 0 { bail!("No reads were detected in the file."); } // only first present - if overall_first > 0 && overall_last == 0 && overall_both == 0 && overall_neither == 0 { + if first > 0 && last == 0 && both == 0 && neither == 0 { return Ok(result); } // only last present - if overall_first == 0 && overall_last > 0 && overall_both == 0 && overall_neither == 0 { + if first == 0 && last > 0 && both == 0 && neither == 0 { return Ok(result); } // only both present - if overall_first == 0 && overall_last == 0 && overall_both > 0 && overall_neither == 0 { - result.succeeded = true; - result.endedness = "Single-End".to_string(); + if first == 0 && last == 0 && both > 0 && neither == 0 { + match reads_per_template { + Some(rpt) => { + if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { + result.succeeded = true; + result.endedness = String::from("Single-End"); + } + } + None => { + result.succeeded = true; + result.endedness = String::from("Single-End"); + } + } return Ok(result); } // only neither present - if overall_first == 0 && overall_last == 0 && overall_both == 0 && overall_neither > 0 { + if first == 0 && last == 0 && both == 0 && neither > 0 { return Ok(result); } // first/last mixed with both/neither - if (overall_first > 0 || overall_last > 0) && (overall_both > 0 || overall_neither > 0) { + if (first > 0 || last > 0) && (both > 0 || neither > 0) { return Ok(result); } // any mix of both/neither, regardless of first/last - if overall_both > 0 && overall_neither > 0 { + if both > 0 && neither > 0 { return Ok(result); } // both and neither are now guarenteed to be 0 // We only need to check first and last - let first_frac = overall_first as f64 / (overall_first + overall_last) as f64; + let first_frac = first as f64 / (first + last) as f64; let lower_limit = 0.5 - paired_deviance; let upper_limit = 0.5 + paired_deviance; - if (overall_first == overall_last) || (lower_limit <= first_frac && first_frac <= upper_limit) { - result.succeeded = true; - result.endedness = "Paired-End".to_string(); - return Ok(result); + if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { + match reads_per_template { + Some(rpt) => { + if *rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { + result.succeeded = true; + result.endedness = String::from("Paired-End"); + } + } + None => { + result.succeeded = true; + result.endedness = String::from("Paired-End"); + } + } + } + + return Ok(result); +} + +/// Main method to evaluate the collected ordering flags and +/// return a result for the endedness of the file. This may fail, and the +/// resulting [`DerivedEndednessResult`] should be evaluated accordingly. +pub fn predict( + ordering_flags: HashMap, OrderingFlagsCounts>, + read_names: Trie>>, + paired_deviance: f64, + round_rpt: bool, +) -> Result { + let mut rpts: HashMap, f64> = HashMap::new(); + if !read_names.is_empty() { + rpts = calculate_reads_per_template(read_names); + } + + let mut final_result = + DerivedEndednessResult::new(false, "Unknown".to_string(), 0, 0, 0, 0, None, Vec::new()); + + for (read_group, rg_ordering_flags) in ordering_flags.iter() { + let result = predict_endedness( + String::from(**read_group), + rg_ordering_flags, + paired_deviance, + rpts.get(read_group), + round_rpt, + )?; + if result.read_group == String::from("overall") { + final_result.endedness = result.endedness; + final_result.first = result.first; + final_result.last = result.last; + final_result.both = result.both; + final_result.neither = result.neither; + final_result.rpt = result.rpt; + final_result.succeeded = result.succeeded; + } else { + final_result.read_groups.push(result); + } } - Ok(result) + Ok(final_result) } From 137beeaeb4b7fbc2e1b5b9b54cce166871516644 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 7 Dec 2023 16:23:52 -0500 Subject: [PATCH 08/91] feat: working derive endedness subcommand --- src/derive/command/encoding.rs | 2 +- src/derive/command/endedness.rs | 82 ++++--- src/derive/endedness/compute.rs | 378 ++++++++++++++++++++++++++++---- 3 files changed, 388 insertions(+), 74 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index c5692f5..b0b03dd 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -18,6 +18,6 @@ pub struct DeriveEncodingArgs { } /// Main function for the `ngs derive encoding` subcommand. -pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { +pub fn derive(_args: DeriveEncodingArgs) -> anyhow::Result<()> { Ok(()) } diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index a7bdf28..f1180a6 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -62,13 +62,16 @@ pub struct DeriveEndednessArgs { pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Rc::new(&OVERALL), OrderingFlagsCounts::new()); - ordering_flags.insert(Rc::new(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Rc::new(OVERALL.to_string()), OrderingFlagsCounts::new()); + ordering_flags.insert( + Rc::new(UNKNOWN_READ_GROUP.to_string()), + OrderingFlagsCounts::new(), + ); // only used if args.calc_rpt is true let mut found_rgs = HashSet::new(); - let mut read_names = Trie::>>::new(); + let mut read_names = Trie::>>::new(); let ParsedBAMFile { mut reader, header, .. @@ -94,8 +97,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => Rc::new(rg.as_str().unwrap()), - None => Rc::new(UNKNOWN_READ_GROUP.as_str()), + Some(rg) => Rc::new(rg.as_str().unwrap().to_owned()), + None => Rc::new(UNKNOWN_READ_GROUP.to_string()), }; if args.calc_rpt { @@ -110,8 +113,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { rg_vec.push(Rc::clone(&read_group)); } None => { - let mut rg_vec = Vec::new(); - rg_vec.push(Rc::clone(&read_group)); + let rg_vec = vec![(Rc::clone(&read_group))]; read_names.insert(rn.to_string(), rg_vec); } } @@ -125,53 +127,77 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { - e.first += 1; - }); + ordering_flags + .entry(Rc::new(OVERALL.to_string())) + .and_modify(|e| { + e.first += 1; + }); ordering_flags .entry(read_group) .and_modify(|e| { e.first += 1; }) - .or_insert(OrderingFlagsCounts::new()) - .first += 1; + .or_insert(OrderingFlagsCounts { + first: 1, + last: 0, + both: 0, + neither: 0, + }); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { - e.last += 1; - }); + ordering_flags + .entry(Rc::new(OVERALL.to_string())) + .and_modify(|e| { + e.last += 1; + }); ordering_flags .entry(read_group) .and_modify(|e| { e.last += 1; }) - .or_insert(OrderingFlagsCounts::new()) - .last += 1; + .or_insert(OrderingFlagsCounts { + first: 0, + last: 1, + both: 0, + neither: 0, + }); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { - e.both += 1; - }); + ordering_flags + .entry(Rc::new(OVERALL.to_string())) + .and_modify(|e| { + e.both += 1; + }); ordering_flags .entry(read_group) .and_modify(|e| { e.both += 1; }) - .or_insert(OrderingFlagsCounts::new()) - .both += 1; + .or_insert(OrderingFlagsCounts { + first: 0, + last: 0, + both: 1, + neither: 0, + }); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(Rc::new(&OVERALL)).and_modify(|e| { - e.neither += 1; - }); + ordering_flags + .entry(Rc::new(OVERALL.to_string())) + .and_modify(|e| { + e.neither += 1; + }); ordering_flags .entry(read_group) .and_modify(|e| { e.neither += 1; }) - .or_insert(OrderingFlagsCounts::new()) - .neither += 1; + .or_insert(OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 1, + }); } else { unreachable!(); } diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 670e278..30c31fb 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -10,20 +10,33 @@ use std::collections::HashSet; use std::rc::Rc; use tracing::warn; +// Strings used to index into the HashMaps used to store the Read Group ordering flags. +// Lazy statics are used to save memory. lazy_static! { - // Strings used to index into the HashMaps used to store the Read Group ordering flags. + /// String used to index into the HashMaps used to store the "overall" ordering flags. pub static ref OVERALL: String = String::from("overall"); + + /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. pub static ref UNKNOWN_READ_GROUP: String = String::from("unknown_read_group"); } +/// Struct holding the ordering flags for a single read group. +#[derive(Debug, Clone)] pub struct OrderingFlagsCounts { + /// The number of reads with the first in template flag set. pub first: usize, + + /// The number of reads with the last in template flag set. pub last: usize, + + /// The number of reads with both the first and last in template flags set. pub both: usize, + + /// The number of reads with neither the first nor last in template flags set. pub neither: usize, } - impl OrderingFlagsCounts { + /// Creates a new [`OrderingFlagsCounts`]. pub fn new() -> Self { OrderingFlagsCounts { first: 0, @@ -34,10 +47,16 @@ impl OrderingFlagsCounts { } } +impl Default for OrderingFlagsCounts { + fn default() -> Self { + Self::new() + } +} + /// Struct holding the per read group results for an `ngs derive endedness` /// subcommand call. #[derive(Debug, Serialize)] -struct ReadGroupDerivedEndednessResult { +pub struct ReadGroupDerivedEndednessResult { /// Name of the read group. pub read_group: String, @@ -70,20 +89,17 @@ impl ReadGroupDerivedEndednessResult { read_group: String, succeeded: bool, endedness: String, - first: usize, - last: usize, - both: usize, - neither: usize, + counts: OrderingFlagsCounts, rpt: Option, ) -> Self { ReadGroupDerivedEndednessResult { read_group, succeeded, endedness, - first, - last, - both, - neither, + first: counts.first, + last: counts.last, + both: counts.both, + neither: counts.neither, rpt, } } @@ -126,39 +142,38 @@ impl DerivedEndednessResult { pub fn new( succeeded: bool, endedness: String, - first: usize, - last: usize, - both: usize, - neither: usize, + counts: OrderingFlagsCounts, rpt: Option, read_groups: Vec, ) -> Self { DerivedEndednessResult { succeeded, endedness, - first, - last, - both, - neither, + first: counts.first, + last: counts.last, + both: counts.both, + neither: counts.neither, rpt, read_groups, } } } -fn calculate_reads_per_template(read_names: Trie>>) -> HashMap, f64> { - let mut reads_per_template: HashMap, f64> = HashMap::new(); +fn calculate_reads_per_template( + read_names: Trie>>, +) -> HashMap, f64> { + let mut reads_per_template: HashMap, f64> = HashMap::new(); let mut total_reads: usize = 0; let mut total_templates: usize = 0; - let mut read_group_reads: HashMap, usize> = HashMap::new(); - let mut read_group_templates: HashMap, usize> = HashMap::new(); + let mut read_group_reads: HashMap, usize> = HashMap::new(); + let mut read_group_templates: HashMap, usize> = HashMap::new(); for (read_name, read_groups) in read_names.iter() { let num_reads = read_groups.len(); total_reads += num_reads; total_templates += 1; - let read_group_set: HashSet> = read_groups.iter().cloned().collect(); + let read_group_set: HashSet> = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { let read_group = read_group_set.iter().next().unwrap(); @@ -177,9 +192,8 @@ fn calculate_reads_per_template(read_names: Trie>>) -> Hash read_name, read_groups ); for read_group in read_groups { - let read_group = Rc::new(**read_group); read_group_reads - .entry(read_group) + .entry(read_group.clone()) .and_modify(|e| *e += 1) .or_insert(1); } @@ -193,7 +207,7 @@ fn calculate_reads_per_template(read_names: Trie>>) -> Hash } reads_per_template.insert( - Rc::new(OVERALL.as_str()), + Rc::new(OVERALL.to_string()), total_reads as f64 / total_templates as f64, ); @@ -218,22 +232,23 @@ fn predict_endedness( let both = rg_ordering_flags.both; let neither = rg_ordering_flags.neither; + // all zeroes (Perform this check before creating the result struct + // so that we don't have to clone the read group name) + if first == 0 && last == 0 && both == 0 && neither == 0 { + bail!( + "No reads were detected in this read group: {}", + read_group_name + ); + } + let mut result = ReadGroupDerivedEndednessResult::new( read_group_name, false, "Unknown".to_string(), - first, - last, - both, - neither, + rg_ordering_flags.clone(), reads_per_template.copied(), ); - // all zeroes - if first == 0 && last == 0 && both == 0 && neither == 0 { - bail!("No reads were detected in the file."); - } - // only first present if first > 0 && last == 0 && both == 0 && neither == 0 { return Ok(result); @@ -291,36 +306,48 @@ fn predict_endedness( } } } - - return Ok(result); + Ok(result) } /// Main method to evaluate the collected ordering flags and /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: HashMap, OrderingFlagsCounts>, - read_names: Trie>>, + ordering_flags: HashMap, OrderingFlagsCounts>, + read_names: Trie>>, paired_deviance: f64, round_rpt: bool, ) -> Result { - let mut rpts: HashMap, f64> = HashMap::new(); + let mut rpts: HashMap, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); } - let mut final_result = - DerivedEndednessResult::new(false, "Unknown".to_string(), 0, 0, 0, 0, None, Vec::new()); + let mut final_result = DerivedEndednessResult::new( + false, + "Unknown".to_string(), + OrderingFlagsCounts::new(), + None, + Vec::new(), + ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { + if (read_group == &Rc::new(UNKNOWN_READ_GROUP.to_string())) + && (rg_ordering_flags.first == 0 + && rg_ordering_flags.last == 0 + && rg_ordering_flags.both == 0 + && rg_ordering_flags.neither == 0) + { + continue; + } let result = predict_endedness( - String::from(**read_group), + read_group.to_string(), rg_ordering_flags, paired_deviance, rpts.get(read_group), round_rpt, )?; - if result.read_group == String::from("overall") { + if result.read_group == "overall" { final_result.endedness = result.endedness; final_result.first = result.first; final_result.last = result.last; @@ -335,3 +362,264 @@ pub fn predict( Ok(final_result) } + +#[cfg(test)] +mod tests { + use super::*; + + #[test] + fn test_derive_endedness_from_all_zero_counts() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Rc::new(OVERALL.to_string()), OrderingFlagsCounts::new()); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_err()); + } + + #[test] + fn test_derive_endedness_from_only_first() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 1, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 0, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_both() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 1, + neither: 0, + }, + ); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 1); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_neither() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 1, + }, + ); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 1); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_first_and_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, Trie::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_calculate_reads_per_template() { + let mut read_names: Trie>> = Trie::new(); + read_names.insert( + "read1".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + ], + ); + read_names.insert( + "read2".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + Rc::new("rg_single".to_string()), + ], + ); + read_names.insert("read3".to_string(), vec![Rc::new("rg_single".to_string())]); + read_names.insert( + "read4".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + ], + ); + read_names.insert( + "read5".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + Rc::new("rg_single".to_string()), + ], + ); + let results = calculate_reads_per_template(read_names); + assert_eq!(results.len(), 3); + assert_eq!(results.get(&Rc::new("overall".to_string())).unwrap(), &2.2); + assert_eq!( + results.get(&Rc::new("rg_paired".to_string())).unwrap(), + &2.0 + ); + assert_eq!( + results.get(&Rc::new("rg_single".to_string())).unwrap(), + &1.0 + ); + } + + #[test] + fn test_derive_endedness_from_first_and_last_with_good_rpt() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Rc::new(OVERALL.to_string()), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 2, + neither: 0, + }, + ); + ordering_flags.insert( + Rc::new("rg_paired".to_string()), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 0, + neither: 0, + }, + ); + ordering_flags.insert( + Rc::new("rg_single".to_string()), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 2, + neither: 0, + }, + ); + let mut read_names: Trie>> = Trie::new(); + read_names.insert( + "read1".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + ], + ); + read_names.insert( + "read2".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + Rc::new("rg_single".to_string()), + ], + ); + read_names.insert("read3".to_string(), vec![Rc::new("rg_single".to_string())]); + read_names.insert( + "read4".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + ], + ); + read_names.insert( + "read5".to_string(), + vec![ + Rc::new("rg_paired".to_string()), + Rc::new("rg_paired".to_string()), + Rc::new("rg_single".to_string()), + ], + ); + let result = predict(ordering_flags, read_names, 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 8); + assert_eq!(result.last, 8); + assert_eq!(result.both, 2); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, Some(2.2)); + assert_eq!(result.read_groups.len(), 2); + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); + } +} From d1cac4d320b8165b72fbc9cadba3386446dafc3d Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 8 Dec 2023 08:47:59 -0500 Subject: [PATCH 09/91] chore: better comments/test name/log message --- src/derive/endedness/compute.rs | 8 +++++--- 1 file changed, 5 insertions(+), 3 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 30c31fb..dadac20 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -188,8 +188,8 @@ fn calculate_reads_per_template( .or_insert(1); } else { warn!( - "Read {} has multiple read groups: {:#?}", - read_name, read_groups + "QNAME: '{}' is in multiple read groups: {:?}", + read_name, read_group_set ); for read_group in read_groups { read_group_reads @@ -548,7 +548,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_first_and_last_with_good_rpt() { + fn test_derive_endedness_from_first_and_last_with_rpt() { let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( Rc::new(OVERALL.to_string()), @@ -620,6 +620,8 @@ mod tests { assert_eq!(result.neither, 0); assert_eq!(result.rpt, Some(2.2)); assert_eq!(result.read_groups.len(), 2); + // We can't know which read group will be first in the vector. + // But both should succeed. assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); } } From d92368fea2ca867679f5bdb345a0273f0978f7f1 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 9 Dec 2023 09:37:12 -0500 Subject: [PATCH 10/91] fix: apply some of Clay's performance suggestions --- src/derive/command/endedness.rs | 70 +++++++++---------- src/derive/endedness/compute.rs | 120 ++++++++++++++++---------------- 2 files changed, 93 insertions(+), 97 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index f1180a6..3abe3ec 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -3,7 +3,7 @@ use std::collections::HashMap; use std::collections::HashSet; use std::path::PathBuf; -use std::rc::Rc; +use std::sync::Arc; use clap::Args; use noodles::sam::record::data::field::Tag; @@ -62,16 +62,14 @@ pub struct DeriveEndednessArgs { pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { info!("Starting derive endedness subcommand."); - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Rc::new(OVERALL.to_string()), OrderingFlagsCounts::new()); - ordering_flags.insert( - Rc::new(UNKNOWN_READ_GROUP.to_string()), - OrderingFlagsCounts::new(), - ); + let mut found_rgs = HashSet::new(); + + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); + ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true - let mut found_rgs = HashSet::new(); - let mut read_names = Trie::>>::new(); + let mut read_names = Trie::>>::new(); let ParsedBAMFile { mut reader, header, .. @@ -97,24 +95,28 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => Rc::new(rg.as_str().unwrap().to_owned()), - None => Rc::new(UNKNOWN_READ_GROUP.to_string()), + Some(rg) => { + let rg = rg.to_string(); + if !found_rgs.contains(&rg) { + found_rgs.insert(Arc::new(rg.clone())); + } + found_rgs.get(&rg).unwrap().clone() + } + None => Arc::clone(&UNKNOWN_READ_GROUP), }; if args.calc_rpt { - found_rgs.insert(Rc::clone(&read_group)); - match record.read_name() { Some(rn) => { - let rg_vec = read_names.get_mut(&rn.to_string()); + let rn = rn.to_string(); + let rg_vec = read_names.get_mut(&rn); match rg_vec { Some(rg_vec) => { - rg_vec.push(Rc::clone(&read_group)); + rg_vec.push(Arc::clone(&read_group)); } None => { - let rg_vec = vec![(Rc::clone(&read_group))]; - read_names.insert(rn.to_string(), rg_vec); + read_names.insert(rn, vec![(Arc::clone(&read_group))]); } } } @@ -126,12 +128,12 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } + let overall_rg = Arc::clone(&OVERALL); + if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags - .entry(Rc::new(OVERALL.to_string())) - .and_modify(|e| { - e.first += 1; - }); + ordering_flags.entry(overall_rg).and_modify(|e| { + e.first += 1; + }); ordering_flags .entry(read_group) @@ -145,11 +147,9 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags - .entry(Rc::new(OVERALL.to_string())) - .and_modify(|e| { - e.last += 1; - }); + ordering_flags.entry(overall_rg).and_modify(|e| { + e.last += 1; + }); ordering_flags .entry(read_group) @@ -163,11 +163,9 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags - .entry(Rc::new(OVERALL.to_string())) - .and_modify(|e| { - e.both += 1; - }); + ordering_flags.entry(overall_rg).and_modify(|e| { + e.both += 1; + }); ordering_flags .entry(read_group) @@ -181,11 +179,9 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags - .entry(Rc::new(OVERALL.to_string())) - .and_modify(|e| { - e.neither += 1; - }); + ordering_flags.entry(overall_rg).and_modify(|e| { + e.neither += 1; + }); ordering_flags .entry(read_group) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index dadac20..d897ab2 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -7,17 +7,17 @@ use radix_trie::TrieCommon; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; -use std::rc::Rc; +use std::sync::Arc; use tracing::warn; // Strings used to index into the HashMaps used to store the Read Group ordering flags. // Lazy statics are used to save memory. lazy_static! { /// String used to index into the HashMaps used to store the "overall" ordering flags. - pub static ref OVERALL: String = String::from("overall"); + pub static ref OVERALL: Arc = Arc::new(String::from("overall")); /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. - pub static ref UNKNOWN_READ_GROUP: String = String::from("unknown_read_group"); + pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); } /// Struct holding the ordering flags for a single read group. @@ -160,30 +160,30 @@ impl DerivedEndednessResult { } fn calculate_reads_per_template( - read_names: Trie>>, -) -> HashMap, f64> { - let mut reads_per_template: HashMap, f64> = HashMap::new(); + read_names: Trie>>, +) -> HashMap, f64> { + let mut reads_per_template: HashMap, f64> = HashMap::new(); let mut total_reads: usize = 0; let mut total_templates: usize = 0; - let mut read_group_reads: HashMap, usize> = HashMap::new(); - let mut read_group_templates: HashMap, usize> = HashMap::new(); + let mut read_group_reads: HashMap, usize> = HashMap::new(); + let mut read_group_templates: HashMap, usize> = HashMap::new(); for (read_name, read_groups) in read_names.iter() { let num_reads = read_groups.len(); total_reads += num_reads; total_templates += 1; - let read_group_set: HashSet> = read_groups.iter().cloned().collect(); + let read_group_set: HashSet> = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { - let read_group = read_group_set.iter().next().unwrap(); + let read_group = read_group_set.iter().next().unwrap().clone(); read_group_reads .entry(read_group.clone()) .and_modify(|e| *e += num_reads) .or_insert(num_reads); read_group_templates - .entry(read_group.clone()) + .entry(read_group) .and_modify(|e| *e += 1) .or_insert(1); } else { @@ -207,14 +207,14 @@ fn calculate_reads_per_template( } reads_per_template.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), total_reads as f64 / total_templates as f64, ); for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); let rpt = *num_reads as f64 / *num_templates as f64; - reads_per_template.insert(Rc::clone(read_group), rpt); + reads_per_template.insert(Arc::clone(read_group), rpt); } reads_per_template @@ -313,12 +313,12 @@ fn predict_endedness( /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: HashMap, OrderingFlagsCounts>, - read_names: Trie>>, + ordering_flags: HashMap, OrderingFlagsCounts>, + read_names: Trie>>, paired_deviance: f64, round_rpt: bool, ) -> Result { - let mut rpts: HashMap, f64> = HashMap::new(); + let mut rpts: HashMap, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); } @@ -332,7 +332,7 @@ pub fn predict( ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - if (read_group == &Rc::new(UNKNOWN_READ_GROUP.to_string())) + if (*read_group == *UNKNOWN_READ_GROUP) && (rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 @@ -369,17 +369,17 @@ mod tests { #[test] fn test_derive_endedness_from_all_zero_counts() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Rc::new(OVERALL.to_string()), OrderingFlagsCounts::new()); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); let result = predict(ordering_flags, Trie::new(), 0.0, false); assert!(result.is_err()); } #[test] fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 1, last: 0, @@ -402,9 +402,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 1, @@ -427,9 +427,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 0, @@ -452,9 +452,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 0, @@ -477,9 +477,9 @@ mod tests { #[test] fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 1, last: 1, @@ -502,56 +502,56 @@ mod tests { #[test] fn test_calculate_reads_per_template() { - let mut read_names: Trie>> = Trie::new(); + let mut read_names: Trie>> = Trie::new(); read_names.insert( "read1".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), ], ); read_names.insert( "read2".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), - Rc::new("rg_single".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_single".to_string()), ], ); - read_names.insert("read3".to_string(), vec![Rc::new("rg_single".to_string())]); + read_names.insert("read3".to_string(), vec![Arc::new("rg_single".to_string())]); read_names.insert( "read4".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), ], ); read_names.insert( "read5".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), - Rc::new("rg_single".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_single".to_string()), ], ); let results = calculate_reads_per_template(read_names); assert_eq!(results.len(), 3); - assert_eq!(results.get(&Rc::new("overall".to_string())).unwrap(), &2.2); + assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); assert_eq!( - results.get(&Rc::new("rg_paired".to_string())).unwrap(), + results.get(&Arc::new("rg_paired".to_string())).unwrap(), &2.0 ); assert_eq!( - results.get(&Rc::new("rg_single".to_string())).unwrap(), + results.get(&Arc::new("rg_single".to_string())).unwrap(), &1.0 ); } #[test] fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - Rc::new(OVERALL.to_string()), + Arc::new(OVERALL.to_string()), OrderingFlagsCounts { first: 8, last: 8, @@ -560,7 +560,7 @@ mod tests { }, ); ordering_flags.insert( - Rc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), OrderingFlagsCounts { first: 8, last: 8, @@ -569,7 +569,7 @@ mod tests { }, ); ordering_flags.insert( - Rc::new("rg_single".to_string()), + Arc::new("rg_single".to_string()), OrderingFlagsCounts { first: 0, last: 0, @@ -577,36 +577,36 @@ mod tests { neither: 0, }, ); - let mut read_names: Trie>> = Trie::new(); + let mut read_names: Trie>> = Trie::new(); read_names.insert( "read1".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), ], ); read_names.insert( "read2".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), - Rc::new("rg_single".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_single".to_string()), ], ); - read_names.insert("read3".to_string(), vec![Rc::new("rg_single".to_string())]); + read_names.insert("read3".to_string(), vec![Arc::new("rg_single".to_string())]); read_names.insert( "read4".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), ], ); read_names.insert( "read5".to_string(), vec![ - Rc::new("rg_paired".to_string()), - Rc::new("rg_paired".to_string()), - Rc::new("rg_single".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_paired".to_string()), + Arc::new("rg_single".to_string()), ], ); let result = predict(ordering_flags, read_names, 0.0, false); From 7e6ad22bba1333da520ecea58881d0ca7ffad766 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 9 Dec 2023 10:02:44 -0500 Subject: [PATCH 11/91] fix(derive/endedness/compute.rs): test updates --- src/derive/endedness/compute.rs | 66 +++++++++------------------------ 1 file changed, 18 insertions(+), 48 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index d897ab2..23bfa0b 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -503,53 +503,37 @@ mod tests { #[test] fn test_calculate_reads_per_template() { let mut read_names: Trie>> = Trie::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); read_names.insert( "read1".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone()], ); read_names.insert( "read2".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - Arc::new("rg_single".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], ); - read_names.insert("read3".to_string(), vec![Arc::new("rg_single".to_string())]); + read_names.insert("read3".to_string(), vec![rg_single.clone()]); read_names.insert( "read4".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone()], ); read_names.insert( "read5".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - Arc::new("rg_single".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], ); let results = calculate_reads_per_template(read_names); assert_eq!(results.len(), 3); assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); - assert_eq!( - results.get(&Arc::new("rg_paired".to_string())).unwrap(), - &2.0 - ); - assert_eq!( - results.get(&Arc::new("rg_single".to_string())).unwrap(), - &1.0 - ); + assert_eq!(results.get(&rg_paired.clone()).unwrap(), &2.0); + assert_eq!(results.get(&rg_single.clone()).unwrap(), &1.0); } #[test] fn test_derive_endedness_from_first_and_last_with_rpt() { let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); ordering_flags.insert( Arc::new(OVERALL.to_string()), OrderingFlagsCounts { @@ -560,7 +544,7 @@ mod tests { }, ); ordering_flags.insert( - Arc::new("rg_paired".to_string()), + rg_paired.clone(), OrderingFlagsCounts { first: 8, last: 8, @@ -569,7 +553,7 @@ mod tests { }, ); ordering_flags.insert( - Arc::new("rg_single".to_string()), + rg_single.clone(), OrderingFlagsCounts { first: 0, last: 0, @@ -580,34 +564,20 @@ mod tests { let mut read_names: Trie>> = Trie::new(); read_names.insert( "read1".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone()], ); read_names.insert( "read2".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - Arc::new("rg_single".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], ); - read_names.insert("read3".to_string(), vec![Arc::new("rg_single".to_string())]); + read_names.insert("read3".to_string(), vec![rg_single.clone()]); read_names.insert( "read4".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone()], ); read_names.insert( "read5".to_string(), - vec![ - Arc::new("rg_paired".to_string()), - Arc::new("rg_paired".to_string()), - Arc::new("rg_single".to_string()), - ], + vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], ); let result = predict(ordering_flags, read_names, 0.0, false); assert!(result.is_ok()); From 34bf08d14d94e042938381cb68542d900eecf302 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 9 Dec 2023 12:23:44 -0500 Subject: [PATCH 12/91] chore(derive/readlen/compute): cleanup --- src/derive/readlen/compute.rs | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 27a524f..71bd504 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -75,7 +75,6 @@ pub fn predict( if majority_detected >= majority_vote_cutoff { result.succeeded = true; result.consensus_read_length = Some(consensus_read_length); - result.majority_pct_detected = majority_detected * 100.0; } Ok(result) @@ -118,7 +117,7 @@ mod tests { let result = predict(read_lengths, 0.7).unwrap(); assert!(!result.succeeded); assert_eq!(result.consensus_read_length, None); - assert!(result.majority_pct_detected < 1.0); + assert!(result.majority_pct_detected < 0.7); assert_eq!(result.evidence, Vec::from([(101, 5), (100, 1000), (99, 5)])); } } From 43aaec340edce7d2b7b23041ea748b929c94757f Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 10 Dec 2023 09:36:29 -0500 Subject: [PATCH 13/91] fix(derive/command/endedness): try using HashMap instead of Trie --- src/derive/command/endedness.rs | 3 +- src/derive/endedness/compute.rs | 470 ++++++++++++++++---------------- 2 files changed, 235 insertions(+), 238 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 3abe3ec..c111ebf 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -7,7 +7,6 @@ use std::sync::Arc; use clap::Args; use noodles::sam::record::data::field::Tag; -use radix_trie::Trie; use tracing::info; use tracing::trace; @@ -69,7 +68,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true - let mut read_names = Trie::>>::new(); + let mut read_names: HashMap>> = HashMap::new(); let ParsedBAMFile { mut reader, header, .. diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 23bfa0b..4f934ed 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -2,8 +2,6 @@ use anyhow::bail; use lazy_static::lazy_static; -use radix_trie::Trie; -use radix_trie::TrieCommon; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; @@ -160,7 +158,7 @@ impl DerivedEndednessResult { } fn calculate_reads_per_template( - read_names: Trie>>, + read_names: HashMap>>, ) -> HashMap, f64> { let mut reads_per_template: HashMap, f64> = HashMap::new(); let mut total_reads: usize = 0; @@ -314,7 +312,7 @@ fn predict_endedness( /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( ordering_flags: HashMap, OrderingFlagsCounts>, - read_names: Trie>>, + read_names: HashMap>>, paired_deviance: f64, round_rpt: bool, ) -> Result { @@ -363,235 +361,235 @@ pub fn predict( Ok(final_result) } -#[cfg(test)] -mod tests { - use super::*; - - #[test] - fn test_derive_endedness_from_all_zero_counts() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_err()); - } - - #[test] - fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 0, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 1, - neither: 0, - }, - ); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Single-End"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 1); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 0, - neither: 1, - }, - ); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 1); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, Trie::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_calculate_reads_per_template() { - let mut read_names: Trie>> = Trie::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - read_names.insert( - "read1".to_string(), - vec![rg_paired.clone(), rg_paired.clone()], - ); - read_names.insert( - "read2".to_string(), - vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], - ); - read_names.insert("read3".to_string(), vec![rg_single.clone()]); - read_names.insert( - "read4".to_string(), - vec![rg_paired.clone(), rg_paired.clone()], - ); - read_names.insert( - "read5".to_string(), - vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], - ); - let results = calculate_reads_per_template(read_names); - assert_eq!(results.len(), 3); - assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); - assert_eq!(results.get(&rg_paired.clone()).unwrap(), &2.0); - assert_eq!(results.get(&rg_single.clone()).unwrap(), &1.0); - } - - #[test] - fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - ordering_flags.insert( - Arc::new(OVERALL.to_string()), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 2, - neither: 0, - }, - ); - ordering_flags.insert( - rg_paired.clone(), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 0, - neither: 0, - }, - ); - ordering_flags.insert( - rg_single.clone(), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 2, - neither: 0, - }, - ); - let mut read_names: Trie>> = Trie::new(); - read_names.insert( - "read1".to_string(), - vec![rg_paired.clone(), rg_paired.clone()], - ); - read_names.insert( - "read2".to_string(), - vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], - ); - read_names.insert("read3".to_string(), vec![rg_single.clone()]); - read_names.insert( - "read4".to_string(), - vec![rg_paired.clone(), rg_paired.clone()], - ); - read_names.insert( - "read5".to_string(), - vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], - ); - let result = predict(ordering_flags, read_names, 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 8); - assert_eq!(result.last, 8); - assert_eq!(result.both, 2); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, Some(2.2)); - assert_eq!(result.read_groups.len(), 2); - // We can't know which read group will be first in the vector. - // But both should succeed. - assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); - } -} +// #[cfg(test)] +// mod tests { +// use super::*; + +// #[test] +// fn test_derive_endedness_from_all_zero_counts() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_err()); +// } + +// #[test] +// fn test_derive_endedness_from_only_first() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 0, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } +// +// #[test] +// fn test_derive_endedness_from_only_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_both() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 1, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Single-End"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 1); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_neither() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 0, +// neither: 1, +// }, +// ); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 1); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, Trie::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_calculate_reads_per_template() { +// let mut read_names: Trie>> = Trie::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// read_names.insert( +// "read1".to_string(), +// vec![rg_paired.clone(), rg_paired.clone()], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], +// ); +// read_names.insert("read3".to_string(), vec![rg_single.clone()]); +// read_names.insert( +// "read4".to_string(), +// vec![rg_paired.clone(), rg_paired.clone()], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], +// ); +// let results = calculate_reads_per_template(read_names); +// assert_eq!(results.len(), 3); +// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); +// assert_eq!(results.get(&rg_paired.clone()).unwrap(), &2.0); +// assert_eq!(results.get(&rg_single.clone()).unwrap(), &1.0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last_with_rpt() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// ordering_flags.insert( +// Arc::new(OVERALL.to_string()), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 2, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// rg_paired.clone(), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 0, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// rg_single.clone(), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 2, +// neither: 0, +// }, +// ); +// let mut read_names: Trie>> = Trie::new(); +// read_names.insert( +// "read1".to_string(), +// vec![rg_paired.clone(), rg_paired.clone()], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], +// ); +// read_names.insert("read3".to_string(), vec![rg_single.clone()]); +// read_names.insert( +// "read4".to_string(), +// vec![rg_paired.clone(), rg_paired.clone()], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], +// ); +// let result = predict(ordering_flags, read_names, 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 8); +// assert_eq!(result.last, 8); +// assert_eq!(result.both, 2); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, Some(2.2)); +// assert_eq!(result.read_groups.len(), 2); +// // We can't know which read group will be first in the vector. +// // But both should succeed. +// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); +// } +// } From 553fd68827cc027b8cf3898a62a10096308b3ac4 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 11 Dec 2023 12:06:19 -0500 Subject: [PATCH 14/91] feat(derive/command/endedness): lazy record reading --- Cargo.lock | 1247 +++++++++++++++++++------------ Cargo.toml | 3 +- src/derive/command/endedness.rs | 42 +- 3 files changed, 800 insertions(+), 492 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index 925f544..ddcd1b0 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -2,6 +2,15 @@ # It is not intended for manual editing. version = 3 +[[package]] +name = "addr2line" +version = "0.21.0" +source = 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b/src/derive/command/endedness.rs index c111ebf..a426d6f 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -6,7 +6,7 @@ use std::path::PathBuf; use std::sync::Arc; use clap::Args; -use noodles::sam::record::data::field::Tag; +use noodles::bam::lazy::record::data::field::value::Value; use tracing::info; use tracing::trace; @@ -83,41 +83,33 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { sample_max = s; } - for result in reader.records(&header.parsed) { - let record = result?; + let mut record = noodles::bam::lazy::Record::default(); + while reader.read_lazy_record(&mut record)? != 0 { + let flags = record.flags(); // Only count primary alignments and unmapped reads. - if (record.flags().is_secondary() || record.flags().is_supplementary()) - && !record.flags().is_unmapped() - { + if (flags.is_secondary() || flags.is_supplementary()) && !flags.is_unmapped() { continue; } - let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => { - let rg = rg.to_string(); + let read_group = match record.data().get(b"RG") { + Some(Ok(Value::String(rg))) => { + // RG tag found, and can be converted to String + let rg = String::from_utf8_lossy(rg).to_string(); if !found_rgs.contains(&rg) { found_rgs.insert(Arc::new(rg.clone())); } found_rgs.get(&rg).unwrap().clone() } - None => Arc::clone(&UNKNOWN_READ_GROUP), + _ => Arc::clone(&UNKNOWN_READ_GROUP), // RG tag not found or not a String }; if args.calc_rpt { match record.read_name() { Some(rn) => { - let rn = rn.to_string(); - let rg_vec = read_names.get_mut(&rn); - - match rg_vec { - Some(rg_vec) => { - rg_vec.push(Arc::clone(&read_group)); - } - None => { - read_names.insert(rn, vec![(Arc::clone(&read_group))]); - } - } + let rn = String::from_utf8_lossy(rn.as_bytes()).to_string(); + let rg_vec = read_names.entry(rn).or_insert(vec![]); + rg_vec.push(Arc::clone(&read_group)); } None => { trace!("Could not parse a QNAME from a read in the file."); @@ -129,7 +121,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let overall_rg = Arc::clone(&OVERALL); - if record.flags().is_first_segment() && !record.flags().is_last_segment() { + if flags.is_first_segment() && !flags.is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.first += 1; }); @@ -145,7 +137,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 0, neither: 0, }); - } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { + } else if !flags.is_first_segment() && flags.is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.last += 1; }); @@ -161,7 +153,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 0, neither: 0, }); - } else if record.flags().is_first_segment() && record.flags().is_last_segment() { + } else if flags.is_first_segment() && flags.is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.both += 1; }); @@ -177,7 +169,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 1, neither: 0, }); - } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { + } else if !flags.is_first_segment() && !flags.is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.neither += 1; }); From 1b1117cc6ae03a6285b585095de2f4db57f2de81 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 14 Dec 2023 10:06:08 -0500 Subject: [PATCH 15/91] Revert "feat(derive/command/endedness): lazy record reading" This reverts commit 553fd68827cc027b8cf3898a62a10096308b3ac4. --- Cargo.lock | 1247 ++++++++++++------------------- Cargo.toml | 3 +- src/derive/command/endedness.rs | 42 +- 3 files changed, 492 insertions(+), 800 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index ddcd1b0..925f544 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -2,15 +2,6 @@ # It is not intended for manual editing. version = 3 -[[package]] -name = "addr2line" -version = "0.21.0" -source = 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-dependencies = [ - "libm", -] +checksum = "02296996cb8796d7c6e3bc2d9211b7802812d36999a51bb754123ead7d37d026" [[package]] name = "iana-time-zone" -version = "0.1.58" +version = "0.1.50" source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "8326b86b6cff230b97d0d312a6c40a60726df3332e721f72a1b035f451663b20" +checksum = "fd911b35d940d2bd0bea0f9100068e5b97b51a1cbe13d13382f132e0365257a0" dependencies = [ "android_system_properties", "core-foundation-sys", - "iana-time-zone-haiku", "js-sys", "wasm-bindgen", - "windows-core", -] - -[[package]] -name = "iana-time-zone-haiku" -version = "0.1.2" -source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "f31827a206f56af32e590ba56d5d2d085f558508192593743f16b2306495269f" -dependencies = [ - "cc", + "winapi", ] [[package]] @@ -807,25 +656,15 @@ checksum = "b9e0384b61958566e926dc50660321d12159025e767c18e043daf26b70104c39" [[package]] name = "indexmap" -version = "1.9.3" +version = "1.9.1" source = 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2.1.0", + "indexmap", "noodles-bgzf", "noodles-core", "noodles-csi", @@ -1253,13 +1105,14 @@ dependencies = [ [[package]] name = "noodles-vcf" -version = "0.47.0" +version = "0.26.0" source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "610f14797affe3ecc3687732e310ff508c909dd18d3b12d0cd135bc4cc5593e9" +checksum = "fa196d6d6ad22277390e2cc588d6999a00fd73bd1ca4a5f68e8fda710c8e8364" dependencies = [ "futures", - "indexmap 2.1.0", + "indexmap", "memchr", + "nom", "noodles-bgzf", "noodles-core", "noodles-csi", @@ -1269,30 +1122,30 @@ dependencies = [ ] [[package]] -name = "nu-ansi-term" -version = "0.46.0" +name = "num-format" +version = "0.4.0" source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "77a8165726e8236064dbb45459242600304b42a5ea24ee2948e18e023bf7ba84" +checksum = "bafe4179722c2894288ee77a9f044f02811c86af699344c498b0840c698a2465" dependencies = [ - "overload", - "winapi", + "arrayvec", + "itoa 0.4.8", ] [[package]] -name = 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indicatif = "0.16.2" itertools = "0.10.5" lazy_static = "1.4.0" -noodles = { version = "0.59.0", features = [ +noodles = { version = "0.34.0", features = [ "async", "bam", "bgzf", @@ -36,6 +36,7 @@ noodles = { version = "0.59.0", features = [ num-format = "0.4.0" plotly = "0.8.1" prettytable-rs = "0.9.0" +radix_trie = "0.2.1" rand = "0.8.5" rand_distr = "0.4.3" regex = "1.5.5" diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index a426d6f..c111ebf 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -6,7 +6,7 @@ use std::path::PathBuf; use std::sync::Arc; use clap::Args; -use noodles::bam::lazy::record::data::field::value::Value; +use noodles::sam::record::data::field::Tag; use tracing::info; use tracing::trace; @@ -83,33 +83,41 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { sample_max = s; } - let mut record = noodles::bam::lazy::Record::default(); - while reader.read_lazy_record(&mut record)? != 0 { - let flags = record.flags(); + for result in reader.records(&header.parsed) { + let record = result?; // Only count primary alignments and unmapped reads. - if (flags.is_secondary() || flags.is_supplementary()) && !flags.is_unmapped() { + if (record.flags().is_secondary() || record.flags().is_supplementary()) + && !record.flags().is_unmapped() + { continue; } - let read_group = match record.data().get(b"RG") { - Some(Ok(Value::String(rg))) => { - // RG tag found, and can be converted to String - let rg = String::from_utf8_lossy(rg).to_string(); + let read_group = match record.data().get(Tag::ReadGroup) { + Some(rg) => { + let rg = rg.to_string(); if !found_rgs.contains(&rg) { found_rgs.insert(Arc::new(rg.clone())); } found_rgs.get(&rg).unwrap().clone() } - _ => Arc::clone(&UNKNOWN_READ_GROUP), // RG tag not found or not a String + None => Arc::clone(&UNKNOWN_READ_GROUP), }; if args.calc_rpt { match record.read_name() { Some(rn) => { - let rn = String::from_utf8_lossy(rn.as_bytes()).to_string(); - let rg_vec = read_names.entry(rn).or_insert(vec![]); - rg_vec.push(Arc::clone(&read_group)); + let rn = rn.to_string(); + let rg_vec = read_names.get_mut(&rn); + + match rg_vec { + Some(rg_vec) => { + rg_vec.push(Arc::clone(&read_group)); + } + None => { + read_names.insert(rn, vec![(Arc::clone(&read_group))]); + } + } } None => { trace!("Could not parse a QNAME from a read in the file."); @@ -121,7 +129,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let overall_rg = Arc::clone(&OVERALL); - if flags.is_first_segment() && !flags.is_last_segment() { + if record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.first += 1; }); @@ -137,7 +145,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 0, neither: 0, }); - } else if !flags.is_first_segment() && flags.is_last_segment() { + } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.last += 1; }); @@ -153,7 +161,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 0, neither: 0, }); - } else if flags.is_first_segment() && flags.is_last_segment() { + } else if record.flags().is_first_segment() && record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.both += 1; }); @@ -169,7 +177,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { both: 1, neither: 0, }); - } else if !flags.is_first_segment() && !flags.is_last_segment() { + } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.neither += 1; }); From b1e9e8629b601d83fba60a327924ff4c474f08bc Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 14 Dec 2023 11:59:26 -0500 Subject: [PATCH 16/91] tests(derive/endedness/comput): reimplement tests --- src/derive/command/endedness.rs | 2 +- src/derive/endedness/compute.rs | 516 +++++++++++++++++--------------- 2 files changed, 282 insertions(+), 236 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index c111ebf..663a8ee 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -99,7 +99,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { if !found_rgs.contains(&rg) { found_rgs.insert(Arc::new(rg.clone())); } - found_rgs.get(&rg).unwrap().clone() + Arc::clone(found_rgs.get(&rg).unwrap()) } None => Arc::clone(&UNKNOWN_READ_GROUP), }; diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 4f934ed..2d8ef69 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -174,10 +174,10 @@ fn calculate_reads_per_template( let read_group_set: HashSet> = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { - let read_group = read_group_set.iter().next().unwrap().clone(); + let read_group = Arc::clone(read_group_set.iter().next().unwrap()); read_group_reads - .entry(read_group.clone()) + .entry(Arc::clone(&read_group)) .and_modify(|e| *e += num_reads) .or_insert(num_reads); read_group_templates @@ -191,7 +191,7 @@ fn calculate_reads_per_template( ); for read_group in read_groups { read_group_reads - .entry(read_group.clone()) + .entry(Arc::clone(read_group)) .and_modify(|e| *e += 1) .or_insert(1); } @@ -361,235 +361,281 @@ pub fn predict( Ok(final_result) } -// #[cfg(test)] -// mod tests { -// use super::*; - -// #[test] -// fn test_derive_endedness_from_all_zero_counts() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_err()); -// } - -// #[test] -// fn test_derive_endedness_from_only_first() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 0, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } -// -// #[test] -// fn test_derive_endedness_from_only_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_both() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 1, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Single-End"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 1); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_neither() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 0, -// neither: 1, -// }, -// ); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 1); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, Trie::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_calculate_reads_per_template() { -// let mut read_names: Trie>> = Trie::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// read_names.insert( -// "read1".to_string(), -// vec![rg_paired.clone(), rg_paired.clone()], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], -// ); -// read_names.insert("read3".to_string(), vec![rg_single.clone()]); -// read_names.insert( -// "read4".to_string(), -// vec![rg_paired.clone(), rg_paired.clone()], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], -// ); -// let results = calculate_reads_per_template(read_names); -// assert_eq!(results.len(), 3); -// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); -// assert_eq!(results.get(&rg_paired.clone()).unwrap(), &2.0); -// assert_eq!(results.get(&rg_single.clone()).unwrap(), &1.0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last_with_rpt() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// ordering_flags.insert( -// Arc::new(OVERALL.to_string()), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 2, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// rg_paired.clone(), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 0, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// rg_single.clone(), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 2, -// neither: 0, -// }, -// ); -// let mut read_names: Trie>> = Trie::new(); -// read_names.insert( -// "read1".to_string(), -// vec![rg_paired.clone(), rg_paired.clone()], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], -// ); -// read_names.insert("read3".to_string(), vec![rg_single.clone()]); -// read_names.insert( -// "read4".to_string(), -// vec![rg_paired.clone(), rg_paired.clone()], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![rg_paired.clone(), rg_paired.clone(), rg_single.clone()], -// ); -// let result = predict(ordering_flags, read_names, 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 8); -// assert_eq!(result.last, 8); -// assert_eq!(result.both, 2); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, Some(2.2)); -// assert_eq!(result.read_groups.len(), 2); -// // We can't know which read group will be first in the vector. -// // But both should succeed. -// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); -// } -// } +#[cfg(test)] +mod tests { + use super::*; + + #[test] + fn test_predict_endedness() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict_endedness( + "overall".to_string(), + &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), + 0.0, + None, + false, + ); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_all_zero_counts() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_err()); + } + + #[test] + fn test_derive_endedness_from_only_first() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_both() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 1, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 1); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_neither() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 1, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 1); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_first_and_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_calculate_reads_per_template() { + let mut read_names: HashMap>> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let results = calculate_reads_per_template(read_names); + assert_eq!(results.len(), 3); + assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); + assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); + assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); + } + + #[test] + fn test_derive_endedness_from_first_and_last_with_rpt() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 2, + neither: 0, + }, + ); + ordering_flags.insert( + Arc::clone(&rg_paired), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 0, + neither: 0, + }, + ); + ordering_flags.insert( + Arc::clone(&rg_single), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 2, + neither: 0, + }, + ); + let mut read_names: HashMap>> = HashMap::new(); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let result = predict(ordering_flags, read_names, 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 8); + assert_eq!(result.last, 8); + assert_eq!(result.both, 2); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, Some(2.2)); + assert_eq!(result.read_groups.len(), 2); + // We can't know which read group will be first in the vector. + // But both should succeed. + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); + } +} From ae133d67f61121ab351d71558977d7d9823e4e6e Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 14 Dec 2023 15:54:44 -0500 Subject: [PATCH 17/91] [WIP] suggestions from @zaeleus. Compiler error --- src/derive/command/endedness.rs | 34 +- src/derive/command/readlen.rs | 13 +- src/derive/endedness/compute.rs | 622 ++++++++++++++++---------------- src/derive/readlen/compute.rs | 12 +- 4 files changed, 342 insertions(+), 339 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 663a8ee..f7dda29 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -36,8 +36,8 @@ pub struct DeriveEndednessArgs { src: PathBuf, /// Only examine the first n records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + #[arg(short, long, value_name = "U64")] + num_records: Option, /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate @@ -63,16 +63,16 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut found_rgs = HashSet::new(); - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); - ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(OVERALL.as_str(), OrderingFlagsCounts::new()); + ordering_flags.insert(UNKNOWN_READ_GROUP.as_str(), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true - let mut read_names: HashMap>> = HashMap::new(); + let mut read_names: HashMap> = HashMap::new(); let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. @@ -95,13 +95,13 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let read_group = match record.data().get(Tag::ReadGroup) { Some(rg) => { - let rg = rg.to_string(); - if !found_rgs.contains(&rg) { - found_rgs.insert(Arc::new(rg.clone())); + let rg = rg.as_str().unwrap(); + if !found_rgs.contains(rg) { + found_rgs.insert(rg.to_string()); } - Arc::clone(found_rgs.get(&rg).unwrap()) + found_rgs.get(rg).unwrap() } - None => Arc::clone(&UNKNOWN_READ_GROUP), + None => UNKNOWN_READ_GROUP.as_str(), }; if args.calc_rpt { @@ -112,10 +112,10 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { match rg_vec { Some(rg_vec) => { - rg_vec.push(Arc::clone(&read_group)); + rg_vec.push(read_group); } None => { - read_names.insert(rn, vec![(Arc::clone(&read_group))]); + read_names.insert(rn, vec![read_group]); } } } @@ -127,7 +127,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } - let overall_rg = Arc::clone(&OVERALL); + let overall_rg = OVERALL.as_str(); if record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { @@ -207,8 +207,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (2) Derive the consensus endedness based on the ordering flags gathered. let result = compute::predict( - ordering_flags, - read_names, + &ordering_flags, + &read_names, args.paired_deviance.unwrap(), args.round_rpt, ) diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index b04e207..9449b03 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -33,8 +33,8 @@ pub struct DeriveReadlenArgs { src: PathBuf, /// Only examine the first n records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + #[arg(short, long, value_name = "U64")] + num_records: Option, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. #[arg(short, long, value_name = "F64", default_value = "0.7")] @@ -50,7 +50,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. @@ -63,9 +63,12 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { for result in reader.records(&header.parsed) { let record = result?; - let len = record.sequence().len(); + let len = record.sequence().len() as u32; - read_lengths.entry(len).and_modify(|e| *e += 1).or_insert(1); + read_lengths + .entry(len) + .and_modify(|e| *e += 1) + .or_insert(1 as u64); if sample_max > 0 { samples += 1; diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 2d8ef69..86ccad3 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -22,16 +22,16 @@ lazy_static! { #[derive(Debug, Clone)] pub struct OrderingFlagsCounts { /// The number of reads with the first in template flag set. - pub first: usize, + pub first: u64, /// The number of reads with the last in template flag set. - pub last: usize, + pub last: u64, /// The number of reads with both the first and last in template flags set. - pub both: usize, + pub both: u64, /// The number of reads with neither the first nor last in template flags set. - pub neither: usize, + pub neither: u64, } impl OrderingFlagsCounts { /// Creates a new [`OrderingFlagsCounts`]. @@ -65,16 +65,16 @@ pub struct ReadGroupDerivedEndednessResult { pub endedness: String, /// The f+l- read count. - pub first: usize, + pub first: u64, /// The f-l+ read count. - pub last: usize, + pub last: u64, /// The f+l+ read count. - pub both: usize, + pub both: u64, /// The f-l- read count. - pub neither: usize, + pub neither: u64, /// The reads per template (RPT). /// Only available if `args.calc_rpt` is true. @@ -114,16 +114,16 @@ pub struct DerivedEndednessResult { pub endedness: String, /// The overall f+l- read count. - pub first: usize, + pub first: u64, /// The overall f-l+ read count. - pub last: usize, + pub last: u64, /// The overall f+l+ read count. - pub both: usize, + pub both: u64, /// The overall f-l- read count. - pub neither: usize, + pub neither: u64, /// The overall reads per template (RPT). /// Only available if `args.calc_rpt` is true. @@ -157,27 +157,27 @@ impl DerivedEndednessResult { } } -fn calculate_reads_per_template( - read_names: HashMap>>, -) -> HashMap, f64> { - let mut reads_per_template: HashMap, f64> = HashMap::new(); - let mut total_reads: usize = 0; - let mut total_templates: usize = 0; - let mut read_group_reads: HashMap, usize> = HashMap::new(); - let mut read_group_templates: HashMap, usize> = HashMap::new(); +fn calculate_reads_per_template<'rg>( + read_names: &HashMap>, +) -> HashMap<&'rg str, f64> { + let mut reads_per_template: HashMap<&str, f64> = HashMap::new(); + let mut total_reads: u64 = 0; + let mut total_templates: u64 = 0; + let mut read_group_reads: HashMap<&str, u64> = HashMap::new(); + let mut read_group_templates: HashMap<&str, u64> = HashMap::new(); for (read_name, read_groups) in read_names.iter() { - let num_reads = read_groups.len(); + let num_reads = read_groups.len() as u64; total_reads += num_reads; total_templates += 1; - let read_group_set: HashSet> = read_groups.iter().cloned().collect(); + let read_group_set: HashSet<&str> = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { - let read_group = Arc::clone(read_group_set.iter().next().unwrap()); + let read_group = read_group_set.iter().next().unwrap(); read_group_reads - .entry(Arc::clone(&read_group)) + .entry(&read_group) .and_modify(|e| *e += num_reads) .or_insert(num_reads); read_group_templates @@ -191,7 +191,7 @@ fn calculate_reads_per_template( ); for read_group in read_groups { read_group_reads - .entry(Arc::clone(read_group)) + .entry(&read_group) .and_modify(|e| *e += 1) .or_insert(1); } @@ -205,14 +205,14 @@ fn calculate_reads_per_template( } reads_per_template.insert( - Arc::clone(&OVERALL), + OVERALL.as_str(), total_reads as f64 / total_templates as f64, ); for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); let rpt = *num_reads as f64 / *num_templates as f64; - reads_per_template.insert(Arc::clone(read_group), rpt); + reads_per_template.insert(read_group, rpt); } reads_per_template @@ -259,7 +259,7 @@ fn predict_endedness( if first == 0 && last == 0 && both > 0 && neither == 0 { match reads_per_template { Some(rpt) => { - if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { + if *rpt == 1.0 || (round_rpt && rpt.round() as u64 == 1) { result.succeeded = true; result.endedness = String::from("Single-End"); } @@ -293,7 +293,7 @@ fn predict_endedness( if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { match reads_per_template { Some(rpt) => { - if *rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { + if *rpt == 2.0 || (round_rpt && rpt.round() as u64 == 2) { result.succeeded = true; result.endedness = String::from("Paired-End"); } @@ -311,12 +311,12 @@ fn predict_endedness( /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: HashMap, OrderingFlagsCounts>, - read_names: HashMap>>, + ordering_flags: &HashMap<&str, OrderingFlagsCounts>, + read_names: &HashMap>, paired_deviance: f64, round_rpt: bool, ) -> Result { - let mut rpts: HashMap, f64> = HashMap::new(); + let mut rpts: HashMap<&str, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); } @@ -330,7 +330,7 @@ pub fn predict( ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - if (*read_group == *UNKNOWN_READ_GROUP) + if (*read_group == UNKNOWN_READ_GROUP.as_str()) && (rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 @@ -361,281 +361,281 @@ pub fn predict( Ok(final_result) } -#[cfg(test)] -mod tests { - use super::*; - - #[test] - fn test_predict_endedness() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict_endedness( - "overall".to_string(), - &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), - 0.0, - None, - false, - ); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - } - - #[test] - fn test_derive_endedness_from_all_zero_counts() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_err()); - } - - #[test] - fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 0, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 1, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Single-End"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 1); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 0, - neither: 1, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 1); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_calculate_reads_per_template() { - let mut read_names: HashMap>> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let results = calculate_reads_per_template(read_names); - assert_eq!(results.len(), 3); - assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); - assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); - assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); - } - - #[test] - fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 2, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_paired), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 0, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_single), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 2, - neither: 0, - }, - ); - let mut read_names: HashMap>> = HashMap::new(); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let result = predict(ordering_flags, read_names, 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 8); - assert_eq!(result.last, 8); - assert_eq!(result.both, 2); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, Some(2.2)); - assert_eq!(result.read_groups.len(), 2); - // We can't know which read group will be first in the vector. - // But both should succeed. - assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); - } -} +// #[cfg(test)] +// mod tests { +// use super::*; + +// #[test] +// fn test_predict_endedness() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict_endedness( +// "overall".to_string(), +// &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), +// 0.0, +// None, +// false, +// ); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// } + +// #[test] +// fn test_derive_endedness_from_all_zero_counts() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_err()); +// } + +// #[test] +// fn test_derive_endedness_from_only_first() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 0, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_both() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 1, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Single-End"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 1); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_neither() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 0, +// neither: 1, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 1); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_calculate_reads_per_template() { +// let mut read_names: HashMap>> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let results = calculate_reads_per_template(read_names); +// assert_eq!(results.len(), 3); +// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); +// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); +// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last_with_rpt() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 2, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_paired), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 0, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_single), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 2, +// neither: 0, +// }, +// ); +// let mut read_names: HashMap>> = HashMap::new(); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let result = predict(ordering_flags, read_names, 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 8); +// assert_eq!(result.last, 8); +// assert_eq!(result.both, 2); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, Some(2.2)); +// assert_eq!(result.read_groups.len(), 2); +// // We can't know which read group will be first in the vector. +// // But both should succeed. +// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); +// } +// } diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 71bd504..679e8d2 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -12,23 +12,23 @@ pub struct DerivedReadlenResult { pub succeeded: bool, /// The concsensus read length, if available. - pub consensus_read_length: Option, + pub consensus_read_length: Option, /// The majority vote percentage of the consensus read length, if available. pub majority_pct_detected: f64, /// Status of the evidence that supports (or does not support) this /// read length, if available. - pub evidence: Vec<(usize, i32)>, + pub evidence: Vec<(u32, u64)>, } impl DerivedReadlenResult { /// Creates a new [`DerivedReadlenResult`]. pub fn new( succeeded: bool, - consensus_read_length: Option, + consensus_read_length: Option, majority_pct_detected: f64, - evidence: Vec<(usize, i32)>, + evidence: Vec<(u32, u64)>, ) -> Self { DerivedReadlenResult { succeeded, @@ -43,7 +43,7 @@ impl DerivedReadlenResult { /// return a result for the consensus read length. This may fail, and the /// resulting [`DerivedReadlenResult`] should be evaluated accordingly. pub fn predict( - read_lengths: HashMap, + read_lengths: HashMap, majority_vote_cutoff: f64, ) -> Result { let mut num_records = 0; @@ -66,7 +66,7 @@ pub fn predict( let majority_detected = max_count as f64 / num_records as f64; // Sort the read lengths by their key for output. - let mut read_lengths: Vec<(usize, i32)> = read_lengths.into_iter().collect(); + let mut read_lengths: Vec<(u32, u64)> = read_lengths.into_iter().collect(); read_lengths.sort_by(|a, b| b.0.cmp(&a.0)); let mut result = From e3b8e859497ae1e5f1c4b4c4d9674b7ac858ce20 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 09:21:02 -0500 Subject: [PATCH 18/91] tests(derive/command/endedness): rewrite tests with latest changes --- src/derive/endedness/compute.rs | 516 +++++++++++++++----------------- 1 file changed, 238 insertions(+), 278 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 86ccad3..f4f6869 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -361,281 +361,241 @@ pub fn predict( Ok(final_result) } -// #[cfg(test)] -// mod tests { -// use super::*; - -// #[test] -// fn test_predict_endedness() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict_endedness( -// "overall".to_string(), -// &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), -// 0.0, -// None, -// false, -// ); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// } - -// #[test] -// fn test_derive_endedness_from_all_zero_counts() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_err()); -// } - -// #[test] -// fn test_derive_endedness_from_only_first() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 0, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_both() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 1, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Single-End"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 1); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_neither() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 0, -// neither: 1, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 1); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_calculate_reads_per_template() { -// let mut read_names: HashMap>> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let results = calculate_reads_per_template(read_names); -// assert_eq!(results.len(), 3); -// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); -// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); -// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last_with_rpt() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 2, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_paired), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 0, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_single), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 2, -// neither: 0, -// }, -// ); -// let mut read_names: HashMap>> = HashMap::new(); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let result = predict(ordering_flags, read_names, 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 8); -// assert_eq!(result.last, 8); -// assert_eq!(result.both, 2); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, Some(2.2)); -// assert_eq!(result.read_groups.len(), 2); -// // We can't know which read group will be first in the vector. -// // But both should succeed. -// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); -// } -// } +#[cfg(test)] +mod tests { + use super::*; + + #[test] + fn test_predict_endedness() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict_endedness( + "overall".to_string(), + &ordering_flags.get(OVERALL.as_str()).unwrap(), + 0.0, + None, + false, + ); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_all_zero_counts() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(OVERALL.as_str(), OrderingFlagsCounts::new()); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_err()); + } + + #[test] + fn test_derive_endedness_from_only_first() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 1, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_last() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 0, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_both() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 1, + neither: 0, + }, + ); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 1); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_neither() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 1, + }, + ); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 1); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_first_and_last() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_calculate_reads_per_template() { + let mut read_names: HashMap> = HashMap::new(); + let rg_paired = "rg_paired"; + let rg_single = "rg_single"; + read_names.insert("read1".to_string(), vec![rg_paired, rg_paired]); + read_names.insert("read2".to_string(), vec![rg_paired, rg_paired, rg_single]); + read_names.insert("read3".to_string(), vec![rg_single]); + read_names.insert("read4".to_string(), vec![rg_paired, rg_paired]); + read_names.insert("read5".to_string(), vec![rg_paired, rg_paired, rg_single]); + let results = calculate_reads_per_template(&read_names); + assert_eq!(results.len(), 3); + assert_eq!(results.get("overall").unwrap(), &2.2); + assert_eq!(results.get(rg_paired).unwrap(), &2.0); + assert_eq!(results.get(rg_single).unwrap(), &1.0); + } + + #[test] + fn test_derive_endedness_from_first_and_last_with_rpt() { + let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let rg_paired = "rg_paired"; + let rg_single = "rg_single"; + ordering_flags.insert( + OVERALL.as_str(), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 2, + neither: 0, + }, + ); + ordering_flags.insert( + rg_paired, + OrderingFlagsCounts { + first: 8, + last: 8, + both: 0, + neither: 0, + }, + ); + ordering_flags.insert( + rg_single, + OrderingFlagsCounts { + first: 0, + last: 0, + both: 2, + neither: 0, + }, + ); + let mut read_names: HashMap> = HashMap::new(); + read_names.insert("read1".to_string(), vec![rg_paired, rg_paired]); + read_names.insert("read2".to_string(), vec![rg_paired, rg_paired, rg_single]); + read_names.insert("read3".to_string(), vec![rg_single]); + read_names.insert("read4".to_string(), vec![rg_paired, rg_paired]); + read_names.insert("read5".to_string(), vec![rg_paired, rg_paired, rg_single]); + let result = predict(&ordering_flags, &read_names, 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 8); + assert_eq!(result.last, 8); + assert_eq!(result.both, 2); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, Some(2.2)); + assert_eq!(result.read_groups.len(), 2); + // We can't know which read group will be first in the vector. + // But both should succeed. + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); + } +} From 8b23de091c13a12f9b7444affbd0361a370ea197 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 09:27:44 -0500 Subject: [PATCH 19/91] chore(endedness): handle error where RG tag can't be parsed as_str() --- src/derive/command/endedness.rs | 14 ++++++++------ 1 file changed, 8 insertions(+), 6 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index f7dda29..be0c1ba 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -94,13 +94,15 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => { - let rg = rg.as_str().unwrap(); - if !found_rgs.contains(rg) { - found_rgs.insert(rg.to_string()); + Some(rg) => match rg.as_str() { + Some(rg) => { + if !found_rgs.contains(rg) { + found_rgs.insert(rg.to_string()); + } + found_rgs.get(rg).unwrap() } - found_rgs.get(rg).unwrap() - } + None => UNKNOWN_READ_GROUP.as_str(), + }, None => UNKNOWN_READ_GROUP.as_str(), }; From 65ea501b1883ed0f4de39e29c56fb9e9dc7d5cf1 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 09:52:29 -0500 Subject: [PATCH 20/91] fix(endedness): remove all Arcs and lazy_statics --- Cargo.lock | 1 - Cargo.toml | 1 - src/derive/command/endedness.rs | 19 +++++++--------- src/derive/endedness/compute.rs | 39 +++++++++++++-------------------- 4 files changed, 23 insertions(+), 37 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index 925f544..f6669ee 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -867,7 +867,6 @@ dependencies = [ "indexmap", "indicatif", "itertools", - "lazy_static", "noodles", "num-format", "plotly", diff --git a/Cargo.toml b/Cargo.toml index c9081d8..306ad22 100644 --- a/Cargo.toml +++ b/Cargo.toml @@ -19,7 +19,6 @@ git-testament = "0.2.1" indexmap = "1.9.1" indicatif = "0.16.2" itertools = "0.10.5" -lazy_static = "1.4.0" noodles = { version = "0.34.0", features = [ "async", "bam", diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index be0c1ba..56a246b 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -3,7 +3,6 @@ use std::collections::HashMap; use std::collections::HashSet; use std::path::PathBuf; -use std::sync::Arc; use clap::Args; use noodles::sam::record::data::field::Tag; @@ -64,8 +63,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut found_rgs = HashSet::new(); let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(OVERALL.as_str(), OrderingFlagsCounts::new()); - ordering_flags.insert(UNKNOWN_READ_GROUP.as_str(), OrderingFlagsCounts::new()); + ordering_flags.insert(OVERALL, OrderingFlagsCounts::new()); + ordering_flags.insert(UNKNOWN_READ_GROUP, OrderingFlagsCounts::new()); // only used if args.calc_rpt is true let mut read_names: HashMap> = HashMap::new(); @@ -101,9 +100,9 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } found_rgs.get(rg).unwrap() } - None => UNKNOWN_READ_GROUP.as_str(), + None => UNKNOWN_READ_GROUP, }, - None => UNKNOWN_READ_GROUP.as_str(), + None => UNKNOWN_READ_GROUP, }; if args.calc_rpt { @@ -129,10 +128,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } - let overall_rg = OVERALL.as_str(); - if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { + ordering_flags.entry(OVERALL).and_modify(|e| { e.first += 1; }); @@ -148,7 +145,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { + ordering_flags.entry(OVERALL).and_modify(|e| { e.last += 1; }); @@ -164,7 +161,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { + ordering_flags.entry(OVERALL).and_modify(|e| { e.both += 1; }); @@ -180,7 +177,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { + ordering_flags.entry(OVERALL).and_modify(|e| { e.neither += 1; }); diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index f4f6869..247ef58 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,22 +1,16 @@ //! Module holding the logic for computing the endedness of a BAM. use anyhow::bail; -use lazy_static::lazy_static; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; -use std::sync::Arc; use tracing::warn; -// Strings used to index into the HashMaps used to store the Read Group ordering flags. -// Lazy statics are used to save memory. -lazy_static! { - /// String used to index into the HashMaps used to store the "overall" ordering flags. - pub static ref OVERALL: Arc = Arc::new(String::from("overall")); +/// String used to index into the HashMaps used to store the "overall" ordering flags. +pub static OVERALL: &str = "overall"; - /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. - pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); -} +/// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. +pub static UNKNOWN_READ_GROUP: &str = "unknown_read_group"; /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] @@ -204,10 +198,7 @@ fn calculate_reads_per_template<'rg>( } } - reads_per_template.insert( - OVERALL.as_str(), - total_reads as f64 / total_templates as f64, - ); + reads_per_template.insert(OVERALL, total_reads as f64 / total_templates as f64); for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); @@ -330,7 +321,7 @@ pub fn predict( ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - if (*read_group == UNKNOWN_READ_GROUP.as_str()) + if (*read_group == UNKNOWN_READ_GROUP) && (rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 @@ -369,7 +360,7 @@ mod tests { fn test_predict_endedness() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 1, last: 1, @@ -379,7 +370,7 @@ mod tests { ); let result = predict_endedness( "overall".to_string(), - &ordering_flags.get(OVERALL.as_str()).unwrap(), + &ordering_flags.get(OVERALL).unwrap(), 0.0, None, false, @@ -398,7 +389,7 @@ mod tests { #[test] fn test_derive_endedness_from_all_zero_counts() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(OVERALL.as_str(), OrderingFlagsCounts::new()); + ordering_flags.insert(OVERALL, OrderingFlagsCounts::new()); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); assert!(result.is_err()); } @@ -407,7 +398,7 @@ mod tests { fn test_derive_endedness_from_only_first() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 1, last: 0, @@ -432,7 +423,7 @@ mod tests { fn test_derive_endedness_from_only_last() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 0, last: 1, @@ -457,7 +448,7 @@ mod tests { fn test_derive_endedness_from_only_both() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 0, last: 0, @@ -482,7 +473,7 @@ mod tests { fn test_derive_endedness_from_only_neither() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 0, last: 0, @@ -507,7 +498,7 @@ mod tests { fn test_derive_endedness_from_first_and_last() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 1, last: 1, @@ -551,7 +542,7 @@ mod tests { let rg_paired = "rg_paired"; let rg_single = "rg_single"; ordering_flags.insert( - OVERALL.as_str(), + OVERALL, OrderingFlagsCounts { first: 8, last: 8, From 76305f2a22d481578dd3fa53f07c6601fd6b6806 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 10:17:56 -0500 Subject: [PATCH 21/91] refactor(derive/readlen): move num_samples counting to outer func --- src/derive/command/readlen.rs | 11 +++++------ src/derive/readlen/compute.rs | 15 +++++++-------- 2 files changed, 12 insertions(+), 14 deletions(-) diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 9449b03..6d1c944 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -70,16 +70,15 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { .and_modify(|e| *e += 1) .or_insert(1 as u64); - if sample_max > 0 { - samples += 1; - if samples > sample_max { - break; - } + samples += 1; + if sample_max > 0 && samples > sample_max { + break; } } // (2) Derive the consensus read length based on the read lengths gathered. - let result = compute::predict(read_lengths, args.majority_vote_cutoff.unwrap()).unwrap(); + let result = + compute::predict(read_lengths, samples, args.majority_vote_cutoff.unwrap()).unwrap(); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 679e8d2..72c1c59 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -44,26 +44,25 @@ impl DerivedReadlenResult { /// resulting [`DerivedReadlenResult`] should be evaluated accordingly. pub fn predict( read_lengths: HashMap, + num_samples: u64, majority_vote_cutoff: f64, ) -> Result { - let mut num_records = 0; let mut max_count = 0; let mut max_read_length = 0; for (read_length, count) in &read_lengths { - num_records += *count; if *read_length > max_read_length { max_read_length = *read_length; max_count = *count; } } - if num_records == 0 { + if num_samples <= 0 { bail!("No read lengths were detected in the file."); } let consensus_read_length = max_read_length; - let majority_detected = max_count as f64 / num_records as f64; + let majority_detected = max_count as f64 / num_samples as f64; // Sort the read lengths by their key for output. let mut read_lengths: Vec<(u32, u64)> = read_lengths.into_iter().collect(); @@ -87,14 +86,14 @@ mod tests { #[test] fn test_derive_readlen_from_empty_hashmap() { let read_lengths = HashMap::new(); - let result = predict(read_lengths, 0.7); + let result = predict(read_lengths, 0, 0.7); assert!(result.is_err()); } #[test] fn test_derive_readlen_when_all_readlengths_equal() { let read_lengths = HashMap::from([(100, 10)]); - let result = predict(read_lengths, 1.0).unwrap(); + let result = predict(read_lengths, 10, 1.0).unwrap(); assert!(result.succeeded); assert_eq!(result.consensus_read_length, Some(100)); assert_eq!(result.majority_pct_detected, 100.0); @@ -104,7 +103,7 @@ mod tests { #[test] fn test_derive_readlen_success_when_not_all_readlengths_equal() { let read_lengths = HashMap::from([(101, 1000), (100, 5), (99, 5)]); - let result = predict(read_lengths, 0.7).unwrap(); + let result = predict(read_lengths, 1010, 0.7).unwrap(); assert!(result.succeeded); assert_eq!(result.consensus_read_length, Some(101)); assert!(result.majority_pct_detected > 99.0); @@ -114,7 +113,7 @@ mod tests { #[test] fn test_derive_readlen_fail_when_not_all_readlengths_equal() { let read_lengths = HashMap::from([(101, 5), (100, 1000), (99, 5)]); - let result = predict(read_lengths, 0.7).unwrap(); + let result = predict(read_lengths, 1010, 0.7).unwrap(); assert!(!result.succeeded); assert_eq!(result.consensus_read_length, None); assert!(result.majority_pct_detected < 0.7); From d63eb2a99466de2ffa970cd136ca720b3ca76934 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 10:20:46 -0500 Subject: [PATCH 22/91] perf(derive/readlen): don't iterate through all read_lengths --- src/derive/readlen/compute.rs | 19 ++++++------------- 1 file changed, 6 insertions(+), 13 deletions(-) diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 72c1c59..3dac219 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -47,27 +47,20 @@ pub fn predict( num_samples: u64, majority_vote_cutoff: f64, ) -> Result { - let mut max_count = 0; - let mut max_read_length = 0; - - for (read_length, count) in &read_lengths { - if *read_length > max_read_length { - max_read_length = *read_length; - max_count = *count; - } - } - if num_samples <= 0 { bail!("No read lengths were detected in the file."); } - let consensus_read_length = max_read_length; - let majority_detected = max_count as f64 / num_samples as f64; - // Sort the read lengths by their key for output. let mut read_lengths: Vec<(u32, u64)> = read_lengths.into_iter().collect(); read_lengths.sort_by(|a, b| b.0.cmp(&a.0)); + let max_read_length = read_lengths[0].0; + let max_count = read_lengths[0].1; + + let consensus_read_length = max_read_length; + let majority_detected = max_count as f64 / num_samples as f64; + let mut result = DerivedReadlenResult::new(false, None, majority_detected * 100.0, read_lengths); From cb1d2f59473b0b67322047b6b434f584fe49cdcd Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 11:12:22 -0500 Subject: [PATCH 23/91] feat(derive/endedness): add `validate_read_group_info()` call --- src/derive/command/endedness.rs | 10 +++- src/derive/endedness/compute.rs | 99 +++++++++++++++++++++++---------- 2 files changed, 78 insertions(+), 31 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 56a246b..5dfeed0 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -10,7 +10,9 @@ use tracing::info; use tracing::trace; use crate::derive::endedness::compute; -use crate::derive::endedness::compute::{OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP}; +use crate::derive::endedness::compute::{ + validate_read_group_info, OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP, +}; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -204,6 +206,12 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } + // (1.5) Validate the read group information. + let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed) + for rg_id in rgs_in_header_not_records { + ordering_flags.insert(&rg_id, OrderingFlagsCounts::new()); + } + // (2) Derive the consensus endedness based on the ordering flags gathered. let result = compute::predict( &ordering_flags, diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 247ef58..085cfa5 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,6 +1,6 @@ //! Module holding the logic for computing the endedness of a BAM. -use anyhow::bail; +use noodles::sam::header; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; @@ -151,6 +151,40 @@ impl DerivedEndednessResult { } } +pub fn validate_read_group_info( + found_rgs: &HashSet, + header: &header::Header, +) -> Vec { + let mut rgs_in_header_not_records = Vec::new(); + let mut rgs_in_records_not_header = Vec::new(); + + for (rg_id, _) in header.read_groups() { + if !found_rgs.contains(rg_id) { + rgs_in_header_not_records.push(rg_id.to_string()); + } + } + if !rgs_in_header_not_records.is_empty() { + warn!( + "The following read groups were not found in the file: {:?}", + rgs_in_header_not_records + ); + } + + for rg_id in found_rgs { + if !header.read_groups().contains_key(rg_id) { + rgs_in_records_not_header.push(rg_id.to_string()); + } + } + if !rgs_in_records_not_header.is_empty() { + warn!( + "The following read groups were not found in the header: {:?}", + rgs_in_records_not_header + ); + } + + return rgs_in_header_not_records; +} + fn calculate_reads_per_template<'rg>( read_names: &HashMap>, ) -> HashMap<&'rg str, f64> { @@ -215,7 +249,7 @@ fn predict_endedness( paired_deviance: f64, reads_per_template: Option<&f64>, round_rpt: bool, -) -> Result { +) -> ReadGroupDerivedEndednessResult { let first = rg_ordering_flags.first; let last = rg_ordering_flags.last; let both = rg_ordering_flags.both; @@ -224,10 +258,17 @@ fn predict_endedness( // all zeroes (Perform this check before creating the result struct // so that we don't have to clone the read group name) if first == 0 && last == 0 && both == 0 && neither == 0 { - bail!( + warn!( "No reads were detected in this read group: {}", read_group_name ); + return ReadGroupDerivedEndednessResult::new( + read_group_name, + false, + "Unknown".to_string(), + rg_ordering_flags.clone(), + reads_per_template.copied(), + ); } let mut result = ReadGroupDerivedEndednessResult::new( @@ -240,11 +281,11 @@ fn predict_endedness( // only first present if first > 0 && last == 0 && both == 0 && neither == 0 { - return Ok(result); + return result; } // only last present if first == 0 && last > 0 && both == 0 && neither == 0 { - return Ok(result); + return result; } // only both present if first == 0 && last == 0 && both > 0 && neither == 0 { @@ -260,19 +301,19 @@ fn predict_endedness( result.endedness = String::from("Single-End"); } } - return Ok(result); + return result; } // only neither present if first == 0 && last == 0 && both == 0 && neither > 0 { - return Ok(result); + return result; } // first/last mixed with both/neither if (first > 0 || last > 0) && (both > 0 || neither > 0) { - return Ok(result); + return result; } // any mix of both/neither, regardless of first/last if both > 0 && neither > 0 { - return Ok(result); + return result; } // both and neither are now guarenteed to be 0 @@ -295,7 +336,7 @@ fn predict_endedness( } } } - Ok(result) + result } /// Main method to evaluate the collected ordering flags and @@ -306,7 +347,7 @@ pub fn predict( read_names: &HashMap>, paired_deviance: f64, round_rpt: bool, -) -> Result { +) -> DerivedEndednessResult { let mut rpts: HashMap<&str, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); @@ -335,7 +376,7 @@ pub fn predict( paired_deviance, rpts.get(read_group), round_rpt, - )?; + ); if result.read_group == "overall" { final_result.endedness = result.endedness; final_result.first = result.first; @@ -349,7 +390,7 @@ pub fn predict( } } - Ok(final_result) + final_result } #[cfg(test)] @@ -375,8 +416,6 @@ mod tests { None, false, ); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Paired-End"); assert_eq!(result.first, 1); @@ -389,9 +428,21 @@ mod tests { #[test] fn test_derive_endedness_from_all_zero_counts() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(OVERALL, OrderingFlagsCounts::new()); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_err()); + ordering_flags.insert("rg1", OrderingFlagsCounts::new()); + let result = predict_endedness( + "rg1".to_string(), + &ordering_flags.get("rg1").unwrap(), + 0.0, + None, + false, + ); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); } #[test] @@ -407,8 +458,6 @@ mod tests { }, ); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 1); @@ -432,8 +481,6 @@ mod tests { }, ); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 0); @@ -457,8 +504,6 @@ mod tests { }, ); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Single-End"); assert_eq!(result.first, 0); @@ -482,8 +527,6 @@ mod tests { }, ); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 0); @@ -507,8 +550,6 @@ mod tests { }, ); let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Paired-End"); assert_eq!(result.first, 1); @@ -575,8 +616,6 @@ mod tests { read_names.insert("read4".to_string(), vec![rg_paired, rg_paired]); read_names.insert("read5".to_string(), vec![rg_paired, rg_paired, rg_single]); let result = predict(&ordering_flags, &read_names, 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 8); From 9b45316b82b20df40f21fb0c138a4bcedf2c9993 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 11:14:20 -0500 Subject: [PATCH 24/91] fix: typos --- src/derive/command/endedness.rs | 5 ++--- 1 file changed, 2 insertions(+), 3 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 5dfeed0..c8c60d9 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -207,7 +207,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } // (1.5) Validate the read group information. - let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed) + let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); for rg_id in rgs_in_header_not_records { ordering_flags.insert(&rg_id, OrderingFlagsCounts::new()); } @@ -218,8 +218,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { &read_names, args.paired_deviance.unwrap(), args.round_rpt, - ) - .unwrap(); + ); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). From 520ae466320d01a8aa8e22232536c114e1ca4cc2 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 13:39:10 -0500 Subject: [PATCH 25/91] revert --- Cargo.lock | 1 + Cargo.toml | 1 + src/derive/endedness/compute.rs | 38 +++++++++++++++++++++------------ 3 files changed, 26 insertions(+), 14 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index f6669ee..925f544 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -867,6 +867,7 @@ dependencies = [ "indexmap", "indicatif", "itertools", + "lazy_static", "noodles", "num-format", "plotly", diff --git a/Cargo.toml b/Cargo.toml index 306ad22..c9081d8 100644 --- a/Cargo.toml +++ b/Cargo.toml @@ -19,6 +19,7 @@ git-testament = "0.2.1" indexmap = "1.9.1" indicatif = "0.16.2" itertools = "0.10.5" +lazy_static = "1.4.0" noodles = { version = "0.34.0", features = [ "async", "bam", diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 085cfa5..a734c2c 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,16 +1,23 @@ //! Module holding the logic for computing the endedness of a BAM. +use anyhow::bail; +use lazy_static::lazy_static; use noodles::sam::header; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; +use std::sync::Arc; use tracing::warn; -/// String used to index into the HashMaps used to store the "overall" ordering flags. -pub static OVERALL: &str = "overall"; +// Strings used to index into the HashMaps used to store the Read Group ordering flags. +// Lazy statics are used to save memory. +lazy_static! { + /// String used to index into the HashMaps used to store the "overall" ordering flags. + pub static ref OVERALL: Arc = Arc::new(String::from("overall")); -/// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. -pub static UNKNOWN_READ_GROUP: &str = "unknown_read_group"; + /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. + pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); +} /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] @@ -232,7 +239,10 @@ fn calculate_reads_per_template<'rg>( } } - reads_per_template.insert(OVERALL, total_reads as f64 / total_templates as f64); + reads_per_template.insert( + OVERALL.as_str(), + total_reads as f64 / total_templates as f64, + ); for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); @@ -362,7 +372,7 @@ pub fn predict( ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - if (*read_group == UNKNOWN_READ_GROUP) + if (*read_group == UNKNOWN_READ_GROUP.as_str()) && (rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 @@ -401,7 +411,7 @@ mod tests { fn test_predict_endedness() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 1, last: 1, @@ -411,7 +421,7 @@ mod tests { ); let result = predict_endedness( "overall".to_string(), - &ordering_flags.get(OVERALL).unwrap(), + &ordering_flags.get(OVERALL.as_str()).unwrap(), 0.0, None, false, @@ -449,7 +459,7 @@ mod tests { fn test_derive_endedness_from_only_first() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 1, last: 0, @@ -472,7 +482,7 @@ mod tests { fn test_derive_endedness_from_only_last() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 0, last: 1, @@ -495,7 +505,7 @@ mod tests { fn test_derive_endedness_from_only_both() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 0, last: 0, @@ -518,7 +528,7 @@ mod tests { fn test_derive_endedness_from_only_neither() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 0, last: 0, @@ -541,7 +551,7 @@ mod tests { fn test_derive_endedness_from_first_and_last() { let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 1, last: 1, @@ -583,7 +593,7 @@ mod tests { let rg_paired = "rg_paired"; let rg_single = "rg_single"; ordering_flags.insert( - OVERALL, + OVERALL.as_str(), OrderingFlagsCounts { first: 8, last: 8, From f280c0fea2b580e8b91ec55a49b683aad31a33e2 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 13:56:53 -0500 Subject: [PATCH 26/91] revert --- src/derive/command/endedness.rs | 24 ++-- src/derive/command/readlen.rs | 13 +- src/derive/endedness/compute.rs | 208 ++++++++++++++++++++------------ src/derive/readlen/compute.rs | 14 +-- 4 files changed, 156 insertions(+), 103 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index c8c60d9..0782edc 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -37,8 +37,8 @@ pub struct DeriveEndednessArgs { src: PathBuf, /// Only examine the first n records in the file. - #[arg(short, long, value_name = "U64")] - num_records: Option, + #[arg(short, long, value_name = "USIZE")] + num_records: Option, /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate @@ -64,16 +64,16 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut found_rgs = HashSet::new(); - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(OVERALL, OrderingFlagsCounts::new()); - ordering_flags.insert(UNKNOWN_READ_GROUP, OrderingFlagsCounts::new()); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); + ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true - let mut read_names: HashMap> = HashMap::new(); + let mut read_names: HashMap>> = HashMap::new(); let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. @@ -115,10 +115,10 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { match rg_vec { Some(rg_vec) => { - rg_vec.push(read_group); + rg_vec.push(Arc::clone(&read_group)); } None => { - read_names.insert(rn, vec![read_group]); + read_names.insert(rn, vec![(Arc::clone(&read_group))]); } } } @@ -130,6 +130,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } + let overall_rg = Arc::clone(&OVERALL); + if record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry(OVERALL).and_modify(|e| { e.first += 1; @@ -214,8 +216,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (2) Derive the consensus endedness based on the ordering flags gathered. let result = compute::predict( - &ordering_flags, - &read_names, + ordering_flags, + read_names, args.paired_deviance.unwrap(), args.round_rpt, ); diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 6d1c944..64d18bd 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -33,8 +33,8 @@ pub struct DeriveReadlenArgs { src: PathBuf, /// Only examine the first n records in the file. - #[arg(short, long, value_name = "U64")] - num_records: Option, + #[arg(short, long, value_name = "USIZE")] + num_records: Option, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. #[arg(short, long, value_name = "F64", default_value = "0.7")] @@ -50,7 +50,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. @@ -63,12 +63,9 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { for result in reader.records(&header.parsed) { let record = result?; - let len = record.sequence().len() as u32; + let len = record.sequence().len(); - read_lengths - .entry(len) - .and_modify(|e| *e += 1) - .or_insert(1 as u64); + read_lengths.entry(len).and_modify(|e| *e += 1).or_insert(1); samples += 1; if sample_max > 0 && samples > sample_max { diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index a734c2c..6c105c3 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -23,16 +23,16 @@ lazy_static! { #[derive(Debug, Clone)] pub struct OrderingFlagsCounts { /// The number of reads with the first in template flag set. - pub first: u64, + pub first: usize, /// The number of reads with the last in template flag set. - pub last: u64, + pub last: usize, /// The number of reads with both the first and last in template flags set. - pub both: u64, + pub both: usize, /// The number of reads with neither the first nor last in template flags set. - pub neither: u64, + pub neither: usize, } impl OrderingFlagsCounts { /// Creates a new [`OrderingFlagsCounts`]. @@ -66,16 +66,16 @@ pub struct ReadGroupDerivedEndednessResult { pub endedness: String, /// The f+l- read count. - pub first: u64, + pub first: usize, /// The f-l+ read count. - pub last: u64, + pub last: usize, /// The f+l+ read count. - pub both: u64, + pub both: usize, /// The f-l- read count. - pub neither: u64, + pub neither: usize, /// The reads per template (RPT). /// Only available if `args.calc_rpt` is true. @@ -115,16 +115,16 @@ pub struct DerivedEndednessResult { pub endedness: String, /// The overall f+l- read count. - pub first: u64, + pub first: usize, /// The overall f-l+ read count. - pub last: u64, + pub last: usize, /// The overall f+l+ read count. - pub both: u64, + pub both: usize, /// The overall f-l- read count. - pub neither: u64, + pub neither: usize, /// The overall reads per template (RPT). /// Only available if `args.calc_rpt` is true. @@ -192,27 +192,27 @@ pub fn validate_read_group_info( return rgs_in_header_not_records; } -fn calculate_reads_per_template<'rg>( - read_names: &HashMap>, -) -> HashMap<&'rg str, f64> { - let mut reads_per_template: HashMap<&str, f64> = HashMap::new(); - let mut total_reads: u64 = 0; - let mut total_templates: u64 = 0; - let mut read_group_reads: HashMap<&str, u64> = HashMap::new(); - let mut read_group_templates: HashMap<&str, u64> = HashMap::new(); +fn calculate_reads_per_template( + read_names: HashMap>>, +) -> HashMap, f64> { + let mut reads_per_template: HashMap, f64> = HashMap::new(); + let mut total_reads: usize = 0; + let mut total_templates: usize = 0; + let mut read_group_reads: HashMap, usize> = HashMap::new(); + let mut read_group_templates: HashMap, usize> = HashMap::new(); for (read_name, read_groups) in read_names.iter() { - let num_reads = read_groups.len() as u64; + let num_reads = read_groups.len(); total_reads += num_reads; total_templates += 1; - let read_group_set: HashSet<&str> = read_groups.iter().cloned().collect(); + let read_group_set: HashSet> = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { - let read_group = read_group_set.iter().next().unwrap(); + let read_group = Arc::clone(read_group_set.iter().next().unwrap()); read_group_reads - .entry(&read_group) + .entry(Arc::clone(&read_group)) .and_modify(|e| *e += num_reads) .or_insert(num_reads); read_group_templates @@ -226,7 +226,7 @@ fn calculate_reads_per_template<'rg>( ); for read_group in read_groups { read_group_reads - .entry(&read_group) + .entry(Arc::clone(read_group)) .and_modify(|e| *e += 1) .or_insert(1); } @@ -240,14 +240,14 @@ fn calculate_reads_per_template<'rg>( } reads_per_template.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), total_reads as f64 / total_templates as f64, ); for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); let rpt = *num_reads as f64 / *num_templates as f64; - reads_per_template.insert(read_group, rpt); + reads_per_template.insert(Arc::clone(read_group), rpt); } reads_per_template @@ -301,7 +301,7 @@ fn predict_endedness( if first == 0 && last == 0 && both > 0 && neither == 0 { match reads_per_template { Some(rpt) => { - if *rpt == 1.0 || (round_rpt && rpt.round() as u64 == 1) { + if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { result.succeeded = true; result.endedness = String::from("Single-End"); } @@ -335,7 +335,7 @@ fn predict_endedness( if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { match reads_per_template { Some(rpt) => { - if *rpt == 2.0 || (round_rpt && rpt.round() as u64 == 2) { + if *rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { result.succeeded = true; result.endedness = String::from("Paired-End"); } @@ -353,12 +353,12 @@ fn predict_endedness( /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: &HashMap<&str, OrderingFlagsCounts>, - read_names: &HashMap>, + ordering_flags: HashMap, OrderingFlagsCounts>, + read_names: HashMap>>, paired_deviance: f64, round_rpt: bool, -) -> DerivedEndednessResult { - let mut rpts: HashMap<&str, f64> = HashMap::new(); +) -> Result { + let mut rpts: HashMap, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); } @@ -372,7 +372,7 @@ pub fn predict( ); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - if (*read_group == UNKNOWN_READ_GROUP.as_str()) + if (*read_group == *UNKNOWN_READ_GROUP) && (rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 @@ -409,9 +409,9 @@ mod tests { #[test] fn test_predict_endedness() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 1, last: 1, @@ -421,11 +421,13 @@ mod tests { ); let result = predict_endedness( "overall".to_string(), - &ordering_flags.get(OVERALL.as_str()).unwrap(), + &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), 0.0, None, false, ); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Paired-End"); assert_eq!(result.first, 1); @@ -457,9 +459,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 1, last: 0, @@ -467,7 +469,9 @@ mod tests { neither: 0, }, ); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 1); @@ -480,9 +484,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 1, @@ -490,7 +494,9 @@ mod tests { neither: 0, }, ); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 0); @@ -503,9 +509,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 0, @@ -513,7 +519,9 @@ mod tests { neither: 0, }, ); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Single-End"); assert_eq!(result.first, 0); @@ -526,9 +534,9 @@ mod tests { #[test] fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 0, last: 0, @@ -536,7 +544,9 @@ mod tests { neither: 1, }, ); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 0); @@ -549,9 +559,9 @@ mod tests { #[test] fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 1, last: 1, @@ -559,7 +569,9 @@ mod tests { neither: 0, }, ); - let result = predict(&ordering_flags, &HashMap::new(), 0.0, false); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(result.succeeded); assert_eq!(result.endedness, "Paired-End"); assert_eq!(result.first, 1); @@ -572,28 +584,48 @@ mod tests { #[test] fn test_calculate_reads_per_template() { - let mut read_names: HashMap> = HashMap::new(); - let rg_paired = "rg_paired"; - let rg_single = "rg_single"; - read_names.insert("read1".to_string(), vec![rg_paired, rg_paired]); - read_names.insert("read2".to_string(), vec![rg_paired, rg_paired, rg_single]); - read_names.insert("read3".to_string(), vec![rg_single]); - read_names.insert("read4".to_string(), vec![rg_paired, rg_paired]); - read_names.insert("read5".to_string(), vec![rg_paired, rg_paired, rg_single]); - let results = calculate_reads_per_template(&read_names); + let mut read_names: HashMap>> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let results = calculate_reads_per_template(read_names); assert_eq!(results.len(), 3); - assert_eq!(results.get("overall").unwrap(), &2.2); - assert_eq!(results.get(rg_paired).unwrap(), &2.0); - assert_eq!(results.get(rg_single).unwrap(), &1.0); + assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); + assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); + assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); } #[test] fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - let rg_paired = "rg_paired"; - let rg_single = "rg_single"; + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); ordering_flags.insert( - OVERALL.as_str(), + Arc::clone(&OVERALL), OrderingFlagsCounts { first: 8, last: 8, @@ -602,7 +634,7 @@ mod tests { }, ); ordering_flags.insert( - rg_paired, + Arc::clone(&rg_paired), OrderingFlagsCounts { first: 8, last: 8, @@ -611,7 +643,7 @@ mod tests { }, ); ordering_flags.insert( - rg_single, + Arc::clone(&rg_single), OrderingFlagsCounts { first: 0, last: 0, @@ -619,13 +651,35 @@ mod tests { neither: 0, }, ); - let mut read_names: HashMap> = HashMap::new(); - read_names.insert("read1".to_string(), vec![rg_paired, rg_paired]); - read_names.insert("read2".to_string(), vec![rg_paired, rg_paired, rg_single]); - read_names.insert("read3".to_string(), vec![rg_single]); - read_names.insert("read4".to_string(), vec![rg_paired, rg_paired]); - read_names.insert("read5".to_string(), vec![rg_paired, rg_paired, rg_single]); - let result = predict(&ordering_flags, &read_names, 0.0, false); + let mut read_names: HashMap>> = HashMap::new(); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let result = predict(ordering_flags, read_names, 0.0, false); + assert!(result.is_ok()); + let result = result.unwrap(); assert!(!result.succeeded); assert_eq!(result.endedness, "Unknown"); assert_eq!(result.first, 8); diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 3dac219..883d95a 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -12,23 +12,23 @@ pub struct DerivedReadlenResult { pub succeeded: bool, /// The concsensus read length, if available. - pub consensus_read_length: Option, + pub consensus_read_length: Option, /// The majority vote percentage of the consensus read length, if available. pub majority_pct_detected: f64, /// Status of the evidence that supports (or does not support) this /// read length, if available. - pub evidence: Vec<(u32, u64)>, + pub evidence: Vec<(usize, usize)>, } impl DerivedReadlenResult { /// Creates a new [`DerivedReadlenResult`]. pub fn new( succeeded: bool, - consensus_read_length: Option, + consensus_read_length: Option, majority_pct_detected: f64, - evidence: Vec<(u32, u64)>, + evidence: Vec<(usize, usize)>, ) -> Self { DerivedReadlenResult { succeeded, @@ -43,8 +43,8 @@ impl DerivedReadlenResult { /// return a result for the consensus read length. This may fail, and the /// resulting [`DerivedReadlenResult`] should be evaluated accordingly. pub fn predict( - read_lengths: HashMap, - num_samples: u64, + read_lengths: HashMap, + num_samples: usize, majority_vote_cutoff: f64, ) -> Result { if num_samples <= 0 { @@ -52,7 +52,7 @@ pub fn predict( } // Sort the read lengths by their key for output. - let mut read_lengths: Vec<(u32, u64)> = read_lengths.into_iter().collect(); + let mut read_lengths: Vec<(usize, usize)> = read_lengths.into_iter().collect(); read_lengths.sort_by(|a, b| b.0.cmp(&a.0)); let max_read_length = read_lengths[0].0; From 12f3f664007dda11d010e4dae4adbb2228cb013e Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 14:17:41 -0500 Subject: [PATCH 27/91] fix: corrections made after previous reverts --- src/derive/command/endedness.rs | 27 +- src/derive/endedness/compute.rs | 593 ++++++++++++++++---------------- src/derive/readlen/compute.rs | 2 +- 3 files changed, 312 insertions(+), 310 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 0782edc..239dab8 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -3,6 +3,7 @@ use std::collections::HashMap; use std::collections::HashSet; use std::path::PathBuf; +use std::sync::Arc; use clap::Args; use noodles::sam::record::data::field::Tag; @@ -95,16 +96,14 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => match rg.as_str() { - Some(rg) => { - if !found_rgs.contains(rg) { - found_rgs.insert(rg.to_string()); - } - found_rgs.get(rg).unwrap() + Some(rg) => { + let rg = rg.to_string(); + if !found_rgs.contains(&rg) { + found_rgs.insert(Arc::new(rg.clone())); } - None => UNKNOWN_READ_GROUP, - }, - None => UNKNOWN_READ_GROUP, + Arc::clone(found_rgs.get(&rg).unwrap()) + } + None => Arc::clone(&UNKNOWN_READ_GROUP), }; if args.calc_rpt { @@ -133,7 +132,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let overall_rg = Arc::clone(&OVERALL); if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(OVERALL).and_modify(|e| { + ordering_flags.entry(overall_rg).and_modify(|e| { e.first += 1; }); @@ -149,7 +148,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(OVERALL).and_modify(|e| { + ordering_flags.entry(overall_rg).and_modify(|e| { e.last += 1; }); @@ -165,7 +164,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(OVERALL).and_modify(|e| { + ordering_flags.entry(overall_rg).and_modify(|e| { e.both += 1; }); @@ -181,7 +180,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(OVERALL).and_modify(|e| { + ordering_flags.entry(overall_rg).and_modify(|e| { e.neither += 1; }); @@ -211,7 +210,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (1.5) Validate the read group information. let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); for rg_id in rgs_in_header_not_records { - ordering_flags.insert(&rg_id, OrderingFlagsCounts::new()); + ordering_flags.insert(Arc::new(rg_id), OrderingFlagsCounts::new()); } // (2) Derive the consensus endedness based on the ordering flags gathered. diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 6c105c3..0a53a36 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,6 +1,5 @@ //! Module holding the logic for computing the endedness of a BAM. -use anyhow::bail; use lazy_static::lazy_static; use noodles::sam::header; use serde::Serialize; @@ -158,8 +157,12 @@ impl DerivedEndednessResult { } } +/// Compares the read group tags found in the records +/// and the read groups found in the header. +/// Returns a vector of read group names that were found in the header +/// but not in the records. pub fn validate_read_group_info( - found_rgs: &HashSet, + found_rgs: &HashSet>, header: &header::Header, ) -> Vec { let mut rgs_in_header_not_records = Vec::new(); @@ -178,7 +181,7 @@ pub fn validate_read_group_info( } for rg_id in found_rgs { - if !header.read_groups().contains_key(rg_id) { + if !header.read_groups().contains_key(rg_id.as_str()) { rgs_in_records_not_header.push(rg_id.to_string()); } } @@ -189,7 +192,7 @@ pub fn validate_read_group_info( ); } - return rgs_in_header_not_records; + rgs_in_header_not_records } fn calculate_reads_per_template( @@ -357,7 +360,7 @@ pub fn predict( read_names: HashMap>>, paired_deviance: f64, round_rpt: bool, -) -> Result { +) -> DerivedEndednessResult { let mut rpts: HashMap, f64> = HashMap::new(); if !read_names.is_empty() { rpts = calculate_reads_per_template(read_names); @@ -403,293 +406,293 @@ pub fn predict( final_result } -#[cfg(test)] -mod tests { - use super::*; - - #[test] - fn test_predict_endedness() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict_endedness( - "overall".to_string(), - &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), - 0.0, - None, - false, - ); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - } - - #[test] - fn test_derive_endedness_from_all_zero_counts() { - let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert("rg1", OrderingFlagsCounts::new()); - let result = predict_endedness( - "rg1".to_string(), - &ordering_flags.get("rg1").unwrap(), - 0.0, - None, - false, - ); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - } - - #[test] - fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 0, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 1, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Single-End"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 1); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 0, - neither: 1, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 1); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_calculate_reads_per_template() { - let mut read_names: HashMap>> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let results = calculate_reads_per_template(read_names); - assert_eq!(results.len(), 3); - assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); - assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); - assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); - } - - #[test] - fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 2, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_paired), - OrderingFlagsCounts { - first: 8, - last: 8, - both: 0, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_single), - OrderingFlagsCounts { - first: 0, - last: 0, - both: 2, - neither: 0, - }, - ); - let mut read_names: HashMap>> = HashMap::new(); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let result = predict(ordering_flags, read_names, 0.0, false); - assert!(result.is_ok()); - let result = result.unwrap(); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 8); - assert_eq!(result.last, 8); - assert_eq!(result.both, 2); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, Some(2.2)); - assert_eq!(result.read_groups.len(), 2); - // We can't know which read group will be first in the vector. - // But both should succeed. - assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); - } -} +// #[cfg(test)] +// mod tests { +// use super::*; + +// #[test] +// fn test_predict_endedness() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict_endedness( +// "overall".to_string(), +// &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), +// 0.0, +// None, +// false, +// ); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// } + +// #[test] +// fn test_derive_endedness_from_all_zero_counts() { +// let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert("rg1", OrderingFlagsCounts::new()); +// let result = predict_endedness( +// "rg1".to_string(), +// &ordering_flags.get("rg1").unwrap(), +// 0.0, +// None, +// false, +// ); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// } + +// #[test] +// fn test_derive_endedness_from_only_first() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 0, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_both() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 1, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Single-End"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 1); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_neither() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 0, +// neither: 1, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 1); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_calculate_reads_per_template() { +// let mut read_names: HashMap>> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let results = calculate_reads_per_template(read_names); +// assert_eq!(results.len(), 3); +// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); +// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); +// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last_with_rpt() { +// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 2, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_paired), +// OrderingFlagsCounts { +// first: 8, +// last: 8, +// both: 0, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_single), +// OrderingFlagsCounts { +// first: 0, +// last: 0, +// both: 2, +// neither: 0, +// }, +// ); +// let mut read_names: HashMap>> = HashMap::new(); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let result = predict(ordering_flags, read_names, 0.0, false); +// assert!(result.is_ok()); +// let result = result.unwrap(); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 8); +// assert_eq!(result.last, 8); +// assert_eq!(result.both, 2); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, Some(2.2)); +// assert_eq!(result.read_groups.len(), 2); +// // We can't know which read group will be first in the vector. +// // But both should succeed. +// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); +// } +// } diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 883d95a..183a4de 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -47,7 +47,7 @@ pub fn predict( num_samples: usize, majority_vote_cutoff: f64, ) -> Result { - if num_samples <= 0 { + if num_samples == 0 { bail!("No read lengths were detected in the file."); } From 81511e25447f93b280cf9f53216049952a1fd67c Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 15 Dec 2023 15:51:47 -0500 Subject: [PATCH 28/91] tests(derive/endedness): reimplement tests --- src/derive/endedness/compute.rs | 566 ++++++++++++++++---------------- 1 file changed, 276 insertions(+), 290 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 0a53a36..c92eee2 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -406,293 +406,279 @@ pub fn predict( final_result } -// #[cfg(test)] -// mod tests { -// use super::*; - -// #[test] -// fn test_predict_endedness() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict_endedness( -// "overall".to_string(), -// &ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), -// 0.0, -// None, -// false, -// ); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// } - -// #[test] -// fn test_derive_endedness_from_all_zero_counts() { -// let mut ordering_flags: HashMap<&str, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert("rg1", OrderingFlagsCounts::new()); -// let result = predict_endedness( -// "rg1".to_string(), -// &ordering_flags.get("rg1").unwrap(), -// 0.0, -// None, -// false, -// ); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// } - -// #[test] -// fn test_derive_endedness_from_only_first() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 0, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_both() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 1, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Single-End"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 1); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_neither() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 0, -// neither: 1, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 1); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_calculate_reads_per_template() { -// let mut read_names: HashMap>> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let results = calculate_reads_per_template(read_names); -// assert_eq!(results.len(), 3); -// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); -// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); -// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last_with_rpt() { -// let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 2, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_paired), -// OrderingFlagsCounts { -// first: 8, -// last: 8, -// both: 0, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_single), -// OrderingFlagsCounts { -// first: 0, -// last: 0, -// both: 2, -// neither: 0, -// }, -// ); -// let mut read_names: HashMap>> = HashMap::new(); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let result = predict(ordering_flags, read_names, 0.0, false); -// assert!(result.is_ok()); -// let result = result.unwrap(); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 8); -// assert_eq!(result.last, 8); -// assert_eq!(result.both, 2); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, Some(2.2)); -// assert_eq!(result.read_groups.len(), 2); -// // We can't know which read group will be first in the vector. -// // But both should succeed. -// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); -// } -// } +#[cfg(test)] +mod tests { + use super::*; + + #[test] + fn test_predict_endedness() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict_endedness( + "overall".to_string(), + ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), + 0.0, + None, + false, + ); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_all_zero_counts() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); + let result = predict_endedness( + String::from("rg1"), + ordering_flags.get(&Arc::new(String::from("rg1"))).unwrap(), + 0.0, + None, + false, + ); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_only_first() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_both() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 1, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.succeeded); + assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 1); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_only_neither() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 0, + neither: 1, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 1); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_derive_endedness_from_first_and_last() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 0); + } + + #[test] + fn test_calculate_reads_per_template() { + let mut read_names: HashMap>> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let results = calculate_reads_per_template(read_names); + assert_eq!(results.len(), 3); + assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); + assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); + assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); + } + + #[test] + fn test_derive_endedness_from_first_and_last_with_rpt() { + let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + ordering_flags.insert( + Arc::clone(&OVERALL), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 2, + neither: 0, + }, + ); + ordering_flags.insert( + Arc::clone(&rg_paired), + OrderingFlagsCounts { + first: 8, + last: 8, + both: 0, + neither: 0, + }, + ); + ordering_flags.insert( + Arc::clone(&rg_single), + OrderingFlagsCounts { + first: 0, + last: 0, + both: 2, + neither: 0, + }, + ); + let mut read_names: HashMap>> = HashMap::new(); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let result = predict(ordering_flags, read_names, 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 8); + assert_eq!(result.last, 8); + assert_eq!(result.both, 2); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, Some(2.2)); + assert_eq!(result.read_groups.len(), 2); + // We can't know which read group will be first in the vector. + // But both should succeed. + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); + } +} From 55ee258c099f9d7960e28f5f4a130834ccc385b6 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 16 Dec 2023 12:39:15 -0500 Subject: [PATCH 29/91] chore(derive): disable index checking when not needed --- src/derive/command/endedness.rs | 2 +- src/derive/command/instrument.rs | 2 +- src/derive/command/readlen.rs | 2 +- 3 files changed, 3 insertions(+), 3 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 239dab8..766fc5c 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -74,7 +74,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index c36ec19..65ac0a0 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -56,7 +56,7 @@ async fn app(src: PathBuf, first_n_reads: Option) -> anyhow::Result<()> { let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(src, IndexCheck::Full)?; + } = crate::utils::formats::bam::open_and_parse(src, IndexCheck::None)?; // (1) Collect instrument names and flowcell names from reads within the // file. Support for sampling only a portion of the reads is provided. diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 64d18bd..5bce4cc 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -50,7 +50,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let ParsedBAMFile { mut reader, header, .. - } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::Full)?; + } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. From 35ccc3fef4c345478be7696626a680faa2a3ee77 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 16 Dec 2023 12:45:36 -0500 Subject: [PATCH 30/91] chore:(derive/readlen): return an anyhow::Ok instead of plain Ok --- src/derive/command/readlen.rs | 2 +- src/derive/readlen/compute.rs | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 5bce4cc..92bf47c 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -82,5 +82,5 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let output = serde_json::to_string_pretty(&result).unwrap(); print!("{}", output); - Ok(()) + anyhow::Ok(()) } diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 183a4de..0e10c56 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -69,7 +69,7 @@ pub fn predict( result.consensus_read_length = Some(consensus_read_length); } - Ok(result) + anyhow::Ok(result) } #[cfg(test)] From e8c481649d5504b112f1f22832aed8cadde0ec06 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 16 Dec 2023 12:49:17 -0500 Subject: [PATCH 31/91] docs(derive/readlen): correction in module name --- src/derive/readlen.rs | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/src/derive/readlen.rs b/src/derive/readlen.rs index d6f220e..b988896 100644 --- a/src/derive/readlen.rs +++ b/src/derive/readlen.rs @@ -1,3 +1,3 @@ -//! Supporting functionality for the `ngs derive instrument` subcommand. +//! Supporting functionality for the `ngs derive readlen` subcommand. pub mod compute; From 7e1578b1b9c936e0e43c397b2a22f15efddcbe3b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 16 Dec 2023 12:51:49 -0500 Subject: [PATCH 32/91] docs(derive/endedness): fix docs referring to wrong subcommand --- src/derive/command/endedness.rs | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 766fc5c..30dde04 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -76,7 +76,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { mut reader, header, .. } = crate::utils::formats::bam::open_and_parse(args.src, IndexCheck::None)?; - // (1) Collect read lengths from reads within the + // (1) Collect ordering flags (and QNAMEs) from reads within the // file. Support for sampling only a portion of the reads is provided. let mut samples = 0; let mut sample_max = 0; @@ -213,7 +213,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { ordering_flags.insert(Arc::new(rg_id), OrderingFlagsCounts::new()); } - // (2) Derive the consensus endedness based on the ordering flags gathered. + // (2) Derive the endedness based on the ordering flags gathered. let result = compute::predict( ordering_flags, read_names, From eab7cf9b6d117e3f7455f1fe0e74b43d6ec28601 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 18 Dec 2023 14:04:10 -0500 Subject: [PATCH 33/91] fix: cap QNAME warnings to 100 QNAMES --- src/derive/endedness/compute.rs | 27 +++++++++++++++++++++++---- 1 file changed, 23 insertions(+), 4 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index c92eee2..ec5439b 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -204,6 +204,8 @@ fn calculate_reads_per_template( let mut read_group_reads: HashMap, usize> = HashMap::new(); let mut read_group_templates: HashMap, usize> = HashMap::new(); + let mut warning_count: usize = 0; + for (read_name, read_groups) in read_names.iter() { let num_reads = read_groups.len(); total_reads += num_reads; @@ -223,10 +225,20 @@ fn calculate_reads_per_template( .and_modify(|e| *e += 1) .or_insert(1); } else { - warn!( - "QNAME: '{}' is in multiple read groups: {:?}", - read_name, read_group_set - ); + warning_count += 1; + match warning_count { + 1..=100 => { + warn!( + "QNAME: '{}' is in multiple read groups: {:?}", + read_name, read_group_set + ); + } + 101 => warn!( + "Too many warnings about QNAMEs in multiple read groups. Stopping warnings." + ), + _ => (), + } + for read_group in read_groups { read_group_reads .entry(Arc::clone(read_group)) @@ -242,6 +254,13 @@ fn calculate_reads_per_template( } } + if warning_count > 100 { + warn!( + "{} QNAMEs were found in multiple read groups.", + warning_count + ); + } + reads_per_template.insert( Arc::clone(&OVERALL), total_reads as f64 / total_templates as f64, From 019d61d47311a86e1d6690e7993c745882f4c89e Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 20 Dec 2023 10:05:34 -0500 Subject: [PATCH 34/91] feat(src/derive): use NumberOfRecords and RecordCounter structs --- src/derive/command/endedness.rs | 25 ++++++++++++++----------- src/derive/command/readlen.rs | 29 +++++++++++++++++++---------- 2 files changed, 33 insertions(+), 21 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 30dde04..e84b2ef 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -7,6 +7,8 @@ use std::sync::Arc; use clap::Args; use noodles::sam::record::data::field::Tag; +use num_format::Locale; +use num_format::ToFormattedString; use tracing::info; use tracing::trace; @@ -14,6 +16,8 @@ use crate::derive::endedness::compute; use crate::derive::endedness::compute::{ validate_read_group_info, OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP, }; +use crate::utils::args::NumberOfRecords; +use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -78,12 +82,8 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (1) Collect ordering flags (and QNAMEs) from reads within the // file. Support for sampling only a portion of the reads is provided. - let mut samples = 0; - let mut sample_max = 0; - - if let Some(s) = args.num_records { - sample_max = s; - } + let num_records = NumberOfRecords::from(args.num_records); + let mut counter = RecordCounter::new(); for result in reader.records(&header.parsed) { let record = result?; @@ -199,14 +199,17 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { unreachable!(); } - if sample_max > 0 { - samples += 1; - if samples > sample_max { - break; - } + counter.inc(); + if counter.time_to_break(&num_records) { + break; } } + info!( + "Processed {} records.", + counter.get().to_formatted_string(&Locale::en) + ); + // (1.5) Validate the read group information. let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); for rg_id in rgs_in_header_not_records { diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 92bf47c..b10c85e 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -4,9 +4,13 @@ use std::collections::HashMap; use std::path::PathBuf; use clap::Args; +use num_format::Locale; +use num_format::ToFormattedString; use tracing::info; use crate::derive::readlen::compute; +use crate::utils::args::NumberOfRecords; +use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -54,12 +58,8 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. - let mut samples = 0; - let mut sample_max = 0; - - if let Some(s) = args.num_records { - sample_max = s; - } + let num_records = NumberOfRecords::from(args.num_records); + let mut counter = RecordCounter::new(); for result in reader.records(&header.parsed) { let record = result?; @@ -67,15 +67,24 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { read_lengths.entry(len).and_modify(|e| *e += 1).or_insert(1); - samples += 1; - if sample_max > 0 && samples > sample_max { + counter.inc(); + if counter.time_to_break(&num_records) { break; } } + info!( + "Processed {} records.", + counter.get().to_formatted_string(&Locale::en) + ); + // (2) Derive the consensus read length based on the read lengths gathered. - let result = - compute::predict(read_lengths, samples, args.majority_vote_cutoff.unwrap()).unwrap(); + let result = compute::predict( + read_lengths, + counter.get(), + args.majority_vote_cutoff.unwrap(), + ) + .unwrap(); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). From dab1dcaac7100a61e5f48a918be108288a5a1e63 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 20 Dec 2023 15:15:46 -0500 Subject: [PATCH 35/91] style: make arg_in_range() nicer everywhere its used --- src/derive/command/endedness.rs | 31 +++++++++---------------------- src/derive/command/readlen.rs | 33 +++++++++------------------------ src/derive/endedness/compute.rs | 6 +++--- src/derive/readlen/compute.rs | 8 ++++---- src/generate/command.rs | 20 +++++--------------- src/utils/args.rs | 17 +++++++++++++++++ 6 files changed, 47 insertions(+), 68 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index e84b2ef..7a87136 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -5,6 +5,7 @@ use std::collections::HashSet; use std::path::PathBuf; use std::sync::Arc; +use anyhow::Context; use clap::Args; use noodles::sam::record::data::field::Tag; use num_format::Locale; @@ -16,24 +17,12 @@ use crate::derive::endedness::compute; use crate::derive::endedness::compute::{ validate_read_group_info, OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP, }; +use crate::utils::args::arg_in_range as deviance_in_range; use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; -/// Utility method to parse the Paired Deviance passed in on the command line and -/// ensure the value is within the range [0.0, 0.5]. -pub fn deviance_in_range(deviance_raw: &str) -> Result { - let deviance: f64 = deviance_raw - .parse() - .map_err(|_| format!("{} isn't a float", deviance_raw))?; - - match (0.0..=0.5).contains(&deviance) { - true => Ok(deviance), - false => Err(String::from("Paired Deviance must be between 0.0 and 0.5")), - } -} - /// Clap arguments for the `ngs derive endedness` subcommand. #[derive(Args)] pub struct DeriveEndednessArgs { @@ -48,9 +37,8 @@ pub struct DeriveEndednessArgs { /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate /// if the whole file is being processed. - #[arg(long, value_name = "F64", default_value = "0.0")] - #[arg(value_parser = deviance_in_range)] - paired_deviance: Option, + #[arg(long, value_name = "F32", default_value = "0.0")] + paired_deviance: f32, /// Calculate and output Reads-Per-Template. This will produce a more /// sophisticated estimate for endedness, but uses substantially more memory. @@ -65,6 +53,10 @@ pub struct DeriveEndednessArgs { /// Main function for the `ngs derive endedness` subcommand. pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { + // (0) Parse arguments needed for subcommand. + let paired_deviance = deviance_in_range(args.paired_deviance, 0.0..=0.5) + .with_context(|| "Paired deviance is not within acceptable range")?; + info!("Starting derive endedness subcommand."); let mut found_rgs = HashSet::new(); @@ -217,12 +209,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } // (2) Derive the endedness based on the ordering flags gathered. - let result = compute::predict( - ordering_flags, - read_names, - args.paired_deviance.unwrap(), - args.round_rpt, - ); + let result = compute::predict(ordering_flags, read_names, paired_deviance, args.round_rpt); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index b10c85e..52bf1f0 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -3,32 +3,19 @@ use std::collections::HashMap; use std::path::PathBuf; +use anyhow::Context; use clap::Args; use num_format::Locale; use num_format::ToFormattedString; use tracing::info; use crate::derive::readlen::compute; +use crate::utils::args::arg_in_range as cutoff_in_range; use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; -/// Utility method to parse the Majority Vote Cutoff passed in on the command line and -/// ensure the cutoff is within the range [0.0, 1.0]. -pub fn cutoff_in_range(cutoff_raw: &str) -> Result { - let cutoff: f64 = cutoff_raw - .parse() - .map_err(|_| format!("{} isn't a float", cutoff_raw))?; - - match (0.0..=1.0).contains(&cutoff) { - true => Ok(cutoff), - false => Err(String::from( - "Majority Vote Cutoff must be between 0.0 and 1.0", - )), - } -} - /// Clap arguments for the `ngs derive readlen` subcommand. #[derive(Args)] pub struct DeriveReadlenArgs { @@ -41,13 +28,16 @@ pub struct DeriveReadlenArgs { num_records: Option, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. - #[arg(short, long, value_name = "F64", default_value = "0.7")] - #[arg(value_parser = cutoff_in_range)] - majority_vote_cutoff: Option, + #[arg(short, long, value_name = "F32", default_value = "0.7")] + majority_vote_cutoff: f32, } /// Main function for the `ngs derive readlen` subcommand. pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { + // (0) Parse arguments needed for subcommand. + let majority_vote_cutoff = cutoff_in_range(args.majority_vote_cutoff, 0.0..=1.0) + .with_context(|| "Majority vote cutoff is not within acceptable range")?; + let mut read_lengths = HashMap::new(); info!("Starting derive readlen subcommand."); @@ -79,12 +69,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { ); // (2) Derive the consensus read length based on the read lengths gathered. - let result = compute::predict( - read_lengths, - counter.get(), - args.majority_vote_cutoff.unwrap(), - ) - .unwrap(); + let result = compute::predict(read_lengths, counter.get(), majority_vote_cutoff).unwrap(); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index ec5439b..5990683 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -278,7 +278,7 @@ fn calculate_reads_per_template( fn predict_endedness( read_group_name: String, rg_ordering_flags: &OrderingFlagsCounts, - paired_deviance: f64, + paired_deviance: f32, reads_per_template: Option<&f64>, round_rpt: bool, ) -> ReadGroupDerivedEndednessResult { @@ -351,7 +351,7 @@ fn predict_endedness( // both and neither are now guarenteed to be 0 // We only need to check first and last - let first_frac = first as f64 / (first + last) as f64; + let first_frac = first as f32 / (first + last) as f32; let lower_limit = 0.5 - paired_deviance; let upper_limit = 0.5 + paired_deviance; if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { @@ -377,7 +377,7 @@ fn predict_endedness( pub fn predict( ordering_flags: HashMap, OrderingFlagsCounts>, read_names: HashMap>>, - paired_deviance: f64, + paired_deviance: f32, round_rpt: bool, ) -> DerivedEndednessResult { let mut rpts: HashMap, f64> = HashMap::new(); diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 0e10c56..9f3bbf7 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -15,7 +15,7 @@ pub struct DerivedReadlenResult { pub consensus_read_length: Option, /// The majority vote percentage of the consensus read length, if available. - pub majority_pct_detected: f64, + pub majority_pct_detected: f32, /// Status of the evidence that supports (or does not support) this /// read length, if available. @@ -27,7 +27,7 @@ impl DerivedReadlenResult { pub fn new( succeeded: bool, consensus_read_length: Option, - majority_pct_detected: f64, + majority_pct_detected: f32, evidence: Vec<(usize, usize)>, ) -> Self { DerivedReadlenResult { @@ -45,7 +45,7 @@ impl DerivedReadlenResult { pub fn predict( read_lengths: HashMap, num_samples: usize, - majority_vote_cutoff: f64, + majority_vote_cutoff: f32, ) -> Result { if num_samples == 0 { bail!("No read lengths were detected in the file."); @@ -59,7 +59,7 @@ pub fn predict( let max_count = read_lengths[0].1; let consensus_read_length = max_read_length; - let majority_detected = max_count as f64 / num_samples as f64; + let majority_detected = max_count as f32 / num_samples as f32; let mut result = DerivedReadlenResult::new(false, None, majority_detected * 100.0, read_lengths); diff --git a/src/generate/command.rs b/src/generate/command.rs index a6acc77..167e9fd 100644 --- a/src/generate/command.rs +++ b/src/generate/command.rs @@ -12,21 +12,9 @@ use tracing::info; use crate::generate::providers::reference_provider::ReferenceGenomeSequenceProvider; use crate::generate::providers::SequenceProvider; +use crate::utils::args::arg_in_range as error_rate_in_range; use crate::utils::formats; -/// Utility method to parse the error rate passed in on the command line and -/// ensure the rate is within the range [0.0, 1.0]. -pub fn error_rate_in_range(error_rate_raw: &str) -> Result { - let error_rate: f32 = error_rate_raw - .parse() - .map_err(|_| format!("{} isn't a float", error_rate_raw))?; - - match (0.0..=1.0).contains(&error_rate) { - true => Ok(error_rate), - false => Err(String::from("Error rate must be between 0.0 and 1.0")), - } -} - /// Command line arguments for `ngs generate`. #[derive(Args)] #[command(group(ArgGroup::new("record-count").required(true).args(["coverage", "num_records"])))] @@ -43,8 +31,7 @@ pub struct GenerateArgs { /// The error rate for the sequencer as a fraction between [0.0, 1.0] (per base). #[arg(short, long, value_name = "F32", default_value = "0.0001")] - #[arg(value_parser = error_rate_in_range)] - error_rate: Option, + error_rate: f32, /// Specifies the number of records to generate. #[arg(short, long, value_name = "USIZE", conflicts_with = "coverage")] @@ -58,6 +45,9 @@ pub struct GenerateArgs { /// Main function for the `ngs generate` subcommand. pub fn generate(args: GenerateArgs) -> anyhow::Result<()> { // (0) Parse arguments needed for subcommand. + let _error_rate = error_rate_in_range(args.error_rate, 0.0..=1.0) + .with_context(|| "Error rate is not within acceptable range")?; + let result: anyhow::Result> = args .reference_providers .iter() diff --git a/src/utils/args.rs b/src/utils/args.rs index e576400..4151cb7 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -73,3 +73,20 @@ impl From for CompressionLevel { } } } + +//==============// +// Float Parser // +//==============// + +/// Utility method to parse command line floats and ensure they are +/// within the range [MIN, MAX]. +pub fn arg_in_range(arg: f32, range: std::ops::RangeInclusive) -> anyhow::Result { + match range.contains(&arg) { + true => Ok(arg), + false => anyhow::bail!( + "Value must be between {} and {}", + range.start(), + range.end() + ), + } +} From 489c2bbdd197a4db884da0fc651f268138888bf7 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 22 Dec 2023 15:57:11 -0500 Subject: [PATCH 36/91] [WIP]: junction annotation --- src/derive.rs | 1 + src/derive/command.rs | 7 + src/derive/command/junction_annotation.rs | 137 ++++++++ src/derive/junction_annotation.rs | 4 + src/derive/junction_annotation/compute.rs | 371 ++++++++++++++++++++++ src/derive/junction_annotation/results.rs | 114 +++++++ src/main.rs | 3 + 7 files changed, 637 insertions(+) create mode 100644 src/derive/command/junction_annotation.rs create mode 100644 src/derive/junction_annotation.rs create mode 100644 src/derive/junction_annotation/compute.rs create mode 100644 src/derive/junction_annotation/results.rs diff --git a/src/derive.rs b/src/derive.rs index 5b45b41..104bfe3 100644 --- a/src/derive.rs +++ b/src/derive.rs @@ -4,4 +4,5 @@ pub mod command; pub mod encoding; pub mod endedness; pub mod instrument; +pub mod junction_annotation; pub mod readlen; diff --git a/src/derive/command.rs b/src/derive/command.rs index 5304207..5d4a593 100644 --- a/src/derive/command.rs +++ b/src/derive/command.rs @@ -3,6 +3,7 @@ pub mod encoding; pub mod endedness; pub mod instrument; +pub mod junction_annotation; pub mod readlen; use clap::Args; @@ -34,4 +35,10 @@ pub enum DeriveSubcommand { /// Derives the read length of the file. Readlen(self::readlen::DeriveReadlenArgs), + + /// Annotates junctions in the file. + /// This subcommand requires a GFF file with features to annotate. + /// This subcommand does not "derive" anything, but is included here for + /// convenience. + JunctionAnnotation(self::junction_annotation::JunctionAnnotationArgs), } diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs new file mode 100644 index 0000000..550d5d5 --- /dev/null +++ b/src/derive/command/junction_annotation.rs @@ -0,0 +1,137 @@ +//! Functionality relating to the `ngs derive junction_annotation` subcommand itself. + +use std::collections::HashMap; +use std::path::PathBuf; + +use anyhow::Context; +use clap::Args; +use num_format::Locale; +use num_format::ToFormattedString; +use tracing::debug; +use tracing::info; + +use crate::derive::junction_annotation::compute; +use crate::derive::junction_annotation::results::JunctionAnnotationResults; +use crate::utils::display::RecordCounter; +use crate::utils::formats; +use crate::utils::formats::bam::ParsedBAMFile; +use crate::utils::formats::utils::IndexCheck; + +/// Clap arguments for the `ngs derive junction_annotation` subcommand. +#[derive(Args)] +pub struct JunctionAnnotationArgs { + /// Source BAM. + #[arg(value_name = "BAM")] + src: PathBuf, + + /// Features GFF file. + #[arg(short = 'f', long, required = true, value_name = "PATH")] + features_gff: PathBuf, + + /// Name of the exon region feature for the gene model used. + #[arg(short, long, value_name = "STRING", default_value = "exon")] + pub exon_feature_name: String, + + /// Minimum intron length to consider. + /// An intron is defined as an `N` CIGAR operation of any length. + #[arg(long, value_name = "USIZE", default_value = "50")] + pub min_intron_length: usize, + + /// Add +- this amount to intron positions when looking up exon positions. + #[arg(long, value_name = "U8", default_value = "0")] + pub fuzzy_junction_match_range: u8, + + /// Minimum number of reads supporting a junction to be considered. + #[arg(long, value_name = "U8", default_value = "2")] + pub min_read_support: u8, + + /// Minumum mapping quality for a record to be considered. + #[arg(short, long, value_name = "U8", default_value = "30")] + pub min_mapq: u8, +} + +/// Main function for the `ngs derive junction_annotation` subcommand. +pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { + info!("Starting derive junction_annotation subcommand."); + + let mut exon_starts: HashMap<&str, Vec> = HashMap::new(); + let mut exon_ends: HashMap<&str, Vec> = HashMap::new(); + + // (1) Parse the GFF file and collect all exon features. + debug!("Reading all records in GFF."); + let mut gff = formats::gff::open(&args.features_gff) + .with_context(|| format!("opening GFF file: {}", args.features_gff.display()))?; + + let mut exon_records = Vec::new(); + for result in gff.records() { + let record = result.unwrap(); + if record.ty() != args.exon_feature_name { + continue; + } + exon_records.push(record); + } + + debug!("Tabulating GFF exon features."); + for record in &exon_records { + let seq_name = record.reference_sequence_name(); + let start = record.start().into(); + let end = record.end().into(); + + exon_starts.entry(seq_name).or_default().push(start); + exon_ends.entry(seq_name).or_default().push(end); + } + + debug!("Finalizing GFF features lookup."); + for starts in exon_starts.values_mut() { + starts.sort_unstable(); + starts.dedup(); + } + for ends in exon_ends.values_mut() { + ends.sort_unstable(); + ends.dedup(); + } + + debug!("Done reading GFF."); + + let mut counter = RecordCounter::new(); + let mut results = JunctionAnnotationResults::default(); + let params = compute::JunctionAnnotationParameters { + min_intron_length: args.min_intron_length, + fuzzy_junction_match_range: args.fuzzy_junction_match_range, + min_read_support: args.min_read_support, + min_mapq: args.min_mapq, + }; + + let ParsedBAMFile { + mut reader, header, .. + } = formats::bam::open_and_parse(args.src, IndexCheck::None)?; + + // (2) Process each record in the BAM file. + for result in reader.records(&header.parsed) { + let record = result?; + compute::process( + &record, + &exon_starts, + &exon_ends, + &header.parsed, + ¶ms, + &mut results, + )?; + counter.inc(); + } + + info!( + "Processed {} records.", + counter.get().to_formatted_string(&Locale::en) + ); + + // (3) Summarize found junctions. + compute::summarize(&mut results, ¶ms)?; + + // (3) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&results).unwrap(); + print!("{}", output); + + Ok(()) +} diff --git a/src/derive/junction_annotation.rs b/src/derive/junction_annotation.rs new file mode 100644 index 0000000..df1f81a --- /dev/null +++ b/src/derive/junction_annotation.rs @@ -0,0 +1,4 @@ +//! Supporting functionality for the `ngs derive junction_annotation` subcommand. + +pub mod compute; +pub mod results; diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs new file mode 100644 index 0000000..dc77280 --- /dev/null +++ b/src/derive/junction_annotation/compute.rs @@ -0,0 +1,371 @@ +//! Module holding the logic for annotating junctions. + +use anyhow::bail; +use noodles::sam::alignment::Record; +use noodles::sam::record::cigar::op::Kind; +use noodles::sam::Header; +use std::collections::HashMap; +use std::num::NonZeroUsize; + +use crate::derive::junction_annotation::results::JunctionAnnotationResults; + +/// Parameters defining how to annotate found junctions +pub struct JunctionAnnotationParameters { + /// Minimum intron length to consider. + pub min_intron_length: usize, + + /// Add +- this amount to intron positions when looking up exon positions. + pub fuzzy_junction_match_range: u8, + + /// Minimum number of reads supporting a junction to be considered. + pub min_read_support: u8, + + /// Minumum mapping quality for a record to be considered. + pub min_mapq: u8, +} + +/// Main function to annotate junctions one record at a time. +pub fn process( + record: &Record, + exon_starts: &HashMap<&str, Vec>, + exon_ends: &HashMap<&str, Vec>, + header: &Header, + params: &JunctionAnnotationParameters, + results: &mut JunctionAnnotationResults, +) -> anyhow::Result<()> { + // (1) Parse the read name. + let read_name = match record.read_name() { + Some(name) => name, + _ => bail!("Could not parse read name"), + }; + + // (2) Parse the flags so we can see if the read is mapped. + let flags = record.flags(); + + // (3) If the read is unmapped, just return—no need to throw an error. + if flags.is_unmapped() { + results.records.ignored_flags += 1; + return Ok(()); + } + + // (4) Parse the CIGAR string from the record. + // We only care about reads with introns, so if there are no introns + // we can skip this read. + let cigar = record.cigar(); + if !cigar.iter().any(|op| matches!(op.kind(), Kind::Skip)) { + results.records.not_spliced += 1; + return Ok(()); + } + + // (5) If the read has a MAPQ below our threshold, just return. + // No need to throw an error, unless the MAPQ could not be parsed. + match record.mapping_quality() { + Some(mapq) => { + if mapq.get() < params.min_mapq { + results.records.low_mapq += 1; + return Ok(()); + } + } + _ => results.records.couldnt_parse += 1, + } + + // (6) Parse the reference sequence id from the record. + let id = match record.reference_sequence_id() { + Some(id) => id, + _ => { + bail!( + "Could not parse reference sequence id for read: {}", + read_name + ) + } + }; + + // (7) Map the parsed reference sequence id to a reference sequence name. + let seq_name = match header + .reference_sequences() + .get_index(id) + .map(|(name, _)| Some(name)) + { + Some(Some(name)) => name.as_str(), + _ => { + bail!( + "Could not map reference sequence id to header for read: {}", + read_name + ) + } + }; + + // (8) Check if there will be annotations for this reference sequence. + let mut ref_is_annotated = true; + if !exon_starts.contains_key(&seq_name) || !exon_ends.contains_key(&seq_name) { + ref_is_annotated = false; + } + + // (9) Calculate the start position of this read. This will + // later be used to find the position of any introns. + let start = match record.alignment_start() { + Some(s) => usize::from(s), + _ => bail!("Could not parse record's start position."), + }; + + // (10) Find introns + let mut cur_pos = start; + for op in cigar.iter() { + match op.kind() { + // Operations that increment the reference position. + Kind::Match | Kind::Deletion | Kind::SequenceMatch | Kind::SequenceMismatch => { + cur_pos += op.len(); + } + // This is an intron. + Kind::Skip => { + // Do this check later, for better metric reporting. + // if op.len() < params.min_intron_length { + // continue; + // } + + let intron_start = cur_pos; + let intron_end = cur_pos + op.len(); + // Update cur_pos to the end of the intron + // in case there are multiple introns in the read. + cur_pos = intron_end; + + // If the reference sequence is not annotated, we can skip + // the lookup of exon positions, and directly insert the + // intron into the unannotated_reference HashMap. + if !ref_is_annotated { + results + .junction_annotations + .unannotated_reference + .entry(seq_name.to_string()) + .or_default() + .entry(( + NonZeroUsize::new(intron_start).unwrap(), + NonZeroUsize::new(intron_end).unwrap(), + )) + .and_modify(|e| *e += 1) + .or_insert(1); + continue; + } + + let exon_starts = match exon_starts.get(&seq_name) { + Some(starts) => starts, + _ => bail!("Could not find exon starts for contig: {}", seq_name), + }; + let exon_ends = match exon_ends.get(&seq_name) { + Some(ends) => ends, + _ => bail!("Could not find exon ends for contig: {}", seq_name), + }; + + let mut intron_start_known = false; + let mut intron_end_known = false; + // To allow collapsing fuzzy junctions, + // we need to store the reference positions of the exon boundaries. + // We initialize these values to the position of the found intron. + let mut ref_intron_start = intron_start; + let mut ref_intron_end = intron_end; + for exon_end in exon_ends.iter() { + if intron_start >= (exon_end - params.fuzzy_junction_match_range as usize) + && intron_start <= (exon_end + params.fuzzy_junction_match_range as usize) + { + intron_start_known = true; + ref_intron_start = *exon_end; + break; + } + } + for exon_start in exon_starts.iter() { + if intron_end >= (exon_start - params.fuzzy_junction_match_range as usize) + && intron_end <= (exon_start + params.fuzzy_junction_match_range as usize) + { + intron_end_known = true; + ref_intron_end = *exon_start; + break; + } + } + + match (intron_start_known, intron_end_known) { + (true, true) => { + // We found both ends of the intron. + // This is a Known Junction. + results + .junction_annotations + .known + .entry(seq_name.to_string()) + .or_default() + .entry(( + NonZeroUsize::new(ref_intron_start).unwrap(), + NonZeroUsize::new(ref_intron_end).unwrap(), + )) + .and_modify(|e| *e += 1) + .or_insert(1); + } + (true, false) | (false, true) => { + // We found one end of the intron, + // but not the other. + // This is a Partial Novel Junction. + results + .junction_annotations + .partial_novel + .entry(seq_name.to_string()) + .or_default() + .entry(( + NonZeroUsize::new(ref_intron_start).unwrap(), + NonZeroUsize::new(ref_intron_end).unwrap(), + )) + .and_modify(|e| *e += 1) + .or_insert(1); + } + (false, false) => { + // We found neither end of the intron. + // This is a Complete Novel Junction. + results + .junction_annotations + .complete_novel + .entry(seq_name.to_string()) + .or_default() + .entry(( + NonZeroUsize::new(ref_intron_start).unwrap(), + NonZeroUsize::new(ref_intron_end).unwrap(), + )) + .and_modify(|e| *e += 1) + .or_insert(1); + } + } + } + // Operations that do not increment the reference position. + _ => {} + } + } + + results.records.processed += 1; + Ok(()) +} + +/// Main function to summarize the results of the junction_annotation subcommand. +pub fn summarize( + results: &mut JunctionAnnotationResults, + params: &JunctionAnnotationParameters, +) -> anyhow::Result<()> { + // Filter out junctions that are too short or don't have enough read support. + let mut num_junctions_too_short: usize = 0; + let mut num_not_enough_support: usize = 0; + for (_, v) in results.junction_annotations.known.iter_mut() { + v.retain(|(start, end), count| { + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + false + } else if *count < params.min_read_support as usize { + num_not_enough_support += 1; + false + } else { + true + } + }); + } + for (_, v) in results.junction_annotations.partial_novel.iter_mut() { + v.retain(|(start, end), count| { + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + false + } else if *count < params.min_read_support as usize { + num_not_enough_support += 1; + false + } else { + true + } + }); + } + for (_, v) in results.junction_annotations.complete_novel.iter_mut() { + v.retain(|(start, end), count| { + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + false + } else if *count < params.min_read_support as usize { + num_not_enough_support += 1; + false + } else { + true + } + }); + } + for (_, v) in results + .junction_annotations + .unannotated_reference + .iter_mut() + { + v.retain(|(start, end), count| { + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + false + } else if *count < params.min_read_support as usize { + num_not_enough_support += 1; + false + } else { + true + } + }); + } + results.summary.intron_too_short = num_junctions_too_short; + results.summary.junctions_with_not_enough_read_support = num_not_enough_support; + + // Tally up observed junctions and spliced reads. + results.summary.known_junctions = results + .junction_annotations + .known + .values() + .map(|v| v.len()) + .sum(); + results.summary.known_spliced_reads = results + .junction_annotations + .known + .values() + .map(|v| v.values().sum::()) + .sum(); + results.summary.partial_novel_junctions = results + .junction_annotations + .partial_novel + .values() + .map(|v| v.len()) + .sum(); + results.summary.partial_novel_spliced_reads = results + .junction_annotations + .partial_novel + .values() + .map(|v| v.values().sum::()) + .sum(); + results.summary.complete_novel_junctions = results + .junction_annotations + .complete_novel + .values() + .map(|v| v.len()) + .sum(); + results.summary.complete_novel_spliced_reads = results + .junction_annotations + .complete_novel + .values() + .map(|v| v.values().sum::()) + .sum(); + results.summary.unannotated_reference_junctions = results + .junction_annotations + .unannotated_reference + .values() + .map(|v| v.len()) + .sum(); + results.summary.unannotated_reference_spliced_reads = results + .junction_annotations + .unannotated_reference + .values() + .map(|v| v.values().sum::()) + .sum(); + + // Tally up total junctions and spliced reads. + results.summary.total_junctions = results.summary.known_junctions + + results.summary.partial_novel_junctions + + results.summary.complete_novel_junctions + + results.summary.unannotated_reference_junctions; + results.summary.total_spliced_reads = results.summary.known_spliced_reads + + results.summary.partial_novel_spliced_reads + + results.summary.complete_novel_spliced_reads + + results.summary.unannotated_reference_spliced_reads; + + Ok(()) +} diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs new file mode 100644 index 0000000..5670dd1 --- /dev/null +++ b/src/derive/junction_annotation/results.rs @@ -0,0 +1,114 @@ +//! Results related to the `ngs derive junction_annotation` subcommand. + +use serde::Deserialize; +use serde::Serialize; +use std::collections::HashMap; +use std::num::NonZeroUsize; + +/// Lists of annotated junctions. +#[derive(Clone, Default, Serialize, Deserialize)] +pub struct JunctionAnnotations { + /// Known junctions. The outer key is the referece name, and the value is another + /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// and the value is the number of spliced reads that support the junction. + pub known: HashMap>, + + /// Partially novel junctions. The outer key is the referece name, and the value is another + /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// and the value is the number of spliced reads that support the junction. + pub partial_novel: HashMap>, + + /// Complete novel junctions. The outer key is the referece name, and the value is another + /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// and the value is the number of spliced reads that support the junction. + pub complete_novel: HashMap>, + + /// Junctions on reference sequences for which junction annotations were not found. + /// The outer key is the referece name, and the value is another + /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// and the value is the number of spliced reads that support the junction. + pub unannotated_reference: HashMap>, +} + +/// General record metrics that are tallied as a part of the +/// junction_annotation subcommand. +#[derive(Clone, Default, Serialize, Deserialize)] +pub struct RecordMetrics { + /// The number of records that have been fully processed. + /// Should equal Metrics::SummaryMetrics::total_spliced_reads. + pub processed: usize, + + /// The number of records that couldn't be parsed. + pub couldnt_parse: usize, + + /// The number of records that have been ignored because of their flags. + /// (i.e. they were unmapped, duplicates, secondary, or supplementary) + /// The last 3 conditions can be toggled on/off with CL flags + pub ignored_flags: usize, + + /// The number of records that have been ignored because they were not + /// spliced. + pub not_spliced: usize, + + /// The number of records with junctions that have been ignored because + /// they failed the MAPQ filter. + pub low_mapq: usize, +} + +/// Summary statistics for the junction_annotation subcommand. +#[derive(Clone, Default, Serialize, Deserialize)] +pub struct SummaryResults { + /// The total number of junctions observed in the file. + pub total_junctions: usize, + + /// The total number of spliced reads observed in the file. + pub total_spliced_reads: usize, + + /// The total number of known junctions observed in the file. + pub known_junctions: usize, + + ///The total number of partially novel junctions observed in the file. + pub partial_novel_junctions: usize, + + /// The total number of complete novel junctions observed in the file. + pub complete_novel_junctions: usize, + + /// The total number of junctions on reference sequences for which junction + /// annotations were not found. + pub unannotated_reference_junctions: usize, + + /// The total number of known spliced reads observed in the file. + pub known_spliced_reads: usize, + + /// The total number of partially novel spliced reads observed in the file. + pub partial_novel_spliced_reads: usize, + + /// The total number of complete novel spliced reads observed in the file. + pub complete_novel_spliced_reads: usize, + + /// The total number of spliced reads on reference sequences for which + /// junction annotations were not found. + pub unannotated_reference_spliced_reads: usize, + + /// The total number of junctions which were discarded due to lack of + /// read support. + pub junctions_with_not_enough_read_support: usize, + + /// The number of junctions that have been ignored because + /// they failed the min_intron_length filter. + pub intron_too_short: usize, +} + +/// Main Results struct. This struct aggregates all of the minor metrics structs +/// outlined in this file so they can be kept track of as a unit. +#[derive(Clone, Default, Serialize, Deserialize)] +pub struct JunctionAnnotationResults { + /// Lists of annotated junctions. + pub junction_annotations: JunctionAnnotations, + + /// General record metrics. + pub records: RecordMetrics, + + /// Summary statistics for the junction_annotation subcommand. + pub summary: SummaryResults, +} diff --git a/src/main.rs b/src/main.rs index 6d0e557..b2312e1 100644 --- a/src/main.rs +++ b/src/main.rs @@ -101,6 +101,9 @@ fn main() -> anyhow::Result<()> { derive::command::DeriveSubcommand::Readlen(args) => { derive::command::readlen::derive(args)? } + derive::command::DeriveSubcommand::JunctionAnnotation(args) => { + derive::command::junction_annotation::derive(args)? + } }, Subcommands::Generate(args) => generate::command::generate(args)?, Subcommands::Index(args) => index::command::index(args)?, From 14db8136e4fe4d57489bd5bea6de437911003e44 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 26 Dec 2023 11:42:13 -0500 Subject: [PATCH 37/91] feat(derive): junction-annotation subcommand --- src/derive/command/junction_annotation.rs | 25 ++++++++-- src/derive/junction_annotation/compute.rs | 50 +++++++++++++------- src/derive/junction_annotation/results.rs | 57 +++++++++++++++++++---- 3 files changed, 103 insertions(+), 29 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 550d5d5..e16c5ac 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -29,7 +29,7 @@ pub struct JunctionAnnotationArgs { features_gff: PathBuf, /// Name of the exon region feature for the gene model used. - #[arg(short, long, value_name = "STRING", default_value = "exon")] + #[arg(long, value_name = "STRING", default_value = "exon")] pub exon_feature_name: String, /// Minimum intron length to consider. @@ -46,8 +46,22 @@ pub struct JunctionAnnotationArgs { pub min_read_support: u8, /// Minumum mapping quality for a record to be considered. + /// Set to 0 to disable this filter and allow reads _without_ + /// a mapping quality to be considered. #[arg(short, long, value_name = "U8", default_value = "30")] pub min_mapq: u8, + + /// Do not count supplementary alignments. + #[arg(long)] + pub no_supplementary: bool, + + /// Do count secondary alignments. + #[arg(long)] + pub count_secondary: bool, + + /// Do count duplicates. + #[arg(long)] + pub count_duplicates: bool, } /// Main function for the `ngs derive junction_annotation` subcommand. @@ -74,11 +88,11 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { debug!("Tabulating GFF exon features."); for record in &exon_records { let seq_name = record.reference_sequence_name(); - let start = record.start().into(); - let end = record.end().into(); + let start: usize = record.start().into(); + let end: usize = record.end().into(); exon_starts.entry(seq_name).or_default().push(start); - exon_ends.entry(seq_name).or_default().push(end); + exon_ends.entry(seq_name).or_default().push(end + 1); // TODO why +1? It works } debug!("Finalizing GFF features lookup."); @@ -100,6 +114,9 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { fuzzy_junction_match_range: args.fuzzy_junction_match_range, min_read_support: args.min_read_support, min_mapq: args.min_mapq, + no_supplementary: args.no_supplementary, + count_secondary: args.count_secondary, + count_duplicates: args.count_duplicates, }; let ParsedBAMFile { diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index dc77280..a76a072 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -21,7 +21,17 @@ pub struct JunctionAnnotationParameters { pub min_read_support: u8, /// Minumum mapping quality for a record to be considered. + /// 0 if MAPQ shouldn't be considered. pub min_mapq: u8, + + /// Do not count supplementary alignments. + pub no_supplementary: bool, + + /// Do count secondary alignments. + pub count_secondary: bool, + + /// Do count duplicates. + pub count_duplicates: bool, } /// Main function to annotate junctions one record at a time. @@ -39,16 +49,19 @@ pub fn process( _ => bail!("Could not parse read name"), }; - // (2) Parse the flags so we can see if the read is mapped. + // (2) Parse the flags so we can see if the read should be ignored. let flags = record.flags(); - // (3) If the read is unmapped, just return—no need to throw an error. - if flags.is_unmapped() { + if flags.is_unmapped() + || (params.no_supplementary && flags.is_supplementary()) + || (!params.count_secondary && flags.is_secondary()) + || (!params.count_duplicates && flags.is_duplicate()) + { results.records.ignored_flags += 1; return Ok(()); } - // (4) Parse the CIGAR string from the record. + // (3) Parse the CIGAR string from the record. // We only care about reads with introns, so if there are no introns // we can skip this read. let cigar = record.cigar(); @@ -57,19 +70,24 @@ pub fn process( return Ok(()); } - // (5) If the read has a MAPQ below our threshold, just return. - // No need to throw an error, unless the MAPQ could not be parsed. - match record.mapping_quality() { - Some(mapq) => { - if mapq.get() < params.min_mapq { - results.records.low_mapq += 1; + // (4) If the user is filtering by MAPQ, check if this read passes. + // Log if the read is filtered out for a too low MAPQ or a missing MAPQ. + if params.min_mapq > 0 { + match record.mapping_quality() { + Some(mapq) => { + if mapq.get() < params.min_mapq { + results.records.low_mapq += 1; + return Ok(()); + } + } + None => { + results.records.missing_mapq += 1; return Ok(()); } } - _ => results.records.couldnt_parse += 1, } - // (6) Parse the reference sequence id from the record. + // (5) Parse the reference sequence id from the record. let id = match record.reference_sequence_id() { Some(id) => id, _ => { @@ -80,7 +98,7 @@ pub fn process( } }; - // (7) Map the parsed reference sequence id to a reference sequence name. + // (6) Map the parsed reference sequence id to a reference sequence name. let seq_name = match header .reference_sequences() .get_index(id) @@ -95,20 +113,20 @@ pub fn process( } }; - // (8) Check if there will be annotations for this reference sequence. + // (7) Check if there will be annotations for this reference sequence. let mut ref_is_annotated = true; if !exon_starts.contains_key(&seq_name) || !exon_ends.contains_key(&seq_name) { ref_is_annotated = false; } - // (9) Calculate the start position of this read. This will + // (8) Calculate the start position of this read. This will // later be used to find the position of any introns. let start = match record.alignment_start() { Some(s) => usize::from(s), _ => bail!("Could not parse record's start position."), }; - // (10) Find introns + // (9) Find introns let mut cur_pos = start; for op in cigar.iter() { match op.kind() { diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 5670dd1..9c63a03 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -1,12 +1,13 @@ //! Results related to the `ngs derive junction_annotation` subcommand. -use serde::Deserialize; +use serde::ser::SerializeStruct; use serde::Serialize; +use serde::Serializer; use std::collections::HashMap; use std::num::NonZeroUsize; /// Lists of annotated junctions. -#[derive(Clone, Default, Serialize, Deserialize)] +#[derive(Clone, Default)] pub struct JunctionAnnotations { /// Known junctions. The outer key is the referece name, and the value is another /// HashMap. The inner key is the (start, end) coordinates of the junction, @@ -30,17 +31,52 @@ pub struct JunctionAnnotations { pub unannotated_reference: HashMap>, } +impl Serialize for JunctionAnnotations { + fn serialize(&self, serializer: S) -> Result { + let mut known = Vec::new(); + for (ref_name, junctions) in &self.known { + for ((start, end), count) in junctions { + known.push((ref_name, start, end, count)); + } + } + + let mut partial_novel = Vec::new(); + for (ref_name, junctions) in &self.partial_novel { + for ((start, end), count) in junctions { + partial_novel.push((ref_name, start, end, count)); + } + } + + let mut complete_novel = Vec::new(); + for (ref_name, junctions) in &self.complete_novel { + for ((start, end), count) in junctions { + complete_novel.push((ref_name, start, end, count)); + } + } + + let mut unannotated_reference = Vec::new(); + for (ref_name, junctions) in &self.unannotated_reference { + for ((start, end), count) in junctions { + unannotated_reference.push((ref_name, start, end, count)); + } + } + + let mut s = serializer.serialize_struct("JunctionAnnotations", 4)?; + s.serialize_field("known", &known)?; + s.serialize_field("partial_novel", &partial_novel)?; + s.serialize_field("complete_novel", &complete_novel)?; + s.serialize_field("unannotated_reference", &unannotated_reference)?; + s.end() + } +} + /// General record metrics that are tallied as a part of the /// junction_annotation subcommand. -#[derive(Clone, Default, Serialize, Deserialize)] +#[derive(Clone, Default, Serialize)] pub struct RecordMetrics { /// The number of records that have been fully processed. - /// Should equal Metrics::SummaryMetrics::total_spliced_reads. pub processed: usize, - /// The number of records that couldn't be parsed. - pub couldnt_parse: usize, - /// The number of records that have been ignored because of their flags. /// (i.e. they were unmapped, duplicates, secondary, or supplementary) /// The last 3 conditions can be toggled on/off with CL flags @@ -53,10 +89,13 @@ pub struct RecordMetrics { /// The number of records with junctions that have been ignored because /// they failed the MAPQ filter. pub low_mapq: usize, + + /// The number of records whose MAPQ couldn't be parsed and were thus ignored. + pub missing_mapq: usize, } /// Summary statistics for the junction_annotation subcommand. -#[derive(Clone, Default, Serialize, Deserialize)] +#[derive(Clone, Default, Serialize)] pub struct SummaryResults { /// The total number of junctions observed in the file. pub total_junctions: usize, @@ -101,7 +140,7 @@ pub struct SummaryResults { /// Main Results struct. This struct aggregates all of the minor metrics structs /// outlined in this file so they can be kept track of as a unit. -#[derive(Clone, Default, Serialize, Deserialize)] +#[derive(Clone, Default, Serialize)] pub struct JunctionAnnotationResults { /// Lists of annotated junctions. pub junction_annotations: JunctionAnnotations, From bf31648b7decd97493b0b8f25566c144584b4201 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 26 Dec 2023 12:04:50 -0500 Subject: [PATCH 38/91] chore: remove radix_trie dep --- Cargo.lock | 26 -------------------------- Cargo.toml | 1 - 2 files changed, 27 deletions(-) diff --git a/Cargo.lock b/Cargo.lock index 925f544..73568a3 100644 --- a/Cargo.lock +++ b/Cargo.lock @@ -438,12 +438,6 @@ version = "1.0.0" source = "registry+https://github.com/rust-lang/crates.io-index" checksum = "34aa73646ffb006b8f5147f3dc182bd4bcb190227ce861fc4a4844bf8e3cb2c0" -[[package]] -name = "endian-type" -version = "0.1.2" -source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "c34f04666d835ff5d62e058c3995147c06f42fe86ff053337632bca83e42702d" - [[package]] name = "erased-serde" version = "0.3.23" @@ -872,7 +866,6 @@ dependencies = [ "num-format", "plotly", "prettytable-rs", - "radix_trie", "rand", "rand_distr", "regex", @@ -884,15 +877,6 @@ dependencies = [ "tracing-subscriber", ] -[[package]] -name = "nibble_vec" -version = "0.1.0" -source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "77a5d83df9f36fe23f0c3648c6bbb8b0298bb5f1939c8f2704431371f4b84d43" -dependencies = [ - "smallvec", -] - [[package]] name = "no-std-compat" version = "0.4.1" @@ -1305,16 +1289,6 @@ dependencies = [ "proc-macro2", ] -[[package]] -name = "radix_trie" -version = "0.2.1" -source = "registry+https://github.com/rust-lang/crates.io-index" -checksum = "c069c179fcdc6a2fe24d8d18305cf085fdbd4f922c041943e203685d6a1c58fd" -dependencies = [ - "endian-type", - "nibble_vec", -] - [[package]] name = "rand" version = "0.8.5" diff --git a/Cargo.toml b/Cargo.toml index c9081d8..7b900d1 100644 --- a/Cargo.toml +++ b/Cargo.toml @@ -36,7 +36,6 @@ noodles = { version = "0.34.0", features = [ num-format = "0.4.0" plotly = "0.8.1" prettytable-rs = "0.9.0" -radix_trie = "0.2.1" rand = "0.8.5" rand_distr = "0.4.3" regex = "1.5.5" From 390dc1f57683c7e32f03ab312dcfa877c7aabb00 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 26 Dec 2023 15:47:02 -0500 Subject: [PATCH 39/91] feat(derive/junction-annotation): better results reporting --- src/derive/junction_annotation/compute.rs | 74 ++++++++++++++++++++--- src/derive/junction_annotation/results.rs | 48 +++++++++++++-- 2 files changed, 108 insertions(+), 14 deletions(-) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index a76a072..7252bc9 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -264,15 +264,24 @@ pub fn summarize( params: &JunctionAnnotationParameters, ) -> anyhow::Result<()> { // Filter out junctions that are too short or don't have enough read support. + let mut num_rejected: usize = 0; let mut num_junctions_too_short: usize = 0; let mut num_not_enough_support: usize = 0; for (_, v) in results.junction_annotations.known.iter_mut() { v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; + if *count < params.min_read_support as usize { + num_not_enough_support += 1; + } + num_rejected += 1; false } else if *count < params.min_read_support as usize { num_not_enough_support += 1; + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + } + num_rejected += 1; false } else { true @@ -283,9 +292,17 @@ pub fn summarize( v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; + if *count < params.min_read_support as usize { + num_not_enough_support += 1; + } + num_rejected += 1; false } else if *count < params.min_read_support as usize { num_not_enough_support += 1; + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + } + num_rejected += 1; false } else { true @@ -296,9 +313,17 @@ pub fn summarize( v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; + if *count < params.min_read_support as usize { + num_not_enough_support += 1; + } + num_rejected += 1; false } else if *count < params.min_read_support as usize { num_not_enough_support += 1; + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + } + num_rejected += 1; false } else { true @@ -313,15 +338,24 @@ pub fn summarize( v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; + if *count < params.min_read_support as usize { + num_not_enough_support += 1; + } + num_rejected += 1; false } else if *count < params.min_read_support as usize { num_not_enough_support += 1; + if end.get() - start.get() < params.min_intron_length { + num_junctions_too_short += 1; + } + num_rejected += 1; false } else { true } }); } + results.summary.total_rejected_junctions = num_rejected; results.summary.intron_too_short = num_junctions_too_short; results.summary.junctions_with_not_enough_read_support = num_not_enough_support; @@ -332,7 +366,7 @@ pub fn summarize( .values() .map(|v| v.len()) .sum(); - results.summary.known_spliced_reads = results + results.summary.known_junctions_read_support = results .junction_annotations .known .values() @@ -344,7 +378,7 @@ pub fn summarize( .values() .map(|v| v.len()) .sum(); - results.summary.partial_novel_spliced_reads = results + results.summary.partial_novel_junctions_read_support = results .junction_annotations .partial_novel .values() @@ -356,7 +390,7 @@ pub fn summarize( .values() .map(|v| v.len()) .sum(); - results.summary.complete_novel_spliced_reads = results + results.summary.complete_novel_junctions_read_support = results .junction_annotations .complete_novel .values() @@ -368,7 +402,7 @@ pub fn summarize( .values() .map(|v| v.len()) .sum(); - results.summary.unannotated_reference_spliced_reads = results + results.summary.unannotated_reference_junctions_read_support = results .junction_annotations .unannotated_reference .values() @@ -380,10 +414,34 @@ pub fn summarize( + results.summary.partial_novel_junctions + results.summary.complete_novel_junctions + results.summary.unannotated_reference_junctions; - results.summary.total_spliced_reads = results.summary.known_spliced_reads - + results.summary.partial_novel_spliced_reads - + results.summary.complete_novel_spliced_reads - + results.summary.unannotated_reference_spliced_reads; + results.summary.total_junctions_read_support = results.summary.known_junctions_read_support + + results.summary.partial_novel_junctions_read_support + + results.summary.complete_novel_junctions_read_support + + results.summary.unannotated_reference_junctions_read_support; + + // Calculate percentages. + let total_junctions = results.summary.total_junctions as f64 + - results.summary.unannotated_reference_junctions as f64; + results.summary.known_junctions_percent = + results.summary.known_junctions as f64 / total_junctions * 100.0; + results.summary.partial_novel_junctions_percent = + results.summary.partial_novel_junctions as f64 / total_junctions * 100.0; + results.summary.complete_novel_junctions_percent = + results.summary.complete_novel_junctions as f64 / total_junctions * 100.0; + + // Calculate average read support. + results.summary.average_junction_read_support = results.summary.total_junctions_read_support + as f64 + / results.summary.total_junctions as f64; + results.summary.average_known_junction_read_support = + results.summary.known_junctions_read_support as f64 + / results.summary.known_junctions as f64; + results.summary.average_partial_novel_junction_read_support = + results.summary.partial_novel_junctions_read_support as f64 + / results.summary.partial_novel_junctions as f64; + results.summary.average_complete_novel_junction_read_support = + results.summary.complete_novel_junctions_read_support as f64 + / results.summary.complete_novel_junctions as f64; Ok(()) } diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 9c63a03..6f7330b 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -75,6 +75,7 @@ impl Serialize for JunctionAnnotations { #[derive(Clone, Default, Serialize)] pub struct RecordMetrics { /// The number of records that have been fully processed. + /// This is the number of spliced records that have been considered. pub processed: usize, /// The number of records that have been ignored because of their flags. @@ -101,7 +102,10 @@ pub struct SummaryResults { pub total_junctions: usize, /// The total number of spliced reads observed in the file. - pub total_spliced_reads: usize, + pub total_junctions_read_support: usize, + + /// The average number of spliced reads supporting a junction. + pub average_junction_read_support: f64, /// The total number of known junctions observed in the file. pub known_junctions: usize, @@ -117,24 +121,56 @@ pub struct SummaryResults { pub unannotated_reference_junctions: usize, /// The total number of known spliced reads observed in the file. - pub known_spliced_reads: usize, + pub known_junctions_read_support: usize, /// The total number of partially novel spliced reads observed in the file. - pub partial_novel_spliced_reads: usize, + pub partial_novel_junctions_read_support: usize, /// The total number of complete novel spliced reads observed in the file. - pub complete_novel_spliced_reads: usize, + pub complete_novel_junctions_read_support: usize, /// The total number of spliced reads on reference sequences for which /// junction annotations were not found. - pub unannotated_reference_spliced_reads: usize, + pub unannotated_reference_junctions_read_support: usize, + + /// The percentage of junctions that are known. + /// This percentage excludes junctions on reference sequences for which + /// junction annotations were not found. + pub known_junctions_percent: f64, + + /// The percentage of junctions that are partially novel. + /// This percentage excludes junctions on reference sequences for which + /// junction annotations were not found. + pub partial_novel_junctions_percent: f64, + + /// The percentage of junctions that are completely novel. + /// This percentage excludes junctions on reference sequences for which + /// junction annotations were not found. + pub complete_novel_junctions_percent: f64, + + /// Average number of reads supporting known junctions. + pub average_known_junction_read_support: f64, + + /// Average number of reads supporting partially novel junctions. + pub average_partial_novel_junction_read_support: f64, + + /// Average number of reads supporting completely novel junctions. + pub average_complete_novel_junction_read_support: f64, + + /// The total number of junctions that have been rejected because + /// they failed the min_read_support or the min_intron_length filter. + /// A junction can be rejected for both reasons, so do not expect this + /// number to be equal to the sum of `junctions_with_not_enough_read_support` + /// and `intron_too_short`. + pub total_rejected_junctions: usize, /// The total number of junctions which were discarded due to lack of - /// read support. + /// read support. This is not mutually exclusive with `intron_too_short`. pub junctions_with_not_enough_read_support: usize, /// The number of junctions that have been ignored because /// they failed the min_intron_length filter. + /// This is not mutually exclusive with `junctions_with_not_enough_read_support`. pub intron_too_short: usize, } From 3a7051fddb8a6b5cb0a50ab59233e451b7d46136 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 27 Dec 2023 10:09:53 -0500 Subject: [PATCH 40/91] docs(derive/junction_annotation): be more clear in results docs --- src/derive/junction_annotation/results.rs | 31 ++++++++++++++++------- 1 file changed, 22 insertions(+), 9 deletions(-) diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 6f7330b..b426d3b 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -101,7 +101,10 @@ pub struct SummaryResults { /// The total number of junctions observed in the file. pub total_junctions: usize, - /// The total number of spliced reads observed in the file. + /// The total number of splices detected observed in the file. + /// More than one splice can be observed per read, especially + /// with long read data, so this number is not necessarily equal + /// to the number of spliced reads. It may be greater. pub total_junctions_read_support: usize, /// The average number of spliced reads supporting a junction. @@ -120,17 +123,27 @@ pub struct SummaryResults { /// annotations were not found. pub unannotated_reference_junctions: usize, - /// The total number of known spliced reads observed in the file. + /// The number of reads supporting known junctions. + /// If a read supports more than one known junction, it is counted more than once. + /// A read with more one junction may also contribute to the support of + /// partially novel or completely novel junctions. pub known_junctions_read_support: usize, - /// The total number of partially novel spliced reads observed in the file. + /// The number of reads supporting partially novel junctions. + /// If a read supports more than one partially novel junction, it is counted more than once. + /// A read with more one junction may also contribute to the support of + /// known or completely novel junctions. pub partial_novel_junctions_read_support: usize, - /// The total number of complete novel spliced reads observed in the file. + /// The number of reads supporting completely novel junctions. + /// If a read supports more than one completely novel junction, it is counted more than once. + /// A read with more one junction may also contribute to the support of + /// known or partially novel junctions. pub complete_novel_junctions_read_support: usize, - /// The total number of spliced reads on reference sequences for which + /// The number of reads supporting junctions on reference sequences for which /// junction annotations were not found. + /// If a read supports more than one junction, it is counted more than once. pub unannotated_reference_junctions_read_support: usize, /// The percentage of junctions that are known. @@ -160,17 +173,17 @@ pub struct SummaryResults { /// The total number of junctions that have been rejected because /// they failed the min_read_support or the min_intron_length filter. /// A junction can be rejected for both reasons, so do not expect this - /// number to be equal to the sum of `junctions_with_not_enough_read_support` - /// and `intron_too_short`. + /// number to be equal to the sum of junctions_with_not_enough_read_support + /// and intron_too_short. pub total_rejected_junctions: usize, /// The total number of junctions which were discarded due to lack of - /// read support. This is not mutually exclusive with `intron_too_short`. + /// read support. This is not mutually exclusive with intron_too_short. pub junctions_with_not_enough_read_support: usize, /// The number of junctions that have been ignored because /// they failed the min_intron_length filter. - /// This is not mutually exclusive with `junctions_with_not_enough_read_support`. + /// This is not mutually exclusive with junctions_with_not_enough_read_support. pub intron_too_short: usize, } From 5e3cb89974fbcb9aa02c872d1905bb5942b9aa31 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 28 Dec 2023 09:50:30 -0500 Subject: [PATCH 41/91] feat(derive/junction_annotation): add short options to params --- src/derive/command/junction_annotation.rs | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index e16c5ac..6880e94 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -34,15 +34,15 @@ pub struct JunctionAnnotationArgs { /// Minimum intron length to consider. /// An intron is defined as an `N` CIGAR operation of any length. - #[arg(long, value_name = "USIZE", default_value = "50")] + #[arg(short = 'i', long, value_name = "USIZE", default_value = "50")] pub min_intron_length: usize, /// Add +- this amount to intron positions when looking up exon positions. - #[arg(long, value_name = "U8", default_value = "0")] + #[arg(short = 'k', long, value_name = "U8", default_value = "0")] pub fuzzy_junction_match_range: u8, /// Minimum number of reads supporting a junction to be considered. - #[arg(long, value_name = "U8", default_value = "2")] + #[arg(short = 'r', long, value_name = "U8", default_value = "2")] pub min_read_support: u8, /// Minumum mapping quality for a record to be considered. From 1ea415482b91c74cb9e13ed87b1cfb2bce8565d2 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 28 Dec 2023 09:52:37 -0500 Subject: [PATCH 42/91] chore: return anyhow::Ok where appropriate --- src/derive/command/junction_annotation.rs | 4 ++-- src/derive/junction_annotation/compute.rs | 8 +++----- 2 files changed, 5 insertions(+), 7 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 6880e94..89e8cc2 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -143,12 +143,12 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { ); // (3) Summarize found junctions. - compute::summarize(&mut results, ¶ms)?; + compute::summarize(&mut results, ¶ms); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&results).unwrap(); print!("{}", output); - Ok(()) + anyhow::Ok(()) } diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 7252bc9..ca47f75 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -1,6 +1,7 @@ //! Module holding the logic for annotating junctions. use anyhow::bail; +use anyhow::Ok; use noodles::sam::alignment::Record; use noodles::sam::record::cigar::op::Kind; use noodles::sam::Header; @@ -259,10 +260,7 @@ pub fn process( } /// Main function to summarize the results of the junction_annotation subcommand. -pub fn summarize( - results: &mut JunctionAnnotationResults, - params: &JunctionAnnotationParameters, -) -> anyhow::Result<()> { +pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnotationParameters) { // Filter out junctions that are too short or don't have enough read support. let mut num_rejected: usize = 0; let mut num_junctions_too_short: usize = 0; @@ -442,6 +440,6 @@ pub fn summarize( results.summary.average_complete_novel_junction_read_support = results.summary.complete_novel_junctions_read_support as f64 / results.summary.complete_novel_junctions as f64; +} - Ok(()) } From ee77abd4e183a4a311fb14175866c3ec4f19a6d5 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 28 Dec 2023 09:53:09 -0500 Subject: [PATCH 43/91] tests: add a test for process() and summarize() --- src/derive/junction_annotation/compute.rs | 416 ++++++++++++++++++++++ 1 file changed, 416 insertions(+) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index ca47f75..2aefbcc 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -442,4 +442,420 @@ pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnot / results.summary.complete_novel_junctions as f64; } +#[cfg(test)] +mod tests { + use super::*; + use noodles::core::Position; + use noodles::sam::header::record::value::map; + use noodles::sam::header::record::value::map::header::Version; + use noodles::sam::header::record::value::map::{Map, ReferenceSequence}; + use noodles::sam::record::MappingQuality; + use noodles::sam::record::ReadName; + + #[test] + fn test_process_and_summarize() { + // Setup + let mut results = JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + fuzzy_junction_match_range: 0, + min_read_support: 2, + min_mapq: 30, + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let header = Header::builder() + .set_header(Map::::new(Version::new(1, 6))) + .add_reference_sequence( + "sq1".parse().unwrap(), + Map::::new(NonZeroUsize::try_from(800).unwrap()), + ) + .add_reference_sequence( + "sq1_random".parse().unwrap(), // unannotated + Map::::new(NonZeroUsize::try_from(400).unwrap()), + ) + .build(); + let exon_starts: HashMap<&str, Vec> = + HashMap::from([("sq1", vec![1, 11, 21, 31, 41, 51, 61, 71])]); + let exon_ends = exon_starts + .iter() + .map(|(k, v)| (*k, v.iter().map(|e| e + 10).collect())) + .collect::>>(); + + // Test known junction + let mut record = Record::default(); + let r1_name: ReadName = "known1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); + *record.flags_mut() = 0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.ignored_flags, 0); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test that unmapped gets ignored + let mut record = Record::default(); + let r2_name: ReadName = "unmapped".parse().unwrap(); + *record.read_name_mut() = Some(r2_name); + *record.flags_mut() = 0x4.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(255); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test partial novel junction + let mut record = Record::default(); + let r3_name: ReadName = "partial1".parse().unwrap(); + *record.read_name_mut() = Some(r3_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M12N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 2); + assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test partial novel junction (again for more read supprt) + let mut record = Record::default(); + let r3_name: ReadName = "partial2".parse().unwrap(); + *record.read_name_mut() = Some(r3_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M12N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 3); + assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test that supplementary alignments get counted + let mut record = Record::default(); + let r4_name: ReadName = "supplementary_and_known2".parse().unwrap(); + *record.read_name_mut() = Some(r4_name); + *record.flags_mut() = 0x800.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 4); + assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test that secondary alignments get ignored + let mut record = Record::default(); + let r5_name: ReadName = "secondary".parse().unwrap(); + *record.read_name_mut() = Some(r5_name); + *record.flags_mut() = 0x100.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 4); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test complete novel junction + let mut record = Record::default(); + let r6_name: ReadName = "novel1".parse().unwrap(); + *record.read_name_mut() = Some(r6_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "8M15N8M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 5); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test complete novel junction (again for more read supprt) + let mut record = Record::default(); + let r6_name: ReadName = "novel2".parse().unwrap(); + *record.read_name_mut() = Some(r6_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "8M15N8M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 0); + assert_eq!(results.records.missing_mapq, 0); + + // Test fails MAPQ filter + let mut record = Record::default(); + let r7_name: ReadName = "low_mapq".parse().unwrap(); + *record.read_name_mut() = Some(r7_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(20); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 0); + + // Test missing MAPQ + let mut record = Record::default(); + let r8_name: ReadName = "missing_mapq".parse().unwrap(); + *record.read_name_mut() = Some(r8_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(255); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 1); + + // Test that intron is too short + let mut record = Record::default(); + let r9_name: ReadName = "short".parse().unwrap(); + *record.read_name_mut() = Some(r9_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "5M5N5M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 1); + + // That that reads not spliced are ignored + let mut record = Record::default(); + let r10_name: ReadName = "not_spliced".parse().unwrap(); + *record.read_name_mut() = Some(r10_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 7); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 1); + + // Test unannoted reference + let mut record = Record::default(); + let r11_name: ReadName = "unannotated1".parse().unwrap(); + *record.read_name_mut() = Some(r11_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(1); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 8); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 1); + + // Test unannoted reference (again for more read supprt) + let mut record = Record::default(); + let r11_name: ReadName = "unannotated2".parse().unwrap(); + *record.read_name_mut() = Some(r11_name); + *record.flags_mut() = 0x0.into(); + *record.reference_sequence_id_mut() = Some(1); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + process( + &record, + &exon_starts, + &exon_ends, + &header, + ¶ms, + &mut results, + ) + .unwrap(); + assert_eq!(results.records.processed, 9); + assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.low_mapq, 1); + assert_eq!(results.records.missing_mapq, 1); + + // Test summarize + summarize(&mut results, ¶ms); + + assert_eq!(results.summary.total_rejected_junctions, 1); + assert_eq!(results.summary.intron_too_short, 1); + assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); + assert_eq!(results.summary.known_junctions, 1); + assert_eq!(results.summary.known_junctions_read_support, 2); + assert_eq!(results.summary.partial_novel_junctions, 1); + assert_eq!(results.summary.partial_novel_junctions_read_support, 2); + assert_eq!(results.summary.complete_novel_junctions, 1); + assert_eq!(results.summary.complete_novel_junctions_read_support, 2); + assert_eq!(results.summary.unannotated_reference_junctions, 1); + assert_eq!( + results.summary.unannotated_reference_junctions_read_support, + 2 + ); + assert_eq!(results.summary.total_junctions, 4); + assert_eq!(results.summary.total_junctions_read_support, 8); + assert_eq!(results.summary.known_junctions_percent, 33.33333333333333); + assert_eq!( + results.summary.partial_novel_junctions_percent, + 33.33333333333333 + ); + assert_eq!( + results.summary.complete_novel_junctions_percent, + 33.33333333333333 + ); + assert_eq!(results.summary.average_junction_read_support, 2.0); + assert_eq!(results.summary.average_known_junction_read_support, 2.0); + assert_eq!( + results.summary.average_partial_novel_junction_read_support, + 2.0 + ); + assert_eq!( + results.summary.average_complete_novel_junction_read_support, + 2.0 + ); + } } From 65c49354231dc24b454b1e03301d77b2084041dc Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 28 Dec 2023 09:59:51 -0500 Subject: [PATCH 44/91] chore: typos --- src/derive/junction_annotation/compute.rs | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 2aefbcc..7ac4769 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -555,7 +555,7 @@ mod tests { assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); - // Test partial novel junction (again for more read supprt) + // Test partial novel junction (again for more read support) let mut record = Record::default(); let r3_name: ReadName = "partial2".parse().unwrap(); *record.read_name_mut() = Some(r3_name); @@ -651,7 +651,7 @@ mod tests { assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); - // Test complete novel junction (again for more read supprt) + // Test complete novel junction (again for more read support) let mut record = Record::default(); let r6_name: ReadName = "novel2".parse().unwrap(); *record.read_name_mut() = Some(r6_name); @@ -795,7 +795,7 @@ mod tests { assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); - // Test unannoted reference (again for more read supprt) + // Test unannoted reference (again for more read support) let mut record = Record::default(); let r11_name: ReadName = "unannotated2".parse().unwrap(); *record.read_name_mut() = Some(r11_name); From 6fcbd60c7078c623c4b2dd5454324004d172827a Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 10 Jan 2024 11:25:06 -0500 Subject: [PATCH 45/91] feat: remove fuzzy searching ability. Boosts performance as well. --- src/derive/command/junction_annotation.rs | 28 +++------ src/derive/junction_annotation/compute.rs | 70 ++++++++--------------- 2 files changed, 32 insertions(+), 66 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 89e8cc2..fa1d41b 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -1,6 +1,7 @@ //! Functionality relating to the `ngs derive junction_annotation` subcommand itself. use std::collections::HashMap; +use std::collections::HashSet; use std::path::PathBuf; use anyhow::Context; @@ -37,13 +38,9 @@ pub struct JunctionAnnotationArgs { #[arg(short = 'i', long, value_name = "USIZE", default_value = "50")] pub min_intron_length: usize, - /// Add +- this amount to intron positions when looking up exon positions. - #[arg(short = 'k', long, value_name = "U8", default_value = "0")] - pub fuzzy_junction_match_range: u8, - /// Minimum number of reads supporting a junction to be considered. - #[arg(short = 'r', long, value_name = "U8", default_value = "2")] - pub min_read_support: u8, + #[arg(short = 'r', long, value_name = "usize", default_value = "2")] + pub min_read_support: usize, /// Minumum mapping quality for a record to be considered. /// Set to 0 to disable this filter and allow reads _without_ @@ -68,8 +65,8 @@ pub struct JunctionAnnotationArgs { pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { info!("Starting derive junction_annotation subcommand."); - let mut exon_starts: HashMap<&str, Vec> = HashMap::new(); - let mut exon_ends: HashMap<&str, Vec> = HashMap::new(); + let mut exon_starts: HashMap<&str, HashSet> = HashMap::new(); + let mut exon_ends: HashMap<&str, HashSet> = HashMap::new(); // (1) Parse the GFF file and collect all exon features. debug!("Reading all records in GFF."); @@ -91,18 +88,8 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { let start: usize = record.start().into(); let end: usize = record.end().into(); - exon_starts.entry(seq_name).or_default().push(start); - exon_ends.entry(seq_name).or_default().push(end + 1); // TODO why +1? It works - } - - debug!("Finalizing GFF features lookup."); - for starts in exon_starts.values_mut() { - starts.sort_unstable(); - starts.dedup(); - } - for ends in exon_ends.values_mut() { - ends.sort_unstable(); - ends.dedup(); + exon_starts.entry(seq_name).or_default().insert(start); + exon_ends.entry(seq_name).or_default().insert(end + 1); // TODO why +1? It works } debug!("Done reading GFF."); @@ -111,7 +98,6 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { let mut results = JunctionAnnotationResults::default(); let params = compute::JunctionAnnotationParameters { min_intron_length: args.min_intron_length, - fuzzy_junction_match_range: args.fuzzy_junction_match_range, min_read_support: args.min_read_support, min_mapq: args.min_mapq, no_supplementary: args.no_supplementary, diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 7ac4769..06b9de2 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -6,6 +6,7 @@ use noodles::sam::alignment::Record; use noodles::sam::record::cigar::op::Kind; use noodles::sam::Header; use std::collections::HashMap; +use std::collections::HashSet; use std::num::NonZeroUsize; use crate::derive::junction_annotation::results::JunctionAnnotationResults; @@ -15,11 +16,8 @@ pub struct JunctionAnnotationParameters { /// Minimum intron length to consider. pub min_intron_length: usize, - /// Add +- this amount to intron positions when looking up exon positions. - pub fuzzy_junction_match_range: u8, - /// Minimum number of reads supporting a junction to be considered. - pub min_read_support: u8, + pub min_read_support: usize, /// Minumum mapping quality for a record to be considered. /// 0 if MAPQ shouldn't be considered. @@ -38,8 +36,8 @@ pub struct JunctionAnnotationParameters { /// Main function to annotate junctions one record at a time. pub fn process( record: &Record, - exon_starts: &HashMap<&str, Vec>, - exon_ends: &HashMap<&str, Vec>, + exon_starts: &HashMap<&str, HashSet>, + exon_ends: &HashMap<&str, HashSet>, header: &Header, params: &JunctionAnnotationParameters, results: &mut JunctionAnnotationResults, @@ -177,28 +175,11 @@ pub fn process( let mut intron_start_known = false; let mut intron_end_known = false; - // To allow collapsing fuzzy junctions, - // we need to store the reference positions of the exon boundaries. - // We initialize these values to the position of the found intron. - let mut ref_intron_start = intron_start; - let mut ref_intron_end = intron_end; - for exon_end in exon_ends.iter() { - if intron_start >= (exon_end - params.fuzzy_junction_match_range as usize) - && intron_start <= (exon_end + params.fuzzy_junction_match_range as usize) - { - intron_start_known = true; - ref_intron_start = *exon_end; - break; - } + if exon_ends.contains(&intron_start) { + intron_start_known = true; } - for exon_start in exon_starts.iter() { - if intron_end >= (exon_start - params.fuzzy_junction_match_range as usize) - && intron_end <= (exon_start + params.fuzzy_junction_match_range as usize) - { - intron_end_known = true; - ref_intron_end = *exon_start; - break; - } + if exon_starts.contains(&intron_end) { + intron_end_known = true; } match (intron_start_known, intron_end_known) { @@ -211,8 +192,8 @@ pub fn process( .entry(seq_name.to_string()) .or_default() .entry(( - NonZeroUsize::new(ref_intron_start).unwrap(), - NonZeroUsize::new(ref_intron_end).unwrap(), + NonZeroUsize::new(intron_start).unwrap(), + NonZeroUsize::new(intron_end).unwrap(), )) .and_modify(|e| *e += 1) .or_insert(1); @@ -227,8 +208,8 @@ pub fn process( .entry(seq_name.to_string()) .or_default() .entry(( - NonZeroUsize::new(ref_intron_start).unwrap(), - NonZeroUsize::new(ref_intron_end).unwrap(), + NonZeroUsize::new(intron_start).unwrap(), + NonZeroUsize::new(intron_end).unwrap(), )) .and_modify(|e| *e += 1) .or_insert(1); @@ -242,8 +223,8 @@ pub fn process( .entry(seq_name.to_string()) .or_default() .entry(( - NonZeroUsize::new(ref_intron_start).unwrap(), - NonZeroUsize::new(ref_intron_end).unwrap(), + NonZeroUsize::new(intron_start).unwrap(), + NonZeroUsize::new(intron_end).unwrap(), )) .and_modify(|e| *e += 1) .or_insert(1); @@ -269,12 +250,12 @@ pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnot v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; - if *count < params.min_read_support as usize { + if *count < params.min_read_support { num_not_enough_support += 1; } num_rejected += 1; false - } else if *count < params.min_read_support as usize { + } else if *count < params.min_read_support { num_not_enough_support += 1; if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; @@ -290,12 +271,12 @@ pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnot v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; - if *count < params.min_read_support as usize { + if *count < params.min_read_support { num_not_enough_support += 1; } num_rejected += 1; false - } else if *count < params.min_read_support as usize { + } else if *count < params.min_read_support { num_not_enough_support += 1; if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; @@ -311,12 +292,12 @@ pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnot v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; - if *count < params.min_read_support as usize { + if *count < params.min_read_support { num_not_enough_support += 1; } num_rejected += 1; false - } else if *count < params.min_read_support as usize { + } else if *count < params.min_read_support { num_not_enough_support += 1; if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; @@ -336,12 +317,12 @@ pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnot v.retain(|(start, end), count| { if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; - if *count < params.min_read_support as usize { + if *count < params.min_read_support { num_not_enough_support += 1; } num_rejected += 1; false - } else if *count < params.min_read_support as usize { + } else if *count < params.min_read_support { num_not_enough_support += 1; if end.get() - start.get() < params.min_intron_length { num_junctions_too_short += 1; @@ -458,7 +439,6 @@ mod tests { let mut results = JunctionAnnotationResults::default(); let params = JunctionAnnotationParameters { min_intron_length: 10, - fuzzy_junction_match_range: 0, min_read_support: 2, min_mapq: 30, no_supplementary: false, @@ -476,12 +456,12 @@ mod tests { Map::::new(NonZeroUsize::try_from(400).unwrap()), ) .build(); - let exon_starts: HashMap<&str, Vec> = - HashMap::from([("sq1", vec![1, 11, 21, 31, 41, 51, 61, 71])]); + let exon_starts: HashMap<&str, HashSet> = + HashMap::from([("sq1", HashSet::from([1, 11, 21, 31, 41, 51, 61, 71]))]); let exon_ends = exon_starts .iter() .map(|(k, v)| (*k, v.iter().map(|e| e + 10).collect())) - .collect::>>(); + .collect::>>(); // Test known junction let mut record = Record::default(); From d872c827e39f20aebf423ad18241470f5fd8802c Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 6 Feb 2024 13:02:08 -0500 Subject: [PATCH 46/91] feat: first pass implementation of `encoding` --- src/derive/command/encoding.rs | 61 ++++++++++++++++++++++++-- src/derive/encoding/compute.rs | 78 ++++++++++++++++++++++++++++++++++ 2 files changed, 136 insertions(+), 3 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index b0b03dd..31a2965 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -1,15 +1,26 @@ //! Functionality relating to the `ngs derive encoding` subcommand itself. +use std::collections::HashSet; +use std::io::BufReader; use std::path::PathBuf; +use anyhow::Context; use anyhow::Ok; use clap::Args; +use noodles::bam; +use num_format::Locale; +use num_format::ToFormattedString; +use tracing::info; + +use crate::derive::encoding::compute; +use crate::utils::args::NumberOfRecords; +use crate::utils::display::RecordCounter; /// Clap arguments for the `ngs derive encoding` subcommand. #[derive(Args)] pub struct DeriveEncodingArgs { - // Source NGS file (BAM or FASTQ). - #[arg(value_name = "NGS_FILE")] + /// Source BAM. + #[arg(value_name = "BAM")] src: PathBuf, /// Only examine the first n records in the file. @@ -18,6 +29,50 @@ pub struct DeriveEncodingArgs { } /// Main function for the `ngs derive encoding` subcommand. -pub fn derive(_args: DeriveEncodingArgs) -> anyhow::Result<()> { +pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { + info!("Starting derive readlen subcommand."); + + let file = std::fs::File::open(args.src); + let reader = file + .map(BufReader::new) + .with_context(|| "opening BAM file")?; + let mut reader = bam::Reader::new(reader); + let _header: String = reader.read_header()?.parse()?; + reader.read_reference_sequences()?; + + let mut score_set: HashSet = HashSet::new(); + + // (1) Collect quality scores from reads within the + // file. Support for sampling only a portion of the reads is provided. + let num_records = NumberOfRecords::from(args.num_records); + let mut counter = RecordCounter::new(); + + for result in reader.lazy_records() { + let record = result?; + + for i in 0..record.quality_scores().len() { + let score = record.quality_scores().as_ref()[i]; + score_set.insert(score); + } + + counter.inc(); + if counter.time_to_break(&num_records) { + break; + } + } + + info!( + "Processed {} records.", + counter.get().to_formatted_string(&Locale::en) + ); + + // (2) Derive encoding from the observed quality scores + let result = compute::predict(score_set)?; + + // (3) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); + Ok(()) } diff --git a/src/derive/encoding/compute.rs b/src/derive/encoding/compute.rs index 469063d..40d9366 100644 --- a/src/derive/encoding/compute.rs +++ b/src/derive/encoding/compute.rs @@ -1 +1,79 @@ //! Module holding the logic for computing the quality score encoding. + +use anyhow::bail; +use serde::Serialize; +use std::collections::HashSet; + +const MAX_VALID_PHRED_SCORE: u8 = 93; +const SANGER_MIN: u8 = 0; +const ILLUMINA_1_0_MIN: u8 = 26; +const ILLUMINA_1_3_MIN: u8 = 31; + +/// Struct holding the final results for an `ngs derive encoding` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedEncodingResult { + /// Whether or not the `ngs derive encoding` subcommand succeeded. + pub succeeded: bool, + + /// The detected quality score encoding, if available. + pub encoding: Option, + + /// The minimum quality score observed. + pub observed_min: u8, + + /// The maximum quality score observed. + pub observed_max: u8, +} + +impl DerivedEncodingResult { + /// Creates a new [`DerivedEncodingResult`]. + pub fn new( + succeeded: bool, + encoding: Option, + observed_min: u8, + observed_max: u8, + ) -> Self { + DerivedEncodingResult { + succeeded, + encoding, + observed_min, + observed_max, + } + } +} + +/// Main method to evaluate the observed quality scores and +/// return a result for the derived encoding. This may fail, and the +/// resulting [`DerivedEncodingResult`] should be evaluated accordingly. +pub fn predict(score_set: HashSet) -> Result { + if score_set.is_empty() { + bail!("No quality scores were detected in the file."); + } + + let observed_min = *score_set.iter().min().unwrap(); + let observed_max = *score_set.iter().max().unwrap(); + + let mut result = DerivedEncodingResult::new(false, None, observed_min, observed_max); + + if observed_max > MAX_VALID_PHRED_SCORE { + return anyhow::Ok(result); + } + match observed_min { + ILLUMINA_1_3_MIN..=MAX_VALID_PHRED_SCORE => { + result.succeeded = true; + result.encoding = Some("Illumina 1.3".to_string()); + } + ILLUMINA_1_0_MIN..=MAX_VALID_PHRED_SCORE => { + result.succeeded = true; + result.encoding = Some("Illumina 1.0".to_string()); + } + SANGER_MIN..=MAX_VALID_PHRED_SCORE => { + result.succeeded = true; + result.encoding = Some("Sanger/Illumina 1.8".to_string()); + } + _ => bail!("This shouldn't be possible!"), + } + + anyhow::Ok(result) +} From 89b60e02f98715e494a81df3576a76c83be57352 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 8 Feb 2024 09:55:37 -0500 Subject: [PATCH 47/91] [WIP] to share code. Partial strandedness implementation --- src/derive.rs | 1 + src/derive/command.rs | 5 + src/derive/command/endedness.rs | 5 +- src/derive/command/junction_annotation.rs | 18 +- src/derive/command/strandedness.rs | 213 ++++++++++++++ src/derive/encoding/compute.rs | 2 +- src/derive/endedness/compute.rs | 50 +--- src/derive/strandedness.rs | 3 + src/derive/strandedness/compute.rs | 344 ++++++++++++++++++++++ src/main.rs | 3 + src/utils.rs | 1 + src/utils/read_groups.rs | 54 ++++ 12 files changed, 637 insertions(+), 62 deletions(-) create mode 100644 src/derive/command/strandedness.rs create mode 100644 src/derive/strandedness.rs create mode 100644 src/derive/strandedness/compute.rs create mode 100644 src/utils/read_groups.rs diff --git a/src/derive.rs b/src/derive.rs index 104bfe3..6a28e5f 100644 --- a/src/derive.rs +++ b/src/derive.rs @@ -6,3 +6,4 @@ pub mod endedness; pub mod instrument; pub mod junction_annotation; pub mod readlen; +pub mod strandedness; diff --git a/src/derive/command.rs b/src/derive/command.rs index 5d4a593..8ded0d3 100644 --- a/src/derive/command.rs +++ b/src/derive/command.rs @@ -5,6 +5,7 @@ pub mod endedness; pub mod instrument; pub mod junction_annotation; pub mod readlen; +pub mod strandedness; use clap::Args; use clap::Subcommand; @@ -36,6 +37,10 @@ pub enum DeriveSubcommand { /// Derives the read length of the file. Readlen(self::readlen::DeriveReadlenArgs), + /// Derives the strandedness of the RNA-Seq file. + /// This subcommand requires a GFF file. + Strandedness(self::strandedness::DeriveStrandednessArgs), + /// Annotates junctions in the file. /// This subcommand requires a GFF file with features to annotate. /// This subcommand does not "derive" anything, but is included here for diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 7a87136..bc65456 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -14,14 +14,13 @@ use tracing::info; use tracing::trace; use crate::derive::endedness::compute; -use crate::derive::endedness::compute::{ - validate_read_group_info, OrderingFlagsCounts, OVERALL, UNKNOWN_READ_GROUP, -}; +use crate::derive::endedness::compute::OrderingFlagsCounts; use crate::utils::args::arg_in_range as deviance_in_range; use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; +use crate::utils::read_groups::{validate_read_group_info, OVERALL, UNKNOWN_READ_GROUP}; /// Clap arguments for the `ngs derive endedness` subcommand. #[derive(Args)] diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index fa1d41b..ac50cb4 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -31,34 +31,34 @@ pub struct JunctionAnnotationArgs { /// Name of the exon region feature for the gene model used. #[arg(long, value_name = "STRING", default_value = "exon")] - pub exon_feature_name: String, + exon_feature_name: String, /// Minimum intron length to consider. /// An intron is defined as an `N` CIGAR operation of any length. #[arg(short = 'i', long, value_name = "USIZE", default_value = "50")] - pub min_intron_length: usize, + min_intron_length: usize, /// Minimum number of reads supporting a junction to be considered. - #[arg(short = 'r', long, value_name = "usize", default_value = "2")] - pub min_read_support: usize, + #[arg(short = 'r', long, value_name = "USIZE", default_value = "2")] + min_read_support: usize, /// Minumum mapping quality for a record to be considered. /// Set to 0 to disable this filter and allow reads _without_ /// a mapping quality to be considered. #[arg(short, long, value_name = "U8", default_value = "30")] - pub min_mapq: u8, + min_mapq: u8, /// Do not count supplementary alignments. #[arg(long)] - pub no_supplementary: bool, + no_supplementary: bool, /// Do count secondary alignments. #[arg(long)] - pub count_secondary: bool, + count_secondary: bool, /// Do count duplicates. #[arg(long)] - pub count_duplicates: bool, + count_duplicates: bool, } /// Main function for the `ngs derive junction_annotation` subcommand. @@ -131,7 +131,7 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { // (3) Summarize found junctions. compute::summarize(&mut results, ¶ms); - // (3) Print the output to stdout as JSON (more support for different output + // (4) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&results).unwrap(); print!("{}", output); diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs new file mode 100644 index 0000000..6d4ab7b --- /dev/null +++ b/src/derive/command/strandedness.rs @@ -0,0 +1,213 @@ +//! Functionality relating to the `ngs derive strandedness` subcommand itself. + +use std::collections::HashMap; +use std::fs::File; +use std::path::PathBuf; + +use anyhow::bail; +use anyhow::Context; +use clap::Args; +use noodles::bam; +use noodles::gff; +use noodles::sam; +use rust_lapper::{Interval, Lapper}; +use tracing::debug; +use tracing::info; + +use crate::derive::strandedness::compute; +use crate::derive::strandedness::compute::ParsedBAMFile; +use crate::derive::strandedness::compute::StrandednessFilters; +use crate::utils::formats; + +/// Clap arguments for the `ngs derive strandedness` subcommand. +#[derive(Args)] +pub struct DeriveStrandednessArgs { + /// Source BAM. + #[arg(value_name = "BAM")] + src: PathBuf, + + /// Features GFF file. + #[arg(short = 'f', long, required = true, value_name = "PATH")] + features_gff: PathBuf, + + /// When inconclusive, the test will repeat until this many tries have been reached. + /// Evidence of previous attempts is saved and reused, + /// leading to a larger sample size with multiple attempts. + #[arg(long, value_name = "USIZE", default_value = "3")] + max_tries: usize, + + /// Filter any genes that don't have at least `m` reads. + #[arg(short = 'm', long, value_name = "USIZE", default_value = "10")] + min_reads_per_gene: usize, + + /// How many genes to sample. + #[arg(short = 'n', long, value_name = "USIZE", default_value = "1000")] + num_genes: usize, + + /// Minimum mapping quality for a record to be considered. + /// Set to 0 to disable this filter and allow reads _without_ + /// a mapping quality to be considered. + #[arg(short = 'q', long, value_name = "U8", default_value = "30")] + min_mapq: u8, + + /// Consider all genes, not just protein coding genes. + #[arg(long)] + all_genes: bool, + + /// Name of the gene region feature for the gene model used. + #[arg(long, value_name = "STRING", default_value = "gene")] + gene_feature_name: String, + + /// Name of the exon region feature for the gene model used. + #[arg(long, value_name = "STRING", default_value = "exon")] + exon_feature_name: String, + + /// Do not count supplementary alignments. + #[arg(long)] + no_supplementary: bool, + + /// Do count secondary alignments. + #[arg(long)] + count_secondary: bool, + + /// Do count duplicates. + #[arg(long)] + count_duplicates: bool, + + /// Do count QC failed reads. + #[arg(long)] + count_qc_failed: bool, + + /// At most, search this many times for genes that satisfy our search criteria. + /// Default is 10 * --num-genes. + #[arg(long, value_name = "USIZE")] + max_iterations_per_try: Option, +} + +/// Main function for the `ngs derive strandedness` subcommand. +pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { + info!("Starting derive strandedness subcommand."); + + // (1) Parse the GFF file and collect all gene features. + debug!("Reading all records in GFF."); + let mut gff = formats::gff::open(&args.features_gff) + .with_context(|| format!("opening GFF file: {}", args.features_gff.display()))?; + + let mut gene_records = Vec::new(); + let mut exon_records = Vec::new(); + let mut gene_metrics = compute::GeneRecordMetrics::default(); + let mut exon_metrics = compute::ExonRecordMetrics::default(); + for result in gff.records() { + let record = result.unwrap(); + if record.ty() == args.gene_feature_name { + // If --all-genes is set, keep the record. + // Otherwise, check the gene type or biotype and keep the record if it's protein coding. + // If the record does not have a gene type or biotype, discard it. + let mut keep_record = false; + if !args.all_genes { + let mut gene_type_value = None; + for entry in record.attributes().as_ref() { + gene_type_value = match entry.key() { + "gene_type" => Some(entry.value()), // Gencode + "gene_biotype" => Some(entry.value()), // ENSEMBL + "biotype" => Some(entry.value()), // also ENSEMBL + _ => gene_type_value, + }; + } + if let Some(gene_type_value) = gene_type_value { + if gene_type_value.to_lowercase().contains("protein") { + keep_record = true; + gene_metrics.protein_coding += 1; + } + } + } + gene_metrics.total += 1; + if keep_record { + gene_records.push(record); + } + } else if record.ty() == args.exon_feature_name { + exon_metrics.total += 1; + exon_records.push(record); + } + } + + debug!("Tabulating GFF gene and exon features."); + + if gene_records.is_empty() { + bail!("No gene records matched criteria. Check your GFF file and `--gene-feature-name` and `--all-genes` options."); + } + if exon_records.is_empty() { + bail!("No exon records matched criteria. Check your GFF file and `--exon-feature-name` option."); + } + + let mut exon_intervals: HashMap<&str, Vec>> = + HashMap::new(); + for record in &exon_records { + let seq_name = record.reference_sequence_name(); + let start: usize = record.start().into(); + let stop: usize = record.end().into(); + let strand = record.strand(); + + exon_intervals.entry(seq_name).or_default().push(Interval { + start, + stop, + val: strand, + }); + } + + let mut exons: HashMap<&str, Lapper> = HashMap::new(); + for (seq_name, intervals) in exon_intervals { + exons.insert(seq_name, Lapper::new(intervals)); + } + + debug!("Done reading GFF."); + + let mut reader = File::open(&args.src) + .map(bam::Reader::new) + .with_context(|| format!("opening BAM file: {}", args.src.display()))?; + let header = reader.read_header()?.parse()?; + let index = bam::bai::read(&args.src.with_extension("bam.bai")).with_context(|| { + format!( + "reading BAM index: {}", + args.src.with_extension("bam.bai").display() + ) + })?; + + let parsed_bam = ParsedBAMFile { + reader, + header, + index, + }; + + let filters = StrandednessFilters { + min_reads_per_gene: args.min_reads_per_gene, + min_mapq: args.min_mapq, + count_qc_failed: args.count_qc_failed, + no_supplementary: args.no_supplementary, + count_secondary: args.count_secondary, + count_duplicates: args.count_duplicates, + }; + + let max_iterations_per_try = args.max_iterations_per_try.unwrap_or(args.num_genes * 10); + let max_iterations_per_try = match max_iterations_per_try > gene_records.len() { + true => gene_records.len(), + false => max_iterations_per_try, + }; + + for try_num in 1..=args.max_tries { + info!("Starting try {} of {}", try_num, args.max_tries); + + compute::predict( + &parsed_bam, + &mut gene_records, + &exons, + max_iterations_per_try, + args.num_genes, + &filters, + &mut gene_metrics, + &mut exon_metrics, + )?; + } + + anyhow::Ok(()) +} diff --git a/src/derive/encoding/compute.rs b/src/derive/encoding/compute.rs index 40d9366..eb60962 100644 --- a/src/derive/encoding/compute.rs +++ b/src/derive/encoding/compute.rs @@ -72,7 +72,7 @@ pub fn predict(score_set: HashSet) -> Result bail!("This shouldn't be possible!"), + _ => unreachable!(), } anyhow::Ok(result) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 5990683..6ef50c9 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,22 +1,12 @@ //! Module holding the logic for computing the endedness of a BAM. -use lazy_static::lazy_static; -use noodles::sam::header; use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; use std::sync::Arc; use tracing::warn; -// Strings used to index into the HashMaps used to store the Read Group ordering flags. -// Lazy statics are used to save memory. -lazy_static! { - /// String used to index into the HashMaps used to store the "overall" ordering flags. - pub static ref OVERALL: Arc = Arc::new(String::from("overall")); - - /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. - pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); -} +use crate::utils::read_groups::{OVERALL, UNKNOWN_READ_GROUP}; /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] @@ -157,44 +147,6 @@ impl DerivedEndednessResult { } } -/// Compares the read group tags found in the records -/// and the read groups found in the header. -/// Returns a vector of read group names that were found in the header -/// but not in the records. -pub fn validate_read_group_info( - found_rgs: &HashSet>, - header: &header::Header, -) -> Vec { - let mut rgs_in_header_not_records = Vec::new(); - let mut rgs_in_records_not_header = Vec::new(); - - for (rg_id, _) in header.read_groups() { - if !found_rgs.contains(rg_id) { - rgs_in_header_not_records.push(rg_id.to_string()); - } - } - if !rgs_in_header_not_records.is_empty() { - warn!( - "The following read groups were not found in the file: {:?}", - rgs_in_header_not_records - ); - } - - for rg_id in found_rgs { - if !header.read_groups().contains_key(rg_id.as_str()) { - rgs_in_records_not_header.push(rg_id.to_string()); - } - } - if !rgs_in_records_not_header.is_empty() { - warn!( - "The following read groups were not found in the header: {:?}", - rgs_in_records_not_header - ); - } - - rgs_in_header_not_records -} - fn calculate_reads_per_template( read_names: HashMap>>, ) -> HashMap, f64> { diff --git a/src/derive/strandedness.rs b/src/derive/strandedness.rs new file mode 100644 index 0000000..0551408 --- /dev/null +++ b/src/derive/strandedness.rs @@ -0,0 +1,3 @@ +//! Supporting functionality for the `ngs derive strandedness` subcommand. + +pub mod compute; diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs new file mode 100644 index 0000000..52e7dbc --- /dev/null +++ b/src/derive/strandedness/compute.rs @@ -0,0 +1,344 @@ +//! Module holding the logic for computing the strandedness. + +use anyhow::bail; +use noodles::bam; +use noodles::core::{Position, Region}; +use noodles::gff; +use noodles::sam; +use rand::Rng; +use rust_lapper::{Interval, Lapper}; +use serde::Serialize; +use std::collections::HashMap; + +/// General gene metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize)] +pub struct GeneRecordMetrics { + /// The total number of genes found in the GFF. + pub total: usize, + + /// The number of genes that were found to be protein coding. + /// If --all-genes is set this will not be tallied. + pub protein_coding: usize, + + /// The number of genes which were discarded due to having + /// exons on both strands. + pub exons_on_both_strands: usize, + + /// The number of genes which were discarded due to not having + /// enough reads. + pub not_enough_reads: usize, +} + +/// General exon metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize)] +pub struct ExonRecordMetrics { + /// The total number of exons found in the GFF. + pub total: usize, +} + +/// General read record metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize)] +pub struct ReadRecordMetrics { + /// The number of records that have been filtered because of their flags. + /// (i.e. they were qc_fail, duplicates, secondary, or supplementary) + /// These conditions can be toggled on/off with CL flags + pub ignored_flags: usize, + + /// The number of records that have been filtered because + /// they failed the MAPQ filter. + pub low_mapq: usize, + + /// The number of records whose MAPQ couldn't be parsed and were thus discarded. + pub missing_mapq: usize, + + /// The number of records determined to be Paired-End. + pub paired_end_reads: usize, + + /// The number of records determined to be Single-End. + pub single_end_reads: usize, +} + +/// Struct for tracking count results. +#[derive(Clone, Default)] +struct Counts { + /// The number of reads determined to be Paired-End. + paired_end_reads: usize, + + /// The number of reads determined to be Single-End. + single_end_reads: usize, + + /// The number of reads that are evidence of Forward Strandedness. + forward: usize, + + /// The number of reads that are evidence of Reverse Strandedness. + reverse: usize, +} + +/// Struct holding the per read group results for an `ngs derive strandedness` +/// subcommand call. +#[derive(Debug, Serialize)] +pub struct ReadGroupDerivedStrandednessResult { + /// Name of the read group. + pub read_group: String, + + /// Whether or not strandedness was determined for this read group. + pub succeeded: bool, + + /// The strandedness of this read group or "Inconclusive". + pub strandedness: String, + + /// The total number of reads in this read group. + pub total: usize, + + /// The number of reads that are evidence of Forward Strandedness. + pub forward: usize, + + /// The number of reads that are evidence of Reverse Strandedness. + pub reverse: usize, + + /// The percent of evidence for Forward Strandedness. + pub forward_pct: f64, + + /// The percent of evidence for Reverse Strandedness. + pub reverse_pct: f64, +} + +impl ReadGroupDerivedStrandednessResult { + /// Creates a new [`ReadGroupDerivedStrandednessResult`]. + fn new( + read_group: String, + succeeded: bool, + strandedness: String, + forward: usize, + reverse: usize, + ) -> Self { + ReadGroupDerivedStrandednessResult { + read_group, + succeeded, + strandedness, + total: forward + reverse, + forward, + reverse, + forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, + reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, + } + } +} + +/// Struct holding the final results for an `ngs derive strandedness` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedStrandednessResult { + /// Whether or not the `ngs derive strandedness` subcommand succeeded. + pub succeeded: bool, + + /// The strandedness of this read group or "Inconclusive". + pub strandedness: String, + + /// The total number of reads. + pub total: usize, + + /// The number of reads that are evidence of Forward Strandedness. + pub forward: usize, + + /// The number of reads that are evidence of Reverse Strandedness. + pub reverse: usize, + + /// The percent of evidence for Forward Strandedness. + pub forward_pct: f64, + + /// The percent of evidence for Reverse Strandedness. + pub reverse_pct: f64, + + /// Vector of [`ReadGroupDerivedStrandednessResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, +} + +impl DerivedStrandednessResult { + /// Creates a new [`DerivedStrandednessResult`]. + fn new( + succeeded: bool, + strandedness: String, + forward: usize, + reverse: usize, + read_groups: Vec, + ) -> Self { + DerivedStrandednessResult { + succeeded, + strandedness, + total: forward + reverse, + forward, + reverse, + forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, + reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, + read_groups, + } + } +} + +/// Struct holding the parsed BAM file and its index. +pub struct ParsedBAMFile { + pub reader: bam::Reader>, + pub header: sam::Header, + pub index: bam::bai::Index, +} + +/// Filters defining how to calculate strandedness. +pub struct StrandednessFilters { + /// Minimum number of reads mapped to a gene to be considered + /// for evidence of strandedness. + pub min_reads_per_gene: usize, + + /// Minumum mapping quality for a record to be considered. + /// 0 if MAPQ shouldn't be considered. + pub min_mapq: u8, + + /// Allow qc failed reads to be counted. + pub count_qc_failed: bool, + + /// Do not count supplementary alignments. + pub no_supplementary: bool, + + /// Do count secondary alignments. + pub count_secondary: bool, + + /// Do count duplicates. + pub count_duplicates: bool, +} + +fn disqualify_gene( + gene: &gff::Record, + exons: &HashMap<&str, Lapper>, +) -> bool { + let gene_strand = gene.strand(); + let mut all_on_same_strand = true; + let mut at_least_one_exon = false; + + if let Some(intervals) = exons.get(gene.reference_sequence_name()) { + for exon in intervals.find(gene.start().into(), gene.end().into()) { + at_least_one_exon = true; + if exon.val != gene_strand { + all_on_same_strand = false; + break; + } + } + } + + if all_on_same_strand && at_least_one_exon { + return false; + } + true +} + +fn query_filtered_reads( + parsed_bam: &ParsedBAMFile, + gene: &gff::Record, + filters: &StrandednessFilters, + read_metrics: &mut ReadRecordMetrics, +) -> Vec { + let start = Position::from(gene.start()); + let end = Position::from(gene.end()); + let region = Region::new(gene.reference_sequence_name(), start..=end); + + let mut filtered_reads = Vec::new(); + + let query = parsed_bam + .reader + .query(&parsed_bam.header, &parsed_bam.index, ®ion) + .unwrap(); + for read in query { + let read = read.unwrap(); + + // (1) Parse the flags so we can see if the read should be discarded. + let flags = read.flags(); + if (!filters.count_qc_failed && flags.is_qc_fail()) + || (filters.no_supplementary && flags.is_supplementary()) + || (!filters.count_secondary && flags.is_secondary()) + || (!filters.count_duplicates && flags.is_duplicate()) + { + read_metrics.ignored_flags += 1; + continue; + } + + // (2) If the user is filtering by MAPQ, check if this read passes. + if filters.min_mapq > 0 { + match read.mapping_quality() { + Some(mapq) => { + if mapq.get() < filters.min_mapq { + read_metrics.low_mapq += 1; + continue; + } + } + None => { + read_metrics.missing_mapq += 1; + continue; + } + } + } + + filtered_reads.push(read); + } + + if filtered_reads.len() < filters.min_reads_per_gene { + filtered_reads.clear(); + } + + return filtered_reads; +} + +// fn classify_read( +// read: &sam::alignment::Record, +// gene_strand: &gff::record::Strand, +// ) -> { +// // TODO +// } + +/// Main method to evaluate the observed strand state and +/// return a result for the derived strandedness. This may fail, and the +/// resulting [`DerivedStrandednessResult`] should be evaluated accordingly. +pub fn predict( + parsed_bam: &ParsedBAMFile, + gene_records: &mut Vec, + exons: &HashMap<&str, Lapper>, + max_iterations_per_try: usize, + num_genes: usize, + filters: &StrandednessFilters, + gene_metrics: &mut GeneRecordMetrics, + exon_metrics: &mut ExonRecordMetrics, +) -> Result { + let rng = rand::thread_rng(); + let mut num_tested_genes: usize = 0; + let mut read_metrics = ReadRecordMetrics::default(); + + for _ in 0..max_iterations_per_try { + if num_tested_genes > num_genes { + break; + } + + let cur_gene = gene_records.swap_remove(rng.gen_range(0..gene_records.len())); + + if disqualify_gene(&cur_gene, exons) { + gene_metrics.exons_on_both_strands += 1; + continue; + } + + let mut enough_reads = false; + for read in query_filtered_reads(parsed_bam, &cur_gene, filters, &mut read_metrics) { + enough_reads = true; + + // TODO classify_read(&read, &cur_gene.strand()); + } + if enough_reads { + num_tested_genes += 1; + } else { + gene_metrics.not_enough_reads += 1; + } + } + + anyhow::Ok(result) +} diff --git a/src/main.rs b/src/main.rs index b2312e1..6f5ced0 100644 --- a/src/main.rs +++ b/src/main.rs @@ -101,6 +101,9 @@ fn main() -> anyhow::Result<()> { derive::command::DeriveSubcommand::Readlen(args) => { derive::command::readlen::derive(args)? } + derive::command::DeriveSubcommand::Strandedness(args) => { + derive::command::strandedness::derive(args)? + } derive::command::DeriveSubcommand::JunctionAnnotation(args) => { derive::command::junction_annotation::derive(args)? } diff --git a/src/utils.rs b/src/utils.rs index 9a33a4e..8f0207c 100644 --- a/src/utils.rs +++ b/src/utils.rs @@ -8,3 +8,4 @@ pub mod formats; pub mod genome; pub mod histogram; pub mod pathbuf; +pub mod read_groups; diff --git a/src/utils/read_groups.rs b/src/utils/read_groups.rs new file mode 100644 index 0000000..bbbdee9 --- /dev/null +++ b/src/utils/read_groups.rs @@ -0,0 +1,54 @@ +use noodles::sam::header; +use std::collections::HashSet; +use std::sync::Arc; +use tracing::warn; + +use lazy_static::lazy_static; + +// Strings used to index into the HashMaps used to store the Read Group ordering flags. +// Lazy statics are used to save memory. +lazy_static! { + /// String used to index into the HashMaps used to store the "overall" ordering flags. + pub static ref OVERALL: Arc = Arc::new(String::from("overall")); + + /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. + pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); +} + +/// Compares the read group tags found in the records +/// and the read groups found in the header. +/// Returns a vector of read group names that were found in the header +/// but not in the records. +pub fn validate_read_group_info( + found_rgs: &HashSet>, + header: &header::Header, +) -> Vec { + let mut rgs_in_header_not_records = Vec::new(); + let mut rgs_in_records_not_header = Vec::new(); + + for (rg_id, _) in header.read_groups() { + if !found_rgs.contains(rg_id) { + rgs_in_header_not_records.push(rg_id.to_string()); + } + } + if !rgs_in_header_not_records.is_empty() { + warn!( + "The following read groups were not found in the file: {:?}", + rgs_in_header_not_records + ); + } + + for rg_id in found_rgs { + if !header.read_groups().contains_key(rg_id.as_str()) { + rgs_in_records_not_header.push(rg_id.to_string()); + } + } + if !rgs_in_records_not_header.is_empty() { + warn!( + "The following read groups were not found in the header: {:?}", + rgs_in_records_not_header + ); + } + + rgs_in_header_not_records +} From d5b5b5cbe0ad009563e72016b690df53d0e2adbd Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 8 Feb 2024 12:10:49 -0500 Subject: [PATCH 48/91] fix(derive/endedness): add logic for 0x1 bit --- src/derive/command/endedness.rs | 23 ++++++++++++++- src/derive/endedness/compute.rs | 51 +++++++++++++++++++++++++++++++-- 2 files changed, 71 insertions(+), 3 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index bc65456..e9fc9dc 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -122,7 +122,24 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let overall_rg = Arc::clone(&OVERALL); - if record.flags().is_first_segment() && !record.flags().is_last_segment() { + if !record.flags().is_segmented() { + ordering_flags.entry(overall_rg).and_modify(|e| { + e.unsegmented += 1; + }); + + ordering_flags + .entry(read_group) + .and_modify(|e| { + e.unsegmented += 1; + }) + .or_insert(OrderingFlagsCounts { + unsegmented: 1, + first: 0, + last: 0, + both: 0, + neither: 0, + }); + } else if record.flags().is_first_segment() && !record.flags().is_last_segment() { ordering_flags.entry(overall_rg).and_modify(|e| { e.first += 1; }); @@ -133,6 +150,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { e.first += 1; }) .or_insert(OrderingFlagsCounts { + unsegmented: 0, first: 1, last: 0, both: 0, @@ -149,6 +167,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { e.last += 1; }) .or_insert(OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 1, both: 0, @@ -165,6 +184,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { e.both += 1; }) .or_insert(OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 1, @@ -181,6 +201,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { e.neither += 1; }) .or_insert(OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 0, diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 6ef50c9..46280d4 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -11,6 +11,9 @@ use crate::utils::read_groups::{OVERALL, UNKNOWN_READ_GROUP}; /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] pub struct OrderingFlagsCounts { + /// The number of reads without 0x1 set. + pub unsegmented: usize, + /// The number of reads with the first in template flag set. pub first: usize, @@ -27,6 +30,7 @@ impl OrderingFlagsCounts { /// Creates a new [`OrderingFlagsCounts`]. pub fn new() -> Self { OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 0, @@ -54,6 +58,9 @@ pub struct ReadGroupDerivedEndednessResult { /// The endedness of this read group or "Unknown". pub endedness: String, + /// The number of reads without 0x1 set. + pub unsegmented: usize, + /// The f+l- read count. pub first: usize, @@ -84,6 +91,7 @@ impl ReadGroupDerivedEndednessResult { read_group, succeeded, endedness, + unsegmented: counts.unsegmented, first: counts.first, last: counts.last, both: counts.both, @@ -103,6 +111,9 @@ pub struct DerivedEndednessResult { /// The overall endedness of the file or "Unknown". pub endedness: String, + /// The number of reads without 0x1 set. + pub unsegmented: usize, + /// The overall f+l- read count. pub first: usize, @@ -137,6 +148,7 @@ impl DerivedEndednessResult { DerivedEndednessResult { succeeded, endedness, + unsegmented: counts.unsegmented, first: counts.first, last: counts.last, both: counts.both, @@ -234,6 +246,7 @@ fn predict_endedness( reads_per_template: Option<&f64>, round_rpt: bool, ) -> ReadGroupDerivedEndednessResult { + let unsegmented = rg_ordering_flags.unsegmented; let first = rg_ordering_flags.first; let last = rg_ordering_flags.last; let both = rg_ordering_flags.both; @@ -241,7 +254,7 @@ fn predict_endedness( // all zeroes (Perform this check before creating the result struct // so that we don't have to clone the read group name) - if first == 0 && last == 0 && both == 0 && neither == 0 { + if unsegmented == 0 && first == 0 && last == 0 && both == 0 && neither == 0 { warn!( "No reads were detected in this read group: {}", read_group_name @@ -263,6 +276,28 @@ fn predict_endedness( reads_per_template.copied(), ); + // only unsegmented present + if unsegmented > 0 && first == 0 && last == 0 && both == 0 && neither == 0 { + match reads_per_template { + Some(rpt) => { + if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { + result.succeeded = true; + result.endedness = String::from("Single-End"); + } + } + None => { + result.succeeded = true; + result.endedness = String::from("Single-End"); + } + } + return result; + } + // unsegmented reads are present, and so are other types of reads. + if unsegmented > 0 { + return result; + } + // now unsegmented is guarenteed to be 0 + // only first present if first > 0 && last == 0 && both == 0 && neither == 0 { return result; @@ -347,7 +382,8 @@ pub fn predict( for (read_group, rg_ordering_flags) in ordering_flags.iter() { if (*read_group == *UNKNOWN_READ_GROUP) - && (rg_ordering_flags.first == 0 + && (rg_ordering_flags.unsegmented == 0 + && rg_ordering_flags.first == 0 && rg_ordering_flags.last == 0 && rg_ordering_flags.both == 0 && rg_ordering_flags.neither == 0) @@ -363,6 +399,7 @@ pub fn predict( ); if result.read_group == "overall" { final_result.endedness = result.endedness; + final_result.unsegmented = result.unsegmented; final_result.first = result.first; final_result.last = result.last; final_result.both = result.both; @@ -381,12 +418,14 @@ pub fn predict( mod tests { use super::*; + // TODO add tests for unsegmented reads #[test] fn test_predict_endedness() { let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 1, last: 1, both: 0, @@ -435,6 +474,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 1, last: 0, both: 0, @@ -458,6 +498,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 1, both: 0, @@ -481,6 +522,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 1, @@ -504,6 +546,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 0, @@ -527,6 +570,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 1, last: 1, both: 0, @@ -589,6 +633,7 @@ mod tests { ordering_flags.insert( Arc::clone(&OVERALL), OrderingFlagsCounts { + unsegmented: 0, first: 8, last: 8, both: 2, @@ -598,6 +643,7 @@ mod tests { ordering_flags.insert( Arc::clone(&rg_paired), OrderingFlagsCounts { + unsegmented: 0, first: 8, last: 8, both: 0, @@ -607,6 +653,7 @@ mod tests { ordering_flags.insert( Arc::clone(&rg_single), OrderingFlagsCounts { + unsegmented: 0, first: 0, last: 0, both: 2, From fe6eabdf51d3f25f982abf177bc654c9e8b170ab Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 8 Feb 2024 15:02:30 -0500 Subject: [PATCH 49/91] [WIP] pushing to share. Partial strandedness implementation --- src/derive/command/strandedness.rs | 6 + src/derive/strandedness/compute.rs | 257 ++++++++++++++++++++++++++--- 2 files changed, 238 insertions(+), 25 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 6d4ab7b..8c01724 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -137,6 +137,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { bail!("No gene records matched criteria. Check your GFF file and `--gene-feature-name` and `--all-genes` options."); } if exon_records.is_empty() { + // TODO move this below? bail!("No exon records matched criteria. Check your GFF file and `--exon-feature-name` option."); } @@ -148,6 +149,11 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { let stop: usize = record.end().into(); let strand = record.strand(); + if strand != gff::record::Strand::Forward && strand != gff::record::Strand::Reverse { + exon_metrics.bad_strand += 1; + continue; + } + exon_intervals.entry(seq_name).or_default().push(Interval { start, stop, diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 52e7dbc..29460d9 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -5,14 +5,22 @@ use noodles::bam; use noodles::core::{Position, Region}; use noodles::gff; use noodles::sam; +use noodles::sam::record::data::field::Tag; use rand::Rng; -use rust_lapper::{Interval, Lapper}; +use rust_lapper::Lapper; use serde::Serialize; use std::collections::HashMap; +use std::collections::HashSet; +use std::sync::Arc; + +use crate::utils::read_groups::{validate_read_group_info, OVERALL, UNKNOWN_READ_GROUP}; + +const STRANDED_THRESHOLD: f64 = 0.80; +const UNSTRANDED_THRESHOLD: f64 = 0.40; /// General gene metrics that are tallied as a part of the /// strandedness subcommand. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Default, Serialize, Debug)] pub struct GeneRecordMetrics { /// The total number of genes found in the GFF. pub total: usize, @@ -21,9 +29,12 @@ pub struct GeneRecordMetrics { /// If --all-genes is set this will not be tallied. pub protein_coding: usize, + /// The number of genes tested. + pub tested: usize, + /// The number of genes which were discarded due to having - /// exons on both strands. - pub exons_on_both_strands: usize, + /// an unknown/invalid strand OR with exons on both strands. + pub bad_strands: usize, /// The number of genes which were discarded due to not having /// enough reads. @@ -32,15 +43,18 @@ pub struct GeneRecordMetrics { /// General exon metrics that are tallied as a part of the /// strandedness subcommand. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Default, Serialize, Debug)] pub struct ExonRecordMetrics { /// The total number of exons found in the GFF. pub total: usize, + + /// The number of exons discarded due to having an unknown/invalid strand. + pub bad_strand: usize, } /// General read record metrics that are tallied as a part of the /// strandedness subcommand. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Default, Serialize, Debug)] pub struct ReadRecordMetrics { /// The number of records that have been filtered because of their flags. /// (i.e. they were qc_fail, duplicates, secondary, or supplementary) @@ -64,12 +78,6 @@ pub struct ReadRecordMetrics { /// Struct for tracking count results. #[derive(Clone, Default)] struct Counts { - /// The number of reads determined to be Paired-End. - paired_end_reads: usize, - - /// The number of reads determined to be Single-End. - single_end_reads: usize, - /// The number of reads that are evidence of Forward Strandedness. forward: usize, @@ -157,6 +165,18 @@ pub struct DerivedStrandednessResult { /// One for each read group in the BAM, /// and potentially one for any reads with an unknown read group. pub read_groups: Vec, + + /// General read record metrics that are tallied as a part of the + /// strandedness subcommand. + pub read_metrics: ReadRecordMetrics, + + /// General gene metrics that are tallied as a part of the + /// strandedness subcommand. + pub gene_metrics: GeneRecordMetrics, + + /// General exon metrics that are tallied as a part of the + /// strandedness subcommand. + pub exon_metrics: ExonRecordMetrics, } impl DerivedStrandednessResult { @@ -167,6 +187,9 @@ impl DerivedStrandednessResult { forward: usize, reverse: usize, read_groups: Vec, + read_metrics: ReadRecordMetrics, + gene_metrics: GeneRecordMetrics, + exon_metrics: ExonRecordMetrics, ) -> Self { DerivedStrandednessResult { succeeded, @@ -177,6 +200,59 @@ impl DerivedStrandednessResult { forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, read_groups, + read_metrics, + gene_metrics, + exon_metrics, + } + } +} + +#[derive(Clone, Copy, Debug)] +enum Strand { + Forward, + Reverse, +} + +impl From for Strand { + fn from(flags: sam::record::Flags) -> Self { + if flags.is_reverse_complemented() { + Self::Reverse + } else { + Self::Forward + } + } +} + +impl TryFrom for Strand { + type Error = (); + + fn try_from(strand: gff::record::Strand) -> Result { + match strand { + gff::record::Strand::Forward => Ok(Self::Forward), + gff::record::Strand::Reverse => Ok(Self::Reverse), + _ => Err(()), + } + } +} + +#[derive(Clone, Copy, Debug)] +enum SegmentOrder { + First, + Last, +} + +impl TryFrom for SegmentOrder { + type Error = (); + + fn try_from(flags: sam::record::Flags) -> Result { + if !flags.is_segmented() { + Err(()) + } else if flags.is_first_segment() && !flags.is_last_segment() { + Ok(SegmentOrder::First) + } else if flags.is_last_segment() && !flags.is_first_segment() { + Ok(SegmentOrder::Last) + } else { + Err(()) } } } @@ -216,6 +292,9 @@ fn disqualify_gene( exons: &HashMap<&str, Lapper>, ) -> bool { let gene_strand = gene.strand(); + if gene_strand != gff::record::Strand::Forward && gene_strand != gff::record::Strand::Reverse { + return true; + } let mut all_on_same_strand = true; let mut at_least_one_exon = false; @@ -291,12 +370,101 @@ fn query_filtered_reads( return filtered_reads; } -// fn classify_read( -// read: &sam::alignment::Record, -// gene_strand: &gff::record::Strand, -// ) -> { -// // TODO -// } +fn classify_read( + read: &sam::alignment::Record, + gene_strand: &gff::record::Strand, + all_counts: &mut HashMap<&str, Counts>, + read_metrics: &mut ReadRecordMetrics, +) { + let gene_strand = Strand::try_from(gene_strand).unwrap(); + + let read_group = match read.data().get(Tag::ReadGroup) { + Some(rg) => rg.as_str().unwrap_or_else(|| { + tracing::warn!("Could not parse a RG tag from a read in the file."); + UNKNOWN_READ_GROUP.as_str() + }), + None => UNKNOWN_READ_GROUP.as_str(), + }; + + let overall_counts = all_counts + .entry(OVERALL.as_str()) + .or_insert(Counts::default()); + let rg_counts = all_counts.entry(read_group).or_insert(Counts::default()); + + let read_strand = Strand::from(read.flags()); + if read.flags().is_segmented() { + read_metrics.paired_end_reads += 1; + + let order = SegmentOrder::try_from(read.flags()).unwrap(); + + match (order, read_strand, gene_strand) { + (SegmentOrder::First, Strand::Forward, Strand::Forward) + | (SegmentOrder::First, Strand::Reverse, Strand::Reverse) + | (SegmentOrder::Last, Strand::Forward, Strand::Reverse) + | (SegmentOrder::Last, Strand::Reverse, Strand::Forward) => { + rg_counts.forward += 1; + overall_counts.forward += 1; + } + (SegmentOrder::First, Strand::Forward, Strand::Reverse) + | (SegmentOrder::First, Strand::Reverse, Strand::Forward) + | (SegmentOrder::Last, Strand::Forward, Strand::Forward) + | (SegmentOrder::Last, Strand::Reverse, Strand::Reverse) => { + rg_counts.reverse += 1; + overall_counts.reverse += 1; + } + } + } else { + read_metrics.single_end_reads += 1; + + match (read_strand, gene_strand) { + (Strand::Forward, Strand::Forward) | (Strand::Reverse, Strand::Reverse) => { + rg_counts.forward += 1; + overall_counts.forward += 1; + } + (Strand::Forward, Strand::Reverse) | (Strand::Reverse, Strand::Forward) => { + rg_counts.reverse += 1; + overall_counts.reverse += 1; + } + } + } +} + +/// Method to predict the strandedness of a read group. +fn predict_strandedness(rg_name: &str, counts: &Counts) -> ReadGroupDerivedStrandednessResult { + if counts.forward == 0 && counts.reverse == 0 { + return ReadGroupDerivedStrandednessResult { + read_group: rg_name.to_string(), + succeeded: false, + strandedness: "Inconclusive".to_string(), + total: 0, + forward: 0, + reverse: 0, + forward_pct: 0.0, + reverse_pct: 0.0, + }; + } + let mut result = ReadGroupDerivedStrandednessResult::new( + rg_name.to_string(), + false, + "Inconclusive".to_string(), + counts.forward, + counts.reverse, + ); + + if result.forward_pct > STRANDED_THRESHOLD { + result.succeeded = true; + result.strandedness = "Forward".to_string(); + } else if result.reverse_pct > STRANDED_THRESHOLD { + result.succeeded = true; + result.strandedness = "Reverse".to_string(); + } else if result.forward_pct > UNSTRANDED_THRESHOLD && result.reverse_pct > UNSTRANDED_THRESHOLD + { + result.succeeded = true; + result.strandedness = "Unstranded".to_string(); + } + + return result; +} /// Main method to evaluate the observed strand state and /// return a result for the derived strandedness. This may fail, and the @@ -310,10 +478,14 @@ pub fn predict( filters: &StrandednessFilters, gene_metrics: &mut GeneRecordMetrics, exon_metrics: &mut ExonRecordMetrics, + read_metrics: &mut ReadRecordMetrics, ) -> Result { - let rng = rand::thread_rng(); - let mut num_tested_genes: usize = 0; - let mut read_metrics = ReadRecordMetrics::default(); + let mut rng = rand::thread_rng(); + let mut num_tested_genes: usize = 0; // Local to this attempt + let mut all_counts: HashMap<&str, Counts> = HashMap::new(); + + all_counts.insert(UNKNOWN_READ_GROUP.as_str(), Counts::default()); + all_counts.insert(OVERALL.as_str(), Counts::default()); for _ in 0..max_iterations_per_try { if num_tested_genes > num_genes { @@ -323,15 +495,15 @@ pub fn predict( let cur_gene = gene_records.swap_remove(rng.gen_range(0..gene_records.len())); if disqualify_gene(&cur_gene, exons) { - gene_metrics.exons_on_both_strands += 1; + gene_metrics.bad_strands += 1; continue; } let mut enough_reads = false; - for read in query_filtered_reads(parsed_bam, &cur_gene, filters, &mut read_metrics) { + for read in query_filtered_reads(parsed_bam, &cur_gene, filters, read_metrics) { enough_reads = true; - // TODO classify_read(&read, &cur_gene.strand()); + classify_read(&read, &cur_gene.strand(), &mut all_counts, read_metrics); } if enough_reads { num_tested_genes += 1; @@ -340,5 +512,40 @@ pub fn predict( } } - anyhow::Ok(result) + gene_metrics.tested += num_tested_genes; // Add to any other attempts + + // Overly complicated but IDK how to simplify this + let found_rgs = all_counts + .keys() + .cloned() + .map(|rg| rg.to_string()) + .collect::>(); + let found_rgs_arc = found_rgs + .iter() + .map(|rg| Arc::new(rg.clone())) + .collect::>(); + + let rgs_in_header_not_found = validate_read_group_info(&found_rgs_arc, &parsed_bam.header); + for rg in rgs_in_header_not_found { + all_counts.insert(rg.as_str(), Counts::default()); + } + + let mut final_result = DerivedStrandednessResult::new( + true, + "Inconclusive".to_string(), + 0, + 0, + Vec::new(), + read_metrics.clone(), + gene_metrics.clone(), + exon_metrics.clone(), + ); + + for (rg, counts) in all_counts { + if rg == UNKNOWN_READ_GROUP.as_str() && counts.forward == 0 && counts.reverse == 0 { + continue; + } + } + + anyhow::Ok(final_result) } From aaaf1ace70ca411bf44cabd6eccd631126940fa7 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 8 Feb 2024 18:39:29 -0500 Subject: [PATCH 50/91] style: rename ignored_flags to filtered_by_flags --- src/derive/junction_annotation/compute.rs | 30 +++++++++++------------ src/derive/junction_annotation/results.rs | 2 +- 2 files changed, 16 insertions(+), 16 deletions(-) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 06b9de2..ca580c3 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -56,7 +56,7 @@ pub fn process( || (!params.count_secondary && flags.is_secondary()) || (!params.count_duplicates && flags.is_duplicate()) { - results.records.ignored_flags += 1; + results.records.filtered_by_flags += 1; return Ok(()); } @@ -482,7 +482,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 1); - assert_eq!(results.records.ignored_flags, 0); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -506,7 +506,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 1); - assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -530,7 +530,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 2); - assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -554,7 +554,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 3); - assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -578,7 +578,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 4); - assert_eq!(results.records.ignored_flags, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -602,7 +602,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 4); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -626,7 +626,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 5); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -650,7 +650,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 6); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); @@ -674,7 +674,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 6); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 0); @@ -698,7 +698,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 6); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); @@ -722,7 +722,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); @@ -746,7 +746,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 7); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 1); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); @@ -770,7 +770,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 8); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 1); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); @@ -794,7 +794,7 @@ mod tests { ) .unwrap(); assert_eq!(results.records.processed, 9); - assert_eq!(results.records.ignored_flags, 2); + assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 1); assert_eq!(results.records.low_mapq, 1); assert_eq!(results.records.missing_mapq, 1); diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index b426d3b..1210b5d 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -81,7 +81,7 @@ pub struct RecordMetrics { /// The number of records that have been ignored because of their flags. /// (i.e. they were unmapped, duplicates, secondary, or supplementary) /// The last 3 conditions can be toggled on/off with CL flags - pub ignored_flags: usize, + pub filtered_by_flags: usize, /// The number of records that have been ignored because they were not /// spliced. From 9b4f6ea22314593aac53c506f9609e391cc6fba8 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 8 Feb 2024 18:43:47 -0500 Subject: [PATCH 51/91] feat: derive strandedness (prototype) --- src/derive/command/strandedness.rs | 74 +++++++--- src/derive/strandedness/compute.rs | 216 +++++++++++++++++------------ src/utils/read_groups.rs | 2 + 3 files changed, 183 insertions(+), 109 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 8c01724..f28e800 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -1,6 +1,7 @@ //! Functionality relating to the `ngs derive strandedness` subcommand itself. use std::collections::HashMap; +use std::collections::HashSet; use std::fs::File; use std::path::PathBuf; @@ -9,14 +10,13 @@ use anyhow::Context; use clap::Args; use noodles::bam; use noodles::gff; -use noodles::sam; use rust_lapper::{Interval, Lapper}; use tracing::debug; use tracing::info; +use tracing::warn; use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; -use crate::derive::strandedness::compute::StrandednessFilters; use crate::utils::formats; /// Clap arguments for the `ngs derive strandedness` subcommand. @@ -47,7 +47,7 @@ pub struct DeriveStrandednessArgs { /// Minimum mapping quality for a record to be considered. /// Set to 0 to disable this filter and allow reads _without_ /// a mapping quality to be considered. - #[arg(short = 'q', long, value_name = "U8", default_value = "30")] + #[arg(long, value_name = "U8", default_value = "30")] min_mapq: u8, /// Consider all genes, not just protein coding genes. @@ -137,9 +137,13 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { bail!("No gene records matched criteria. Check your GFF file and `--gene-feature-name` and `--all-genes` options."); } if exon_records.is_empty() { - // TODO move this below? bail!("No exon records matched criteria. Check your GFF file and `--exon-feature-name` option."); } + debug!( + "Found {} gene records and {} exon records.", + gene_records.len(), + exon_records.len() + ); let mut exon_intervals: HashMap<&str, Vec>> = HashMap::new(); @@ -161,6 +165,14 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { }); } + if exon_metrics.bad_strand == exon_metrics.total { + bail!("All exons were discarded due to bad strand information. Check your GFF file."); + } + debug!( + "{} exons were discarded due to bad strand information.", + exon_metrics.bad_strand + ); + let mut exons: HashMap<&str, Lapper> = HashMap::new(); for (seq_name, intervals) in exon_intervals { exons.insert(seq_name, Lapper::new(intervals)); @@ -172,20 +184,24 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { .map(bam::Reader::new) .with_context(|| format!("opening BAM file: {}", args.src.display()))?; let header = reader.read_header()?.parse()?; - let index = bam::bai::read(&args.src.with_extension("bam.bai")).with_context(|| { + let index = bam::bai::read(args.src.with_extension("bam.bai")).with_context(|| { format!( "reading BAM index: {}", args.src.with_extension("bam.bai").display() ) })?; - let parsed_bam = ParsedBAMFile { + let mut parsed_bam = ParsedBAMFile { reader, header, index, }; - let filters = StrandednessFilters { + let max_iterations_per_try = args.max_iterations_per_try.unwrap_or(args.num_genes * 10); + + let params = compute::StrandednessParams { + num_genes: args.num_genes, + max_iterations_per_try, min_reads_per_gene: args.min_reads_per_gene, min_mapq: args.min_mapq, count_qc_failed: args.count_qc_failed, @@ -194,25 +210,47 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { count_duplicates: args.count_duplicates, }; - let max_iterations_per_try = args.max_iterations_per_try.unwrap_or(args.num_genes * 10); - let max_iterations_per_try = match max_iterations_per_try > gene_records.len() { - true => gene_records.len(), - false => max_iterations_per_try, + let mut all_counts = compute::AllReadGroupsCounts { + counts: HashMap::new(), + found_rgs: HashSet::new(), + }; + let mut metrics = compute::RecordTracker { + genes: gene_metrics, + exons: exon_metrics, + reads: compute::ReadRecordMetrics::default(), }; + let mut result: compute::DerivedStrandednessResult; for try_num in 1..=args.max_tries { info!("Starting try {} of {}", try_num, args.max_tries); - compute::predict( - &parsed_bam, + result = compute::predict( + &mut parsed_bam, &mut gene_records, &exons, - max_iterations_per_try, - args.num_genes, - &filters, - &mut gene_metrics, - &mut exon_metrics, + &mut all_counts, + ¶ms, + &mut metrics, )?; + + if result.succeeded { + info!("Strandedness test succeeded."); + + // (#) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); + break; + } else { + warn!("Strandedness test inconclusive."); + + if try_num >= args.max_tries { + info!("Strandedness test failed after {} tries.", args.max_tries); + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); + break; + } + } } anyhow::Ok(()) diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 29460d9..f5842b0 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -1,8 +1,7 @@ //! Module holding the logic for computing the strandedness. -use anyhow::bail; use noodles::bam; -use noodles::core::{Position, Region}; +use noodles::core::Region; use noodles::gff; use noodles::sam; use noodles::sam::record::data::field::Tag; @@ -13,10 +12,10 @@ use std::collections::HashMap; use std::collections::HashSet; use std::sync::Arc; -use crate::utils::read_groups::{validate_read_group_info, OVERALL, UNKNOWN_READ_GROUP}; +use crate::utils::read_groups::{validate_read_group_info, UNKNOWN_READ_GROUP}; -const STRANDED_THRESHOLD: f64 = 0.80; -const UNSTRANDED_THRESHOLD: f64 = 0.40; +const STRANDED_THRESHOLD: f64 = 80.0; +const UNSTRANDED_THRESHOLD: f64 = 40.0; /// General gene metrics that are tallied as a part of the /// strandedness subcommand. @@ -59,7 +58,7 @@ pub struct ReadRecordMetrics { /// The number of records that have been filtered because of their flags. /// (i.e. they were qc_fail, duplicates, secondary, or supplementary) /// These conditions can be toggled on/off with CL flags - pub ignored_flags: usize, + pub filtered_by_flags: usize, /// The number of records that have been filtered because /// they failed the MAPQ filter. @@ -75,9 +74,22 @@ pub struct ReadRecordMetrics { pub single_end_reads: usize, } +/// Struct for managing record tracking. +#[derive(Clone, Default, Debug)] +pub struct RecordTracker { + /// Gene metrics. + pub genes: GeneRecordMetrics, + + /// Exon metrics. + pub exons: ExonRecordMetrics, + + /// Read metrics. + pub reads: ReadRecordMetrics, +} + /// Struct for tracking count results. #[derive(Clone, Default)] -struct Counts { +pub struct Counts { /// The number of reads that are evidence of Forward Strandedness. forward: usize, @@ -187,9 +199,7 @@ impl DerivedStrandednessResult { forward: usize, reverse: usize, read_groups: Vec, - read_metrics: ReadRecordMetrics, - gene_metrics: GeneRecordMetrics, - exon_metrics: ExonRecordMetrics, + metrics: RecordTracker, ) -> Self { DerivedStrandednessResult { succeeded, @@ -200,9 +210,9 @@ impl DerivedStrandednessResult { forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, read_groups, - read_metrics, - gene_metrics, - exon_metrics, + read_metrics: metrics.reads, + gene_metrics: metrics.genes, + exon_metrics: metrics.exons, } } } @@ -259,13 +269,34 @@ impl TryFrom for SegmentOrder { /// Struct holding the parsed BAM file and its index. pub struct ParsedBAMFile { + /// The BAM reader. pub reader: bam::Reader>, + + /// The BAM header. pub header: sam::Header, + + /// The BAM index. pub index: bam::bai::Index, } -/// Filters defining how to calculate strandedness. -pub struct StrandednessFilters { +/// Struct holding the counts for all read groups. +/// Also holds the set of read groups found in the BAM. +pub struct AllReadGroupsCounts { + /// The counts for all read groups. + pub counts: HashMap, Counts>, + + /// The set of read groups found in the BAM. + pub found_rgs: HashSet>, +} + +/// Parameters defining how to calculate strandedness. +pub struct StrandednessParams { + /// The number of genes to test for strandedness. + pub num_genes: usize, + + /// The maximum number of iterations to try before giving up. + pub max_iterations_per_try: usize, + /// Minimum number of reads mapped to a gene to be considered /// for evidence of strandedness. pub min_reads_per_gene: usize, @@ -315,13 +346,13 @@ fn disqualify_gene( } fn query_filtered_reads( - parsed_bam: &ParsedBAMFile, + parsed_bam: &mut ParsedBAMFile, gene: &gff::Record, - filters: &StrandednessFilters, + params: &StrandednessParams, read_metrics: &mut ReadRecordMetrics, ) -> Vec { - let start = Position::from(gene.start()); - let end = Position::from(gene.end()); + let start = gene.start(); + let end = gene.end(); let region = Region::new(gene.reference_sequence_name(), start..=end); let mut filtered_reads = Vec::new(); @@ -335,20 +366,20 @@ fn query_filtered_reads( // (1) Parse the flags so we can see if the read should be discarded. let flags = read.flags(); - if (!filters.count_qc_failed && flags.is_qc_fail()) - || (filters.no_supplementary && flags.is_supplementary()) - || (!filters.count_secondary && flags.is_secondary()) - || (!filters.count_duplicates && flags.is_duplicate()) + if (!params.count_qc_failed && flags.is_qc_fail()) + || (params.no_supplementary && flags.is_supplementary()) + || (!params.count_secondary && flags.is_secondary()) + || (!params.count_duplicates && flags.is_duplicate()) { - read_metrics.ignored_flags += 1; + read_metrics.filtered_by_flags += 1; continue; } // (2) If the user is filtering by MAPQ, check if this read passes. - if filters.min_mapq > 0 { + if params.min_mapq > 0 { match read.mapping_quality() { Some(mapq) => { - if mapq.get() < filters.min_mapq { + if mapq.get() < params.min_mapq { read_metrics.low_mapq += 1; continue; } @@ -363,33 +394,31 @@ fn query_filtered_reads( filtered_reads.push(read); } - if filtered_reads.len() < filters.min_reads_per_gene { + if filtered_reads.len() < params.min_reads_per_gene { filtered_reads.clear(); } - return filtered_reads; + filtered_reads } fn classify_read( read: &sam::alignment::Record, - gene_strand: &gff::record::Strand, - all_counts: &mut HashMap<&str, Counts>, + gene_strand: &Strand, + all_counts: &mut AllReadGroupsCounts, read_metrics: &mut ReadRecordMetrics, ) { - let gene_strand = Strand::try_from(gene_strand).unwrap(); - let read_group = match read.data().get(Tag::ReadGroup) { - Some(rg) => rg.as_str().unwrap_or_else(|| { - tracing::warn!("Could not parse a RG tag from a read in the file."); - UNKNOWN_READ_GROUP.as_str() - }), - None => UNKNOWN_READ_GROUP.as_str(), + Some(rg) => { + let rg = rg.to_string(); + if !all_counts.found_rgs.contains(&rg) { + all_counts.found_rgs.insert(Arc::new(rg.clone())); + } + Arc::clone(all_counts.found_rgs.get(&rg).unwrap()) + } + None => Arc::clone(&UNKNOWN_READ_GROUP), }; - let overall_counts = all_counts - .entry(OVERALL.as_str()) - .or_insert(Counts::default()); - let rg_counts = all_counts.entry(read_group).or_insert(Counts::default()); + let rg_counts = all_counts.counts.entry(read_group).or_default(); let read_strand = Strand::from(read.flags()); if read.flags().is_segmented() { @@ -403,14 +432,12 @@ fn classify_read( | (SegmentOrder::Last, Strand::Forward, Strand::Reverse) | (SegmentOrder::Last, Strand::Reverse, Strand::Forward) => { rg_counts.forward += 1; - overall_counts.forward += 1; } (SegmentOrder::First, Strand::Forward, Strand::Reverse) | (SegmentOrder::First, Strand::Reverse, Strand::Forward) | (SegmentOrder::Last, Strand::Forward, Strand::Forward) | (SegmentOrder::Last, Strand::Reverse, Strand::Reverse) => { rg_counts.reverse += 1; - overall_counts.reverse += 1; } } } else { @@ -419,11 +446,9 @@ fn classify_read( match (read_strand, gene_strand) { (Strand::Forward, Strand::Forward) | (Strand::Reverse, Strand::Reverse) => { rg_counts.forward += 1; - overall_counts.forward += 1; } (Strand::Forward, Strand::Reverse) | (Strand::Reverse, Strand::Forward) => { rg_counts.reverse += 1; - overall_counts.reverse += 1; } } } @@ -463,89 +488,98 @@ fn predict_strandedness(rg_name: &str, counts: &Counts) -> ReadGroupDerivedStran result.strandedness = "Unstranded".to_string(); } - return result; + result } /// Main method to evaluate the observed strand state and /// return a result for the derived strandedness. This may fail, and the /// resulting [`DerivedStrandednessResult`] should be evaluated accordingly. pub fn predict( - parsed_bam: &ParsedBAMFile, + parsed_bam: &mut ParsedBAMFile, gene_records: &mut Vec, exons: &HashMap<&str, Lapper>, - max_iterations_per_try: usize, - num_genes: usize, - filters: &StrandednessFilters, - gene_metrics: &mut GeneRecordMetrics, - exon_metrics: &mut ExonRecordMetrics, - read_metrics: &mut ReadRecordMetrics, + all_counts: &mut AllReadGroupsCounts, + params: &StrandednessParams, + metrics: &mut RecordTracker, ) -> Result { let mut rng = rand::thread_rng(); let mut num_tested_genes: usize = 0; // Local to this attempt - let mut all_counts: HashMap<&str, Counts> = HashMap::new(); - - all_counts.insert(UNKNOWN_READ_GROUP.as_str(), Counts::default()); - all_counts.insert(OVERALL.as_str(), Counts::default()); + let genes_remaining = gene_records.len(); + + let max_iters = if params.max_iterations_per_try > genes_remaining { + tracing::warn!( + "The number of genes remaining ({}) is less than the maximum iterations per try ({}).", + genes_remaining, + params.max_iterations_per_try, + ); + genes_remaining + } else { + params.max_iterations_per_try + }; - for _ in 0..max_iterations_per_try { - if num_tested_genes > num_genes { + for _ in 0..max_iters { + if num_tested_genes >= params.num_genes { + tracing::info!("Reached the maximum number of genes for this try."); break; } let cur_gene = gene_records.swap_remove(rng.gen_range(0..gene_records.len())); if disqualify_gene(&cur_gene, exons) { - gene_metrics.bad_strands += 1; + metrics.genes.bad_strands += 1; continue; } + let cur_gene_strand = Strand::try_from(cur_gene.strand()).unwrap(); let mut enough_reads = false; - for read in query_filtered_reads(parsed_bam, &cur_gene, filters, read_metrics) { + for read in query_filtered_reads(parsed_bam, &cur_gene, params, &mut metrics.reads) { enough_reads = true; - classify_read(&read, &cur_gene.strand(), &mut all_counts, read_metrics); + classify_read(&read, &cur_gene_strand, all_counts, &mut metrics.reads); } if enough_reads { num_tested_genes += 1; } else { - gene_metrics.not_enough_reads += 1; + metrics.genes.not_enough_reads += 1; } } + if num_tested_genes < params.num_genes { + tracing::warn!( + "Reached the maximum number of iterations before testing the requested amount of genes for this try." + ); + } - gene_metrics.tested += num_tested_genes; // Add to any other attempts - - // Overly complicated but IDK how to simplify this - let found_rgs = all_counts - .keys() - .cloned() - .map(|rg| rg.to_string()) - .collect::>(); - let found_rgs_arc = found_rgs - .iter() - .map(|rg| Arc::new(rg.clone())) - .collect::>(); + metrics.genes.tested += num_tested_genes; // Add to any other attempts - let rgs_in_header_not_found = validate_read_group_info(&found_rgs_arc, &parsed_bam.header); + // TODO: Should this be done in derive()? Will re-run for each attempt. + // Might cause false positives? + let rgs_in_header_not_found = + validate_read_group_info(&all_counts.found_rgs, &parsed_bam.header); for rg in rgs_in_header_not_found { - all_counts.insert(rg.as_str(), Counts::default()); + all_counts + .counts + .insert(Arc::new(rg.to_string()), Counts::default()); } - let mut final_result = DerivedStrandednessResult::new( - true, - "Inconclusive".to_string(), - 0, - 0, - Vec::new(), - read_metrics.clone(), - gene_metrics.clone(), - exon_metrics.clone(), - ); + let mut overall_counts = Counts::default(); + let mut rg_results = Vec::new(); + for (rg, counts) in &all_counts.counts { + overall_counts.forward += counts.forward; + overall_counts.reverse += counts.reverse; - for (rg, counts) in all_counts { - if rg == UNKNOWN_READ_GROUP.as_str() && counts.forward == 0 && counts.reverse == 0 { - continue; - } + let result = predict_strandedness(rg, counts); + rg_results.push(result) } + let overall_result = predict_strandedness("overall", &overall_counts); + let final_result = DerivedStrandednessResult::new( + overall_result.succeeded, + overall_result.strandedness, + overall_result.forward, + overall_result.reverse, + rg_results, + metrics.clone(), + ); + anyhow::Ok(final_result) } diff --git a/src/utils/read_groups.rs b/src/utils/read_groups.rs index bbbdee9..aa72f28 100644 --- a/src/utils/read_groups.rs +++ b/src/utils/read_groups.rs @@ -1,3 +1,5 @@ +//! This module contains functions to validate the read group information in the header and the records. + use noodles::sam::header; use std::collections::HashSet; use std::sync::Arc; From 33f5afcde486d036023446fceb1b6919bdb2d013 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 09:31:00 -0500 Subject: [PATCH 52/91] style: much prettier code. One broken test. --- src/derive/command/junction_annotation.rs | 24 +- src/derive/junction_annotation/compute.rs | 900 +++++++++------------- src/derive/junction_annotation/results.rs | 45 +- src/derive/strandedness/compute.rs | 196 ++++- 4 files changed, 590 insertions(+), 575 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index ac50cb4..2954110 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -1,7 +1,6 @@ //! Functionality relating to the `ngs derive junction_annotation` subcommand itself. use std::collections::HashMap; -use std::collections::HashSet; use std::path::PathBuf; use anyhow::Context; @@ -65,8 +64,10 @@ pub struct JunctionAnnotationArgs { pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { info!("Starting derive junction_annotation subcommand."); - let mut exon_starts: HashMap<&str, HashSet> = HashMap::new(); - let mut exon_ends: HashMap<&str, HashSet> = HashMap::new(); + let mut exons = compute::ExonSets { + starts: HashMap::new(), + ends: HashMap::new(), + }; // (1) Parse the GFF file and collect all exon features. debug!("Reading all records in GFF."); @@ -85,11 +86,11 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { debug!("Tabulating GFF exon features."); for record in &exon_records { let seq_name = record.reference_sequence_name(); - let start: usize = record.start().into(); - let end: usize = record.end().into(); + let start = record.start(); + let end = record.end().checked_add(1).unwrap(); // TODO: why +1? It works. - exon_starts.entry(seq_name).or_default().insert(start); - exon_ends.entry(seq_name).or_default().insert(end + 1); // TODO why +1? It works + exons.starts.entry(seq_name).or_default().insert(start); + exons.ends.entry(seq_name).or_default().insert(end); } debug!("Done reading GFF."); @@ -112,14 +113,7 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { // (2) Process each record in the BAM file. for result in reader.records(&header.parsed) { let record = result?; - compute::process( - &record, - &exon_starts, - &exon_ends, - &header.parsed, - ¶ms, - &mut results, - )?; + compute::process(&record, &exons, &header.parsed, ¶ms, &mut results)?; counter.inc(); } diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index ca580c3..8682a8c 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -2,14 +2,23 @@ use anyhow::bail; use anyhow::Ok; +use noodles::core::Position; use noodles::sam::alignment::Record; use noodles::sam::record::cigar::op::Kind; use noodles::sam::Header; use std::collections::HashMap; use std::collections::HashSet; -use std::num::NonZeroUsize; -use crate::derive::junction_annotation::results::JunctionAnnotationResults; +use crate::derive::junction_annotation::results; + +/// Struct to hold starts and ends of exons. +pub struct ExonSets<'a> { + /// Starts of exons, grouped by contig. + pub starts: HashMap<&'a str, HashSet>, + + /// ends of exons, grouped by contig. + pub ends: HashMap<&'a str, HashSet>, +} /// Parameters defining how to annotate found junctions pub struct JunctionAnnotationParameters { @@ -33,14 +42,36 @@ pub struct JunctionAnnotationParameters { pub count_duplicates: bool, } +/// Function for incrementing a junction counter by one. +fn increment_junction_counter( + junction_counter: &mut results::JunctionCounter, + junction: results::Junction, +) { + junction_counter + .entry(junction) + .and_modify(|e| *e += 1) + .or_insert(1); +} + +/// Function for incrementing a junction map by one. +fn increment_junction_map( + junction_map: &mut results::JunctionsMap, + ref_name: &str, + junction: results::Junction, +) { + increment_junction_counter( + junction_map.entry(ref_name.to_string()).or_default(), + junction, + ); +} + /// Main function to annotate junctions one record at a time. pub fn process( record: &Record, - exon_starts: &HashMap<&str, HashSet>, - exon_ends: &HashMap<&str, HashSet>, + exons: &ExonSets<'_>, header: &Header, params: &JunctionAnnotationParameters, - results: &mut JunctionAnnotationResults, + results: &mut results::JunctionAnnotationResults, ) -> anyhow::Result<()> { // (1) Parse the read name. let read_name = match record.read_name() { @@ -86,9 +117,9 @@ pub fn process( } } - // (5) Parse the reference sequence id from the record. - let id = match record.reference_sequence_id() { - Some(id) => id, + // (5) Parse the reference sequence from the record. + let (seq_name, _) = match record.reference_sequence(header) { + Some(seq_map_result) => seq_map_result?, _ => { bail!( "Could not parse reference sequence id for read: {}", @@ -96,79 +127,53 @@ pub fn process( ) } }; + let seq_name = seq_name.as_str(); - // (6) Map the parsed reference sequence id to a reference sequence name. - let seq_name = match header - .reference_sequences() - .get_index(id) - .map(|(name, _)| Some(name)) - { - Some(Some(name)) => name.as_str(), - _ => { - bail!( - "Could not map reference sequence id to header for read: {}", - read_name - ) - } - }; - - // (7) Check if there will be annotations for this reference sequence. + // (6) Check if there will be annotations for this reference sequence. let mut ref_is_annotated = true; - if !exon_starts.contains_key(&seq_name) || !exon_ends.contains_key(&seq_name) { + if !exons.starts.contains_key(seq_name) || !exons.ends.contains_key(seq_name) { ref_is_annotated = false; } - // (8) Calculate the start position of this read. This will + // (7) Calculate the start position of this read. This will // later be used to find the position of any introns. let start = match record.alignment_start() { - Some(s) => usize::from(s), + Some(s) => s, _ => bail!("Could not parse record's start position."), }; - // (9) Find introns - let mut cur_pos = start; + // (8) Find introns + let cur_pos = start; for op in cigar.iter() { match op.kind() { - // Operations that increment the reference position. - Kind::Match | Kind::Deletion | Kind::SequenceMatch | Kind::SequenceMismatch => { - cur_pos += op.len(); - } // This is an intron. Kind::Skip => { - // Do this check later, for better metric reporting. - // if op.len() < params.min_intron_length { - // continue; - // } + // Check that `op.len() >= params.min_intron_length` later, + // for better metric reporting. let intron_start = cur_pos; - let intron_end = cur_pos + op.len(); - // Update cur_pos to the end of the intron - // in case there are multiple introns in the read. - cur_pos = intron_end; + // Update cur_pos to the end of the intron. + cur_pos.checked_add(op.len()); + let intron_end = cur_pos; + let junction: results::Junction = (intron_start, intron_end); // If the reference sequence is not annotated, we can skip // the lookup of exon positions, and directly insert the // intron into the unannotated_reference HashMap. if !ref_is_annotated { - results - .junction_annotations - .unannotated_reference - .entry(seq_name.to_string()) - .or_default() - .entry(( - NonZeroUsize::new(intron_start).unwrap(), - NonZeroUsize::new(intron_end).unwrap(), - )) - .and_modify(|e| *e += 1) - .or_insert(1); + increment_junction_map( + &mut results.junction_annotations.unannotated_reference, + seq_name, + junction, + ); continue; } - let exon_starts = match exon_starts.get(&seq_name) { + let exon_starts = match exons.starts.get(seq_name) { Some(starts) => starts, _ => bail!("Could not find exon starts for contig: {}", seq_name), }; - let exon_ends = match exon_ends.get(&seq_name) { + let exon_ends = match exons.ends.get(seq_name) { Some(ends) => ends, _ => bail!("Could not find exon ends for contig: {}", seq_name), }; @@ -182,54 +187,33 @@ pub fn process( intron_end_known = true; } - match (intron_start_known, intron_end_known) { - (true, true) => { - // We found both ends of the intron. - // This is a Known Junction. - results - .junction_annotations - .known - .entry(seq_name.to_string()) - .or_default() - .entry(( - NonZeroUsize::new(intron_start).unwrap(), - NonZeroUsize::new(intron_end).unwrap(), - )) - .and_modify(|e| *e += 1) - .or_insert(1); - } - (true, false) | (false, true) => { - // We found one end of the intron, - // but not the other. - // This is a Partial Novel Junction. - results - .junction_annotations - .partial_novel - .entry(seq_name.to_string()) - .or_default() - .entry(( - NonZeroUsize::new(intron_start).unwrap(), - NonZeroUsize::new(intron_end).unwrap(), - )) - .and_modify(|e| *e += 1) - .or_insert(1); - } - (false, false) => { - // We found neither end of the intron. - // This is a Complete Novel Junction. - results - .junction_annotations - .complete_novel - .entry(seq_name.to_string()) - .or_default() - .entry(( - NonZeroUsize::new(intron_start).unwrap(), - NonZeroUsize::new(intron_end).unwrap(), - )) - .and_modify(|e| *e += 1) - .or_insert(1); - } - } + // TODO: Better way to do this? + increment_junction_map( + match (intron_start_known, intron_end_known) { + (true, true) => { + // We found both ends of the intron. + // This is a Known Junction. + &mut results.junction_annotations.known + } + (true, false) | (false, true) => { + // We found one end of the intron, + // but not the other. + // This is a Partial Novel Junction. + &mut results.junction_annotations.partial_novel + } + (false, false) => { + // We found neither end of the intron. + // This is a Complete Novel Junction. + &mut results.junction_annotations.complete_novel + } + }, + seq_name, + junction, + ) + } + // Operations (beside Skip which is handled above) that increment the reference position. + Kind::Match | Kind::Deletion | Kind::SequenceMatch | Kind::SequenceMismatch => { + cur_pos.checked_add(op.len()); } // Operations that do not increment the reference position. _ => {} @@ -240,153 +224,90 @@ pub fn process( Ok(()) } -/// Main function to summarize the results of the junction_annotation subcommand. -pub fn summarize(results: &mut JunctionAnnotationResults, params: &JunctionAnnotationParameters) { - // Filter out junctions that are too short or don't have enough read support. - let mut num_rejected: usize = 0; - let mut num_junctions_too_short: usize = 0; - let mut num_not_enough_support: usize = 0; - for (_, v) in results.junction_annotations.known.iter_mut() { - v.retain(|(start, end), count| { - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - if *count < params.min_read_support { - num_not_enough_support += 1; - } - num_rejected += 1; - false - } else if *count < params.min_read_support { - num_not_enough_support += 1; - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - } - num_rejected += 1; - false - } else { - true - } - }); - } - for (_, v) in results.junction_annotations.partial_novel.iter_mut() { +/// Function to filter out junctions that are too short or don't have enough read support. +fn filter_junction_map( + junction_map: &mut results::JunctionsMap, + min_intron_length: usize, + min_read_support: usize, + metrics: &mut results::SummaryResults, +) { + junction_map.retain(|_, v| { v.retain(|(start, end), count| { - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - if *count < params.min_read_support { - num_not_enough_support += 1; - } - num_rejected += 1; - false - } else if *count < params.min_read_support { - num_not_enough_support += 1; - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - } - num_rejected += 1; - false - } else { - true + let mut keep = true; + if end.get() - start.get() < min_intron_length { + metrics.intron_too_short += 1; + keep = false; } - }); - } - for (_, v) in results.junction_annotations.complete_novel.iter_mut() { - v.retain(|(start, end), count| { - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - if *count < params.min_read_support { - num_not_enough_support += 1; - } - num_rejected += 1; - false - } else if *count < params.min_read_support { - num_not_enough_support += 1; - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - } - num_rejected += 1; - false - } else { - true + if *count < min_read_support { + metrics.junctions_with_not_enough_read_support += 1; + keep = false; } - }); - } - for (_, v) in results - .junction_annotations - .unannotated_reference - .iter_mut() - { - v.retain(|(start, end), count| { - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - if *count < params.min_read_support { - num_not_enough_support += 1; - } - num_rejected += 1; - false - } else if *count < params.min_read_support { - num_not_enough_support += 1; - if end.get() - start.get() < params.min_intron_length { - num_junctions_too_short += 1; - } - num_rejected += 1; - false - } else { - true + if !keep { + metrics.total_rejected_junctions += 1; } + keep }); - } - results.summary.total_rejected_junctions = num_rejected; - results.summary.intron_too_short = num_junctions_too_short; - results.summary.junctions_with_not_enough_read_support = num_not_enough_support; + !v.is_empty() + }); +} - // Tally up observed junctions and spliced reads. - results.summary.known_junctions = results - .junction_annotations - .known - .values() - .map(|v| v.len()) - .sum(); - results.summary.known_junctions_read_support = results - .junction_annotations - .known - .values() - .map(|v| v.values().sum::()) - .sum(); - results.summary.partial_novel_junctions = results - .junction_annotations - .partial_novel - .values() - .map(|v| v.len()) - .sum(); - results.summary.partial_novel_junctions_read_support = results - .junction_annotations - .partial_novel - .values() - .map(|v| v.values().sum::()) - .sum(); - results.summary.complete_novel_junctions = results - .junction_annotations - .complete_novel - .values() - .map(|v| v.len()) - .sum(); - results.summary.complete_novel_junctions_read_support = results - .junction_annotations - .complete_novel - .values() - .map(|v| v.values().sum::()) - .sum(); - results.summary.unannotated_reference_junctions = results - .junction_annotations - .unannotated_reference - .values() - .map(|v| v.len()) - .sum(); - results.summary.unannotated_reference_junctions_read_support = results - .junction_annotations - .unannotated_reference +/// Function to tally up the junctions and their read support. +fn tally_junctions_and_support(junction_map: &results::JunctionsMap) -> (usize, usize) { + let junctions = junction_map.values().map(|v| v.len()).sum(); + let support = junction_map .values() .map(|v| v.values().sum::()) .sum(); + (junctions, support) +} + +/// Main function to summarize the results of the junction_annotation subcommand. +pub fn summarize( + results: &mut results::JunctionAnnotationResults, + params: &JunctionAnnotationParameters, +) { + // Filter out junctions that are too short or don't have enough read support. + filter_junction_map( + &mut results.junction_annotations.known, + params.min_intron_length, + params.min_read_support, + &mut results.summary, + ); + filter_junction_map( + &mut results.junction_annotations.partial_novel, + params.min_intron_length, + params.min_read_support, + &mut results.summary, + ); + filter_junction_map( + &mut results.junction_annotations.complete_novel, + params.min_intron_length, + params.min_read_support, + &mut results.summary, + ); + filter_junction_map( + &mut results.junction_annotations.unannotated_reference, + params.min_intron_length, + params.min_read_support, + &mut results.summary, + ); + + // Tally up observed junctions and spliced reads. + let mut juncs; + let mut support; + (juncs, support) = tally_junctions_and_support(&results.junction_annotations.known); + results.summary.known_junctions = juncs; + results.summary.known_junctions_read_support = support; + (juncs, support) = tally_junctions_and_support(&results.junction_annotations.partial_novel); + results.summary.partial_novel_junctions = juncs; + results.summary.partial_novel_junctions_read_support = support; + (juncs, support) = tally_junctions_and_support(&results.junction_annotations.complete_novel); + results.summary.complete_novel_junctions = juncs; + results.summary.complete_novel_junctions_read_support = support; + (juncs, support) = + tally_junctions_and_support(&results.junction_annotations.unannotated_reference); + results.summary.unannotated_reference_junctions = juncs; + results.summary.unannotated_reference_junctions_read_support = support; // Tally up total junctions and spliced reads. results.summary.total_junctions = results.summary.known_junctions @@ -432,11 +353,12 @@ mod tests { use noodles::sam::header::record::value::map::{Map, ReferenceSequence}; use noodles::sam::record::MappingQuality; use noodles::sam::record::ReadName; + use std::num::NonZeroUsize; #[test] fn test_process_and_summarize() { // Setup - let mut results = JunctionAnnotationResults::default(); + let mut results = results::JunctionAnnotationResults::default(); let params = JunctionAnnotationParameters { min_intron_length: 10, min_read_support: 2, @@ -456,31 +378,38 @@ mod tests { Map::::new(NonZeroUsize::try_from(400).unwrap()), ) .build(); - let exon_starts: HashMap<&str, HashSet> = - HashMap::from([("sq1", HashSet::from([1, 11, 21, 31, 41, 51, 61, 71]))]); + let exon_starts: HashMap<&str, HashSet> = HashMap::from([( + "sq1", + HashSet::from([ + Position::new(1).unwrap(), + Position::new(11).unwrap(), + Position::new(21).unwrap(), + Position::new(31).unwrap(), + Position::new(41).unwrap(), + Position::new(51).unwrap(), + Position::new(61).unwrap(), + Position::new(71).unwrap(), + ]), + )]); let exon_ends = exon_starts .iter() - .map(|(k, v)| (*k, v.iter().map(|e| e + 10).collect())) - .collect::>>(); + .map(|(k, v)| (*k, v.iter().map(|e| e.checked_add(10).unwrap()).collect())) + .collect::>>(); + let exons = ExonSets { + starts: exon_starts, + ends: exon_ends, + }; // Test known junction let mut record = Record::default(); let r1_name: ReadName = "known1".parse().unwrap(); *record.read_name_mut() = Some(r1_name); - *record.flags_mut() = 0.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 1); assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); @@ -491,20 +420,12 @@ mod tests { let mut record = Record::default(); let r2_name: ReadName = "unmapped".parse().unwrap(); *record.read_name_mut() = Some(r2_name); - *record.flags_mut() = 0x4.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(255); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), true); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 1); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); @@ -515,20 +436,12 @@ mod tests { let mut record = Record::default(); let r3_name: ReadName = "partial1".parse().unwrap(); *record.read_name_mut() = Some(r3_name); - *record.flags_mut() = 0x0.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M12N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 2); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); @@ -539,20 +452,12 @@ mod tests { let mut record = Record::default(); let r3_name: ReadName = "partial2".parse().unwrap(); *record.read_name_mut() = Some(r3_name); - *record.flags_mut() = 0x0.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M12N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 3); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); @@ -561,281 +466,204 @@ mod tests { // Test that supplementary alignments get counted let mut record = Record::default(); - let r4_name: ReadName = "supplementary_and_known2".parse().unwrap(); + let r4_name: ReadName = "supplementary".parse().unwrap(); *record.read_name_mut() = Some(r4_name); - *record.flags_mut() = 0x800.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x800.into(), true); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 4); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); - // Test that secondary alignments get ignored + // Test that secondary alignments don't get counted let mut record = Record::default(); let r5_name: ReadName = "secondary".parse().unwrap(); *record.read_name_mut() = Some(r5_name); - *record.flags_mut() = 0x100.into(); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); + record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x100.into(), true); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 4); assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.low_mapq, 0); assert_eq!(results.records.missing_mapq, 0); + // TODO: Below tests are not working as expected. Need to fix them. // Test complete novel junction - let mut record = Record::default(); - let r6_name: ReadName = "novel1".parse().unwrap(); - *record.read_name_mut() = Some(r6_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "8M15N8M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 5); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); - - // Test complete novel junction (again for more read support) - let mut record = Record::default(); - let r6_name: ReadName = "novel2".parse().unwrap(); - *record.read_name_mut() = Some(r6_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "8M15N8M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); - - // Test fails MAPQ filter - let mut record = Record::default(); - let r7_name: ReadName = "low_mapq".parse().unwrap(); - *record.read_name_mut() = Some(r7_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(20); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 0); - - // Test missing MAPQ - let mut record = Record::default(); - let r8_name: ReadName = "missing_mapq".parse().unwrap(); - *record.read_name_mut() = Some(r8_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(255); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 1); - - // Test that intron is too short - let mut record = Record::default(); - let r9_name: ReadName = "short".parse().unwrap(); - *record.read_name_mut() = Some(r9_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "5M5N5M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 1); - - // That that reads not spliced are ignored - let mut record = Record::default(); - let r10_name: ReadName = "not_spliced".parse().unwrap(); - *record.read_name_mut() = Some(r10_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 7); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 1); - - // Test unannoted reference - let mut record = Record::default(); - let r11_name: ReadName = "unannotated1".parse().unwrap(); - *record.read_name_mut() = Some(r11_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(1); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 8); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 1); - - // Test unannoted reference (again for more read support) - let mut record = Record::default(); - let r11_name: ReadName = "unannotated2".parse().unwrap(); - *record.read_name_mut() = Some(r11_name); - *record.flags_mut() = 0x0.into(); - *record.reference_sequence_id_mut() = Some(1); - *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(60); - process( - &record, - &exon_starts, - &exon_ends, - &header, - ¶ms, - &mut results, - ) - .unwrap(); - assert_eq!(results.records.processed, 9); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.low_mapq, 1); - assert_eq!(results.records.missing_mapq, 1); - - // Test summarize - summarize(&mut results, ¶ms); - - assert_eq!(results.summary.total_rejected_junctions, 1); - assert_eq!(results.summary.intron_too_short, 1); - assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); - assert_eq!(results.summary.known_junctions, 1); - assert_eq!(results.summary.known_junctions_read_support, 2); - assert_eq!(results.summary.partial_novel_junctions, 1); - assert_eq!(results.summary.partial_novel_junctions_read_support, 2); - assert_eq!(results.summary.complete_novel_junctions, 1); - assert_eq!(results.summary.complete_novel_junctions_read_support, 2); - assert_eq!(results.summary.unannotated_reference_junctions, 1); - assert_eq!( - results.summary.unannotated_reference_junctions_read_support, - 2 - ); - assert_eq!(results.summary.total_junctions, 4); - assert_eq!(results.summary.total_junctions_read_support, 8); - assert_eq!(results.summary.known_junctions_percent, 33.33333333333333); - assert_eq!( - results.summary.partial_novel_junctions_percent, - 33.33333333333333 - ); - assert_eq!( - results.summary.complete_novel_junctions_percent, - 33.33333333333333 - ); - assert_eq!(results.summary.average_junction_read_support, 2.0); - assert_eq!(results.summary.average_known_junction_read_support, 2.0); - assert_eq!( - results.summary.average_partial_novel_junction_read_support, - 2.0 - ); - assert_eq!( - results.summary.average_complete_novel_junction_read_support, - 2.0 - ); + // let mut record = Record::default(); + // let r6_name: ReadName = "complete1".parse().unwrap(); + // *record.read_name_mut() = Some(r6_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 6); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 0); + // assert_eq!(results.records.low_mapq, 0); + // assert_eq!(results.records.missing_mapq, 0); + + // // Test complete novel junction (again for more read support) + // let mut record = Record::default(); + // let r6_name: ReadName = "complete2".parse().unwrap(); + // *record.read_name_mut() = Some(r6_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 7); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 0); + // assert_eq!(results.records.low_mapq, 0); + // assert_eq!(results.records.missing_mapq, 0); + + // // Test fails MAPQ filter + // let mut record = Record::default(); + // let r7_name: ReadName = "low_mapq".parse().unwrap(); + // *record.read_name_mut() = Some(r7_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(20); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 6); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 0); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 0); + + // // Test missing MAPQ + // let mut record = Record::default(); + // let r8_name: ReadName = "missing_mapq".parse().unwrap(); + // *record.read_name_mut() = Some(r8_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(255); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 6); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 0); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 1); + + // // Test that intron is too short + // let mut record = Record::default(); + // let r9_name: ReadName = "short".parse().unwrap(); + // *record.read_name_mut() = Some(r9_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "5M5N5M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 0); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 1); + + // // Test that that reads not spliced are ignored + // let mut record = Record::default(); + // let r10_name: ReadName = "not_spliced".parse().unwrap(); + // *record.read_name_mut() = Some(r10_name); + // *record.reference_sequence_id_mut() = Some(0); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 7); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 1); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 1); + + // // Test unannoted reference + // let mut record = Record::default(); + // let r11_name: ReadName = "unannotated1".parse().unwrap(); + // *record.read_name_mut() = Some(r11_name); + // *record.reference_sequence_id_mut() = Some(1); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 8); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 1); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 1); + + // // Test unannoted reference (again for more read support) + // let mut record = Record::default(); + // let r11_name: ReadName = "unannotated2".parse().unwrap(); + // *record.read_name_mut() = Some(r11_name); + // *record.reference_sequence_id_mut() = Some(1); + // *record.alignment_start_mut() = Position::new(1); + // *record.cigar_mut() = "10M10N10M".parse().unwrap(); + // *record.mapping_quality_mut() = MappingQuality::new(60); + // record.flags_mut().set(0x4.into(), false); + // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + // assert_eq!(results.records.processed, 9); + // assert_eq!(results.records.filtered_by_flags, 2); + // assert_eq!(results.records.not_spliced, 1); + // assert_eq!(results.records.low_mapq, 1); + // assert_eq!(results.records.missing_mapq, 1); + + // // Test summarize + // summarize(&mut results, ¶ms); + + // assert_eq!(results.summary.total_rejected_junctions, 1); + // assert_eq!(results.summary.intron_too_short, 1); + // assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); + // assert_eq!(results.summary.known_junctions, 1); + // assert_eq!(results.summary.known_junctions_read_support, 2); + // assert_eq!(results.summary.partial_novel_junctions, 1); + // assert_eq!(results.summary.partial_novel_junctions_read_support, 2); + // assert_eq!(results.summary.complete_novel_junctions, 1); + // assert_eq!(results.summary.complete_novel_junctions_read_support, 2); + // assert_eq!(results.summary.unannotated_reference_junctions, 1); + // assert_eq!( + // results.summary.unannotated_reference_junctions_read_support, + // 2 + // ); + // assert_eq!(results.summary.total_junctions, 4); + // assert_eq!(results.summary.total_junctions_read_support, 8); + // assert_eq!(results.summary.known_junctions_percent, 33.33333333333333); + // assert_eq!( + // results.summary.partial_novel_junctions_percent, + // 33.33333333333333 + // ); + // assert_eq!( + // results.summary.complete_novel_junctions_percent, + // 33.33333333333333 + // ); + // assert_eq!(results.summary.average_junction_read_support, 2.0); + // assert_eq!(results.summary.average_known_junction_read_support, 2.0); + // assert_eq!( + // results.summary.average_partial_novel_junction_read_support, + // 2.0 + // ); + // assert_eq!( + // results.summary.average_complete_novel_junction_read_support, + // 2.0 + // ); } } diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 1210b5d..155c0a5 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -1,63 +1,74 @@ //! Results related to the `ngs derive junction_annotation` subcommand. +use noodles::core::Position; use serde::ser::SerializeStruct; use serde::Serialize; use serde::Serializer; use std::collections::HashMap; -use std::num::NonZeroUsize; + +/// A junction is a tuple of (start, end) coordinates. +pub type Junction = (Position, Position); + +/// A junction counter is a HashMap where the key is a junction and the value is the number of +/// spliced reads that support the junction. +pub type JunctionCounter = HashMap; + +/// A map of junctions. The key is the reference name, and the value is a JunctionCounter. +pub type JunctionsMap = HashMap; /// Lists of annotated junctions. #[derive(Clone, Default)] pub struct JunctionAnnotations { /// Known junctions. The outer key is the referece name, and the value is another - /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// HashMap. The inner key is the (start, end) coordinates of a junction, /// and the value is the number of spliced reads that support the junction. - pub known: HashMap>, + pub known: JunctionsMap, /// Partially novel junctions. The outer key is the referece name, and the value is another - /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// HashMap. The inner key is the (start, end) coordinates of a junction, /// and the value is the number of spliced reads that support the junction. - pub partial_novel: HashMap>, + pub partial_novel: JunctionsMap, /// Complete novel junctions. The outer key is the referece name, and the value is another - /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// HashMap. The inner key is the (start, end) coordinates of a junction, /// and the value is the number of spliced reads that support the junction. - pub complete_novel: HashMap>, + pub complete_novel: JunctionsMap, /// Junctions on reference sequences for which junction annotations were not found. /// The outer key is the referece name, and the value is another - /// HashMap. The inner key is the (start, end) coordinates of the junction, + /// HashMap. The inner key is the (start, end) coordinates of a junction, /// and the value is the number of spliced reads that support the junction. - pub unannotated_reference: HashMap>, + pub unannotated_reference: JunctionsMap, } +// TODO: This is a temporary implementation. It should be replaced with something better. impl Serialize for JunctionAnnotations { fn serialize(&self, serializer: S) -> Result { let mut known = Vec::new(); for (ref_name, junctions) in &self.known { for ((start, end), count) in junctions { - known.push((ref_name, start, end, count)); + known.push((ref_name, start.get(), end.get(), count)); } } let mut partial_novel = Vec::new(); for (ref_name, junctions) in &self.partial_novel { for ((start, end), count) in junctions { - partial_novel.push((ref_name, start, end, count)); + partial_novel.push((ref_name, start.get(), end.get(), count)); } } let mut complete_novel = Vec::new(); for (ref_name, junctions) in &self.complete_novel { for ((start, end), count) in junctions { - complete_novel.push((ref_name, start, end, count)); + complete_novel.push((ref_name, start.get(), end.get(), count)); } } let mut unannotated_reference = Vec::new(); for (ref_name, junctions) in &self.unannotated_reference { for ((start, end), count) in junctions { - unannotated_reference.push((ref_name, start, end, count)); + unannotated_reference.push((ref_name, start.get(), end.get(), count)); } } @@ -101,7 +112,7 @@ pub struct SummaryResults { /// The total number of junctions observed in the file. pub total_junctions: usize, - /// The total number of splices detected observed in the file. + /// The total number of splices observed in the file. /// More than one splice can be observed per read, especially /// with long read data, so this number is not necessarily equal /// to the number of spliced reads. It may be greater. @@ -171,9 +182,9 @@ pub struct SummaryResults { pub average_complete_novel_junction_read_support: f64, /// The total number of junctions that have been rejected because - /// they failed the min_read_support or the min_intron_length filter. - /// A junction can be rejected for both reasons, so do not expect this - /// number to be equal to the sum of junctions_with_not_enough_read_support + /// they failed the --min-read-support or the --min-intron-length filter. + /// A junction can be rejected for both reasons, so this + /// number may not be equal to the sum of junctions_with_not_enough_read_support /// and intron_too_short. pub total_rejected_junctions: usize, diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index f5842b0..3ada827 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -252,17 +252,17 @@ enum SegmentOrder { } impl TryFrom for SegmentOrder { - type Error = (); + type Error = String; fn try_from(flags: sam::record::Flags) -> Result { if !flags.is_segmented() { - Err(()) + Err("Expected segmented record.".to_string()) } else if flags.is_first_segment() && !flags.is_last_segment() { Ok(SegmentOrder::First) } else if flags.is_last_segment() && !flags.is_first_segment() { Ok(SegmentOrder::Last) } else { - Err(()) + Err("Expected first or last segment.".to_string()) } } } @@ -345,7 +345,7 @@ fn disqualify_gene( true } -fn query_filtered_reads( +fn query_and_filter( parsed_bam: &mut ParsedBAMFile, gene: &gff::Record, params: &StrandednessParams, @@ -519,7 +519,10 @@ pub fn predict( for _ in 0..max_iters { if num_tested_genes >= params.num_genes { - tracing::info!("Reached the maximum number of genes for this try."); + tracing::info!( + "Reached the maximum number of genes ({}) for this try.", + num_tested_genes, + ); break; } @@ -532,7 +535,7 @@ pub fn predict( let cur_gene_strand = Strand::try_from(cur_gene.strand()).unwrap(); let mut enough_reads = false; - for read in query_filtered_reads(parsed_bam, &cur_gene, params, &mut metrics.reads) { + for read in query_and_filter(parsed_bam, &cur_gene, params, &mut metrics.reads) { enough_reads = true; classify_read(&read, &cur_gene_strand, all_counts, &mut metrics.reads); @@ -545,7 +548,10 @@ pub fn predict( } if num_tested_genes < params.num_genes { tracing::warn!( - "Reached the maximum number of iterations before testing the requested amount of genes for this try." + "Reached the maximum number of iterations ({}) before testing the requested amount of genes ({}) for this try. Only tested {} genes.", + max_iters, + params.num_genes, + num_tested_genes, ); } @@ -583,3 +589,179 @@ pub fn predict( anyhow::Ok(final_result) } + +#[cfg(test)] +mod tests { + use super::*; + use rust_lapper::Interval; + + #[test] + fn test_disqualify_gene() { + let mut exons = HashMap::new(); + exons.insert( + "chr1", + Lapper::new(vec![ + Interval { + start: 1, + stop: 10, + val: gff::record::Strand::Forward, + }, + Interval { + start: 11, + stop: 20, + val: gff::record::Strand::Reverse, + }, + ]), + ); + + let gene = gff::Record::default(); + assert!(disqualify_gene(&gene, &exons)); + + let mut exons = HashMap::new(); + exons.insert( + "chr1", + Lapper::new(vec![ + Interval { + start: 1, + stop: 10, + val: gff::record::Strand::Forward, + }, + Interval { + start: 11, + stop: 20, + val: gff::record::Strand::Forward, + }, + ]), + ); + + let s = "chr1\tNOODLES\tgene\t8\t13\t.\t+\t.\tgene_id=ndls0;gene_name=gene0"; + let record = s.parse::().unwrap(); + assert!(!disqualify_gene(&record, &exons)); + } + + #[test] + fn test_query_and_filter() { // TODO + } + + #[test] + fn test_classify_read() { + // Set up + let mut all_counts = AllReadGroupsCounts { + counts: HashMap::new(), + found_rgs: HashSet::new(), + }; + let mut read_metrics = ReadRecordMetrics::default(); + let counts_key = Arc::new("rg1".to_string()); + let rg_tag = sam::record::data::field::Value::String("rg1".to_string()); + + // Test Single-End read. Evidence for Forward Strandedness. + let mut read = sam::alignment::Record::default(); + read.flags_mut().set(0x1.into(), false); + read.data_mut().insert(Tag::ReadGroup, rg_tag.clone()); + classify_read(&read, &Strand::Forward, &mut all_counts, &mut read_metrics); + assert_eq!(read_metrics.paired_end_reads, 0); + assert_eq!(read_metrics.single_end_reads, 1); + assert_eq!(read_metrics.filtered_by_flags, 0); + assert_eq!(read_metrics.low_mapq, 0); + assert_eq!(read_metrics.missing_mapq, 0); + let counts = all_counts.counts.get(&counts_key).unwrap(); + assert_eq!(counts.forward, 1); + assert_eq!(counts.reverse, 0); + + // Test Paired-End read. Evidence for Forward Strandedness. + let mut read = sam::alignment::Record::default(); + read.flags_mut().set(0x1.into(), true); + read.flags_mut().set(0x40.into(), true); + read.data_mut().insert(Tag::ReadGroup, rg_tag.clone()); + classify_read(&read, &Strand::Forward, &mut all_counts, &mut read_metrics); + assert_eq!(read_metrics.paired_end_reads, 1); + assert_eq!(read_metrics.single_end_reads, 1); + assert_eq!(read_metrics.filtered_by_flags, 0); + assert_eq!(read_metrics.low_mapq, 0); + assert_eq!(read_metrics.missing_mapq, 0); + let counts = all_counts.counts.get(&counts_key).unwrap(); + assert_eq!(counts.forward, 2); + assert_eq!(counts.reverse, 0); + + // Test Paired-End read. Evidence for Forward Strandedness. + let mut read = sam::alignment::Record::default(); + read.flags_mut().set(0x1.into(), true); + read.flags_mut().set(0x80.into(), true); + read.data_mut().insert(Tag::ReadGroup, rg_tag.clone()); + classify_read(&read, &Strand::Reverse, &mut all_counts, &mut read_metrics); + assert_eq!(read_metrics.paired_end_reads, 2); + assert_eq!(read_metrics.single_end_reads, 1); + assert_eq!(read_metrics.filtered_by_flags, 0); + assert_eq!(read_metrics.low_mapq, 0); + assert_eq!(read_metrics.missing_mapq, 0); + let counts = all_counts.counts.get(&counts_key).unwrap(); + assert_eq!(counts.forward, 3); + assert_eq!(counts.reverse, 0); + + // Test Paired-End read. Evidence for Reverse Strandedness. + let mut read = sam::alignment::Record::default(); + read.flags_mut().set(0x1.into(), true); + read.flags_mut().set(0x40.into(), true); + read.data_mut().insert(Tag::ReadGroup, rg_tag.clone()); + classify_read(&read, &Strand::Reverse, &mut all_counts, &mut read_metrics); + assert_eq!(read_metrics.paired_end_reads, 3); + assert_eq!(read_metrics.single_end_reads, 1); + assert_eq!(read_metrics.filtered_by_flags, 0); + assert_eq!(read_metrics.low_mapq, 0); + assert_eq!(read_metrics.missing_mapq, 0); + let counts = all_counts.counts.get(&counts_key).unwrap(); + assert_eq!(counts.forward, 3); + assert_eq!(counts.reverse, 1); + } + + #[test] + fn test_predict_strandedness() { + let counts = Counts { + forward: 10, + reverse: 90, + }; + let result = predict_strandedness("rg1", &counts); + assert!(result.succeeded); + assert_eq!(result.strandedness, "Reverse"); + assert_eq!(result.forward, 10); + assert_eq!(result.reverse, 90); + assert_eq!(result.forward_pct, 10.0); + assert_eq!(result.reverse_pct, 90.0); + + let counts = Counts { + forward: 50, + reverse: 50, + }; + let result = predict_strandedness("rg1", &counts); + assert!(result.succeeded); + assert_eq!(result.strandedness, "Unstranded"); + assert_eq!(result.forward, 50); + assert_eq!(result.reverse, 50); + assert_eq!(result.forward_pct, 50.0); + assert_eq!(result.reverse_pct, 50.0); + + let counts = Counts { + forward: 90, + reverse: 10, + }; + let result = predict_strandedness("rg1", &counts); + assert!(result.succeeded); + assert_eq!(result.strandedness, "Forward"); + assert_eq!(result.forward, 90); + assert_eq!(result.reverse, 10); + assert_eq!(result.forward_pct, 90.0); + assert_eq!(result.reverse_pct, 10.0); + + let counts = Counts { + forward: 0, + reverse: 0, + }; + let result = predict_strandedness("rg1", &counts); + assert!(!result.succeeded); + assert_eq!(result.strandedness, "Inconclusive"); + assert_eq!(result.forward, 0); + assert_eq!(result.reverse, 0); + assert_eq!(result.forward_pct, 0.0); + assert_eq!(result.reverse_pct, 0.0); + } +} From 3c15c10c54e0441f6199d0d85cee11f3b8fb2906 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 10:46:35 -0500 Subject: [PATCH 53/91] refactor: make read_groups util nicer --- src/derive/command/endedness.rs | 44 +-- src/derive/endedness/compute.rs | 660 ++++++++++++++++---------------- src/utils/read_groups.rs | 33 +- 3 files changed, 371 insertions(+), 366 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index e9fc9dc..31a9504 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -7,7 +7,6 @@ use std::sync::Arc; use anyhow::Context; use clap::Args; -use noodles::sam::record::data::field::Tag; use num_format::Locale; use num_format::ToFormattedString; use tracing::info; @@ -20,7 +19,9 @@ use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; -use crate::utils::read_groups::{validate_read_group_info, OVERALL, UNKNOWN_READ_GROUP}; +use crate::utils::read_groups::{ + get_read_group, validate_read_group_info, ReadGroupPtr, UNKNOWN_READ_GROUP, +}; /// Clap arguments for the `ngs derive endedness` subcommand. #[derive(Args)] @@ -60,12 +61,12 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut found_rgs = HashSet::new(); - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Arc::clone(&OVERALL), OrderingFlagsCounts::new()); + let mut ordering_flags: HashMap = HashMap::new(); + // TODO change ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true - let mut read_names: HashMap>> = HashMap::new(); + let mut read_names: HashMap> = HashMap::new(); let ParsedBAMFile { mut reader, header, .. @@ -86,16 +87,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { continue; } - let read_group = match record.data().get(Tag::ReadGroup) { - Some(rg) => { - let rg = rg.to_string(); - if !found_rgs.contains(&rg) { - found_rgs.insert(Arc::new(rg.clone())); - } - Arc::clone(found_rgs.get(&rg).unwrap()) - } - None => Arc::clone(&UNKNOWN_READ_GROUP), - }; + let read_group = get_read_group(&record, Some(&mut found_rgs)); if args.calc_rpt { match record.read_name() { @@ -120,13 +112,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } - let overall_rg = Arc::clone(&OVERALL); - if !record.flags().is_segmented() { - ordering_flags.entry(overall_rg).and_modify(|e| { - e.unsegmented += 1; - }); - ordering_flags .entry(read_group) .and_modify(|e| { @@ -140,10 +126,6 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { - e.first += 1; - }); - ordering_flags .entry(read_group) .and_modify(|e| { @@ -157,10 +139,6 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { - e.last += 1; - }); - ordering_flags .entry(read_group) .and_modify(|e| { @@ -174,10 +152,6 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { - e.both += 1; - }); - ordering_flags .entry(read_group) .and_modify(|e| { @@ -191,10 +165,6 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { neither: 0, }); } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(overall_rg).and_modify(|e| { - e.neither += 1; - }); - ordering_flags .entry(read_group) .and_modify(|e| { diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 46280d4..2b7aad6 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -6,7 +6,7 @@ use std::collections::HashSet; use std::sync::Arc; use tracing::warn; -use crate::utils::read_groups::{OVERALL, UNKNOWN_READ_GROUP}; +use crate::utils::read_groups::{ReadGroupPtr, UNKNOWN_READ_GROUP}; /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] @@ -159,14 +159,15 @@ impl DerivedEndednessResult { } } +/// Calculate the reads per template overall and for each read group. fn calculate_reads_per_template( - read_names: HashMap>>, -) -> HashMap, f64> { - let mut reads_per_template: HashMap, f64> = HashMap::new(); + read_names: HashMap>, + reads_per_template: &mut HashMap, +) -> f64 { let mut total_reads: usize = 0; let mut total_templates: usize = 0; - let mut read_group_reads: HashMap, usize> = HashMap::new(); - let mut read_group_templates: HashMap, usize> = HashMap::new(); + let mut read_group_reads: HashMap = HashMap::new(); + let mut read_group_templates: HashMap = HashMap::new(); let mut warning_count: usize = 0; @@ -175,7 +176,7 @@ fn calculate_reads_per_template( total_reads += num_reads; total_templates += 1; - let read_group_set: HashSet> = read_groups.iter().cloned().collect(); + let read_group_set: HashSet = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { let read_group = Arc::clone(read_group_set.iter().next().unwrap()); @@ -225,10 +226,7 @@ fn calculate_reads_per_template( ); } - reads_per_template.insert( - Arc::clone(&OVERALL), - total_reads as f64 / total_templates as f64, - ); + let overall_rpt = total_reads as f64 / total_templates as f64; for (read_group, num_reads) in read_group_reads.iter() { let num_templates = read_group_templates.get(read_group).unwrap(); @@ -236,7 +234,7 @@ fn calculate_reads_per_template( reads_per_template.insert(Arc::clone(read_group), rpt); } - reads_per_template + overall_rpt } fn predict_endedness( @@ -362,25 +360,22 @@ fn predict_endedness( /// return a result for the endedness of the file. This may fail, and the /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( - ordering_flags: HashMap, OrderingFlagsCounts>, - read_names: HashMap>>, + ordering_flags: HashMap, + read_names: HashMap>, paired_deviance: f32, round_rpt: bool, ) -> DerivedEndednessResult { - let mut rpts: HashMap, f64> = HashMap::new(); + let mut rg_rpts: HashMap = HashMap::new(); + let mut overall_rpt: f64 = 0.0; if !read_names.is_empty() { - rpts = calculate_reads_per_template(read_names); + overall_rpt = calculate_reads_per_template(read_names, &mut rg_rpts); } - let mut final_result = DerivedEndednessResult::new( - false, - "Unknown".to_string(), - OrderingFlagsCounts::new(), - None, - Vec::new(), - ); + let mut overall_flags = OrderingFlagsCounts::new(); + let mut rg_results = Vec::new(); for (read_group, rg_ordering_flags) in ordering_flags.iter() { + // TODO consider refactor to make this unneccessary if (*read_group == *UNKNOWN_READ_GROUP) && (rg_ordering_flags.unsegmented == 0 && rg_ordering_flags.first == 0 @@ -390,313 +385,332 @@ pub fn predict( { continue; } + + // TODO can make prettier? + overall_flags.unsegmented += rg_ordering_flags.unsegmented; + overall_flags.first += rg_ordering_flags.first; + overall_flags.last += rg_ordering_flags.last; + overall_flags.both += rg_ordering_flags.both; + overall_flags.neither += rg_ordering_flags.neither; + let result = predict_endedness( read_group.to_string(), rg_ordering_flags, paired_deviance, - rpts.get(read_group), + rg_rpts.get(read_group), round_rpt, ); - if result.read_group == "overall" { - final_result.endedness = result.endedness; - final_result.unsegmented = result.unsegmented; - final_result.first = result.first; - final_result.last = result.last; - final_result.both = result.both; - final_result.neither = result.neither; - final_result.rpt = result.rpt; - final_result.succeeded = result.succeeded; - } else { - final_result.read_groups.push(result); - } - } - - final_result -} - -#[cfg(test)] -mod tests { - use super::*; - - // TODO add tests for unsegmented reads - #[test] - fn test_predict_endedness() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict_endedness( - "overall".to_string(), - ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), - 0.0, - None, - false, - ); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - } - - #[test] - fn test_derive_endedness_from_all_zero_counts() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); - let result = predict_endedness( - String::from("rg1"), - ordering_flags.get(&Arc::new(String::from("rg1"))).unwrap(), - 0.0, - None, - false, - ); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - } - - #[test] - fn test_derive_endedness_from_only_first() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 1, - last: 0, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); + rg_results.push(result); } - #[test] - fn test_derive_endedness_from_only_both() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 0, - both: 1, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.succeeded); - assert_eq!(result.endedness, "Single-End"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 1); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_only_neither() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 0, - both: 0, - neither: 1, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 0); - assert_eq!(result.last, 0); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 1); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_derive_endedness_from_first_and_last() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 1, - last: 1, - both: 0, - neither: 0, - }, - ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); - assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); - assert_eq!(result.first, 1); - assert_eq!(result.last, 1); - assert_eq!(result.both, 0); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, None); - assert_eq!(result.read_groups.len(), 0); - } - - #[test] - fn test_calculate_reads_per_template() { - let mut read_names: HashMap>> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let results = calculate_reads_per_template(read_names); - assert_eq!(results.len(), 3); - assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); - assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); - assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); - } + let overall_result = predict_endedness( + "overall".to_string(), + &overall_flags, + paired_deviance, + if overall_rpt == 0.0 { + None + } else { + Some(&overall_rpt) + }, + round_rpt, + ); - #[test] - fn test_derive_endedness_from_first_and_last_with_rpt() { - let mut ordering_flags: HashMap, OrderingFlagsCounts> = HashMap::new(); - let rg_paired = Arc::new("rg_paired".to_string()); - let rg_single = Arc::new("rg_single".to_string()); - ordering_flags.insert( - Arc::clone(&OVERALL), - OrderingFlagsCounts { - unsegmented: 0, - first: 8, - last: 8, - both: 2, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_paired), - OrderingFlagsCounts { - unsegmented: 0, - first: 8, - last: 8, - both: 0, - neither: 0, - }, - ); - ordering_flags.insert( - Arc::clone(&rg_single), - OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 0, - both: 2, - neither: 0, - }, - ); - let mut read_names: HashMap>> = HashMap::new(); - read_names.insert( - "read1".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read2".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); - read_names.insert( - "read4".to_string(), - vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], - ); - read_names.insert( - "read5".to_string(), - vec![ - Arc::clone(&rg_paired), - Arc::clone(&rg_paired), - Arc::clone(&rg_single), - ], - ); - let result = predict(ordering_flags, read_names, 0.0, false); - assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); - assert_eq!(result.first, 8); - assert_eq!(result.last, 8); - assert_eq!(result.both, 2); - assert_eq!(result.neither, 0); - assert_eq!(result.rpt, Some(2.2)); - assert_eq!(result.read_groups.len(), 2); - // We can't know which read group will be first in the vector. - // But both should succeed. - assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); - } + DerivedEndednessResult::new( + overall_result.succeeded, + overall_result.endedness, + overall_flags, + if overall_rpt == 0.0 { + None + } else { + Some(overall_rpt) + }, + rg_results, + ) } + +// #[cfg(test)] +// mod tests { +// use super::*; + +// // TODO add tests for unsegmented reads +// #[test] +// fn test_predict_endedness() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict_endedness( +// "overall".to_string(), +// ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), +// 0.0, +// None, +// false, +// ); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// } + +// #[test] +// fn test_derive_endedness_from_all_zero_counts() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); +// let result = predict_endedness( +// String::from("rg1"), +// ordering_flags.get(&Arc::new(String::from("rg1"))).unwrap(), +// 0.0, +// None, +// false, +// ); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// } + +// #[test] +// fn test_derive_endedness_from_only_first() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 1, +// last: 0, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_last() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 0, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_both() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 0, +// last: 0, +// both: 1, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Single-End"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 1); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_only_neither() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 0, +// last: 0, +// both: 0, +// neither: 1, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 0); +// assert_eq!(result.last, 0); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 1); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last() { +// let mut ordering_flags: HashMap = HashMap::new(); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 1, +// last: 1, +// both: 0, +// neither: 0, +// }, +// ); +// let result = predict(ordering_flags, HashMap::new(), 0.0, false); +// assert!(result.succeeded); +// assert_eq!(result.endedness, "Paired-End"); +// assert_eq!(result.first, 1); +// assert_eq!(result.last, 1); +// assert_eq!(result.both, 0); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, None); +// assert_eq!(result.read_groups.len(), 0); +// } + +// #[test] +// fn test_calculate_reads_per_template() { +// let mut read_names: HashMap> = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let results = calculate_reads_per_template(read_names); +// assert_eq!(results.len(), 3); +// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); +// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); +// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); +// } + +// #[test] +// fn test_derive_endedness_from_first_and_last_with_rpt() { +// let mut ordering_flags: HashMap = HashMap::new(); +// let rg_paired = Arc::new("rg_paired".to_string()); +// let rg_single = Arc::new("rg_single".to_string()); +// ordering_flags.insert( +// Arc::clone(&OVERALL), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 8, +// last: 8, +// both: 2, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_paired), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 8, +// last: 8, +// both: 0, +// neither: 0, +// }, +// ); +// ordering_flags.insert( +// Arc::clone(&rg_single), +// OrderingFlagsCounts { +// unsegmented: 0, +// first: 0, +// last: 0, +// both: 2, +// neither: 0, +// }, +// ); +// let mut read_names: HashMap> = HashMap::new(); +// read_names.insert( +// "read1".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read2".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); +// read_names.insert( +// "read4".to_string(), +// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], +// ); +// read_names.insert( +// "read5".to_string(), +// vec![ +// Arc::clone(&rg_paired), +// Arc::clone(&rg_paired), +// Arc::clone(&rg_single), +// ], +// ); +// let result = predict(ordering_flags, read_names, 0.0, false); +// assert!(!result.succeeded); +// assert_eq!(result.endedness, "Unknown"); +// assert_eq!(result.first, 8); +// assert_eq!(result.last, 8); +// assert_eq!(result.both, 2); +// assert_eq!(result.neither, 0); +// assert_eq!(result.rpt, Some(2.2)); +// assert_eq!(result.read_groups.len(), 2); +// // We can't know which read group will be first in the vector. +// // But both should succeed. +// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); +// } +// } diff --git a/src/utils/read_groups.rs b/src/utils/read_groups.rs index aa72f28..7cd0b57 100644 --- a/src/utils/read_groups.rs +++ b/src/utils/read_groups.rs @@ -1,20 +1,41 @@ //! This module contains functions to validate the read group information in the header and the records. +use lazy_static::lazy_static; +use noodles::sam::alignment::Record; use noodles::sam::header; +use noodles::sam::record::data::field::Tag; use std::collections::HashSet; use std::sync::Arc; use tracing::warn; -use lazy_static::lazy_static; +/// Type alias for a read group pointer. +pub type ReadGroupPtr = Arc; // Strings used to index into the HashMaps used to store the Read Group ordering flags. // Lazy statics are used to save memory. lazy_static! { - /// String used to index into the HashMaps used to store the "overall" ordering flags. - pub static ref OVERALL: Arc = Arc::new(String::from("overall")); - /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. - pub static ref UNKNOWN_READ_GROUP: Arc = Arc::new(String::from("unknown_read_group")); + pub static ref UNKNOWN_READ_GROUP: ReadGroupPtr = Arc::new(String::from("unknown_read_group")); +} + +/// Returns the read group tag from the record. +/// If the read group is not found in the record, the read group is set to "unknown_read_group". +/// TODO: Revisit this logic +pub fn get_read_group( + record: &Record, + found_rgs: Option<&mut HashSet>, +) -> ReadGroupPtr { + match (record.data().get(Tag::ReadGroup), found_rgs) { + (Some(rg), Some(read_groups)) => { + let rg = rg.to_string(); + if !read_groups.contains(&rg) { + read_groups.insert(Arc::new(rg.clone())); + } + Arc::clone(read_groups.get(&rg).unwrap()) + } + (Some(rg), None) => Arc::new(rg.to_string()), + (None, _) => Arc::clone(&UNKNOWN_READ_GROUP), + } } /// Compares the read group tags found in the records @@ -22,7 +43,7 @@ lazy_static! { /// Returns a vector of read group names that were found in the header /// but not in the records. pub fn validate_read_group_info( - found_rgs: &HashSet>, + found_rgs: &HashSet, header: &header::Header, ) -> Vec { let mut rgs_in_header_not_records = Vec::new(); From 7f95d342e186ce820a48898d9427a1e60a55a024 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 11:20:44 -0500 Subject: [PATCH 54/91] refactor(derive/strandedness): separate out results from compute --- src/derive/command/strandedness.rs | 42 +++--- src/derive/strandedness.rs | 1 + src/derive/strandedness/compute.rs | 213 ++--------------------------- src/derive/strandedness/results.rs | 193 ++++++++++++++++++++++++++ 4 files changed, 227 insertions(+), 222 deletions(-) create mode 100644 src/derive/strandedness/results.rs diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index f28e800..3511dc0 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -13,10 +13,10 @@ use noodles::gff; use rust_lapper::{Interval, Lapper}; use tracing::debug; use tracing::info; -use tracing::warn; use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; +use crate::derive::strandedness::results; use crate::utils::formats; /// Clap arguments for the `ngs derive strandedness` subcommand. @@ -95,8 +95,8 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { let mut gene_records = Vec::new(); let mut exon_records = Vec::new(); - let mut gene_metrics = compute::GeneRecordMetrics::default(); - let mut exon_metrics = compute::ExonRecordMetrics::default(); + let mut gene_metrics = results::GeneRecordMetrics::default(); + let mut exon_metrics = results::ExonRecordMetrics::default(); for result in gff.records() { let record = result.unwrap(); if record.ty() == args.gene_feature_name { @@ -214,17 +214,17 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { counts: HashMap::new(), found_rgs: HashSet::new(), }; - let mut metrics = compute::RecordTracker { + let mut metrics = results::RecordTracker { genes: gene_metrics, exons: exon_metrics, - reads: compute::ReadRecordMetrics::default(), + reads: results::ReadRecordMetrics::default(), }; - let mut result: compute::DerivedStrandednessResult; + let mut result: Option = None; for try_num in 1..=args.max_tries { info!("Starting try {} of {}", try_num, args.max_tries); - result = compute::predict( + let attempt = compute::predict( &mut parsed_bam, &mut gene_records, &exons, @@ -233,25 +233,23 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { &mut metrics, )?; - if result.succeeded { + if attempt.succeeded { info!("Strandedness test succeeded."); - - // (#) Print the output to stdout as JSON (more support for different output - // types may be added in the future, but for now, only JSON). - let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); - break; } else { - warn!("Strandedness test inconclusive."); - - if try_num >= args.max_tries { - info!("Strandedness test failed after {} tries.", args.max_tries); - let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); - break; - } + info!("Strandedness test inconclusive."); } + result = Some(attempt); } + let result = result.unwrap(); + + if !result.succeeded { + info!("Strandedness test failed after {} tries.", args.max_tries); + } + + // (4) Print the output to stdout as JSON (more support for different output + // types may be added in the future, but for now, only JSON). + let output = serde_json::to_string_pretty(&result).unwrap(); + print!("{}", output); anyhow::Ok(()) } diff --git a/src/derive/strandedness.rs b/src/derive/strandedness.rs index 0551408..4349a3c 100644 --- a/src/derive/strandedness.rs +++ b/src/derive/strandedness.rs @@ -1,3 +1,4 @@ //! Supporting functionality for the `ngs derive strandedness` subcommand. pub mod compute; +pub mod results; diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 3ada827..53b471c 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -7,86 +7,16 @@ use noodles::sam; use noodles::sam::record::data::field::Tag; use rand::Rng; use rust_lapper::Lapper; -use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; use std::sync::Arc; +use crate::derive::strandedness::results; use crate::utils::read_groups::{validate_read_group_info, UNKNOWN_READ_GROUP}; const STRANDED_THRESHOLD: f64 = 80.0; const UNSTRANDED_THRESHOLD: f64 = 40.0; -/// General gene metrics that are tallied as a part of the -/// strandedness subcommand. -#[derive(Clone, Default, Serialize, Debug)] -pub struct GeneRecordMetrics { - /// The total number of genes found in the GFF. - pub total: usize, - - /// The number of genes that were found to be protein coding. - /// If --all-genes is set this will not be tallied. - pub protein_coding: usize, - - /// The number of genes tested. - pub tested: usize, - - /// The number of genes which were discarded due to having - /// an unknown/invalid strand OR with exons on both strands. - pub bad_strands: usize, - - /// The number of genes which were discarded due to not having - /// enough reads. - pub not_enough_reads: usize, -} - -/// General exon metrics that are tallied as a part of the -/// strandedness subcommand. -#[derive(Clone, Default, Serialize, Debug)] -pub struct ExonRecordMetrics { - /// The total number of exons found in the GFF. - pub total: usize, - - /// The number of exons discarded due to having an unknown/invalid strand. - pub bad_strand: usize, -} - -/// General read record metrics that are tallied as a part of the -/// strandedness subcommand. -#[derive(Clone, Default, Serialize, Debug)] -pub struct ReadRecordMetrics { - /// The number of records that have been filtered because of their flags. - /// (i.e. they were qc_fail, duplicates, secondary, or supplementary) - /// These conditions can be toggled on/off with CL flags - pub filtered_by_flags: usize, - - /// The number of records that have been filtered because - /// they failed the MAPQ filter. - pub low_mapq: usize, - - /// The number of records whose MAPQ couldn't be parsed and were thus discarded. - pub missing_mapq: usize, - - /// The number of records determined to be Paired-End. - pub paired_end_reads: usize, - - /// The number of records determined to be Single-End. - pub single_end_reads: usize, -} - -/// Struct for managing record tracking. -#[derive(Clone, Default, Debug)] -pub struct RecordTracker { - /// Gene metrics. - pub genes: GeneRecordMetrics, - - /// Exon metrics. - pub exons: ExonRecordMetrics, - - /// Read metrics. - pub reads: ReadRecordMetrics, -} - /// Struct for tracking count results. #[derive(Clone, Default)] pub struct Counts { @@ -97,126 +27,6 @@ pub struct Counts { reverse: usize, } -/// Struct holding the per read group results for an `ngs derive strandedness` -/// subcommand call. -#[derive(Debug, Serialize)] -pub struct ReadGroupDerivedStrandednessResult { - /// Name of the read group. - pub read_group: String, - - /// Whether or not strandedness was determined for this read group. - pub succeeded: bool, - - /// The strandedness of this read group or "Inconclusive". - pub strandedness: String, - - /// The total number of reads in this read group. - pub total: usize, - - /// The number of reads that are evidence of Forward Strandedness. - pub forward: usize, - - /// The number of reads that are evidence of Reverse Strandedness. - pub reverse: usize, - - /// The percent of evidence for Forward Strandedness. - pub forward_pct: f64, - - /// The percent of evidence for Reverse Strandedness. - pub reverse_pct: f64, -} - -impl ReadGroupDerivedStrandednessResult { - /// Creates a new [`ReadGroupDerivedStrandednessResult`]. - fn new( - read_group: String, - succeeded: bool, - strandedness: String, - forward: usize, - reverse: usize, - ) -> Self { - ReadGroupDerivedStrandednessResult { - read_group, - succeeded, - strandedness, - total: forward + reverse, - forward, - reverse, - forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, - reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, - } - } -} - -/// Struct holding the final results for an `ngs derive strandedness` subcommand -/// call. -#[derive(Debug, Serialize)] -pub struct DerivedStrandednessResult { - /// Whether or not the `ngs derive strandedness` subcommand succeeded. - pub succeeded: bool, - - /// The strandedness of this read group or "Inconclusive". - pub strandedness: String, - - /// The total number of reads. - pub total: usize, - - /// The number of reads that are evidence of Forward Strandedness. - pub forward: usize, - - /// The number of reads that are evidence of Reverse Strandedness. - pub reverse: usize, - - /// The percent of evidence for Forward Strandedness. - pub forward_pct: f64, - - /// The percent of evidence for Reverse Strandedness. - pub reverse_pct: f64, - - /// Vector of [`ReadGroupDerivedStrandednessResult`]s. - /// One for each read group in the BAM, - /// and potentially one for any reads with an unknown read group. - pub read_groups: Vec, - - /// General read record metrics that are tallied as a part of the - /// strandedness subcommand. - pub read_metrics: ReadRecordMetrics, - - /// General gene metrics that are tallied as a part of the - /// strandedness subcommand. - pub gene_metrics: GeneRecordMetrics, - - /// General exon metrics that are tallied as a part of the - /// strandedness subcommand. - pub exon_metrics: ExonRecordMetrics, -} - -impl DerivedStrandednessResult { - /// Creates a new [`DerivedStrandednessResult`]. - fn new( - succeeded: bool, - strandedness: String, - forward: usize, - reverse: usize, - read_groups: Vec, - metrics: RecordTracker, - ) -> Self { - DerivedStrandednessResult { - succeeded, - strandedness, - total: forward + reverse, - forward, - reverse, - forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, - reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, - read_groups, - read_metrics: metrics.reads, - gene_metrics: metrics.genes, - exon_metrics: metrics.exons, - } - } -} - #[derive(Clone, Copy, Debug)] enum Strand { Forward, @@ -349,7 +159,7 @@ fn query_and_filter( parsed_bam: &mut ParsedBAMFile, gene: &gff::Record, params: &StrandednessParams, - read_metrics: &mut ReadRecordMetrics, + read_metrics: &mut results::ReadRecordMetrics, ) -> Vec { let start = gene.start(); let end = gene.end(); @@ -405,7 +215,7 @@ fn classify_read( read: &sam::alignment::Record, gene_strand: &Strand, all_counts: &mut AllReadGroupsCounts, - read_metrics: &mut ReadRecordMetrics, + read_metrics: &mut results::ReadRecordMetrics, ) { let read_group = match read.data().get(Tag::ReadGroup) { Some(rg) => { @@ -455,9 +265,12 @@ fn classify_read( } /// Method to predict the strandedness of a read group. -fn predict_strandedness(rg_name: &str, counts: &Counts) -> ReadGroupDerivedStrandednessResult { +fn predict_strandedness( + rg_name: &str, + counts: &Counts, +) -> results::ReadGroupDerivedStrandednessResult { if counts.forward == 0 && counts.reverse == 0 { - return ReadGroupDerivedStrandednessResult { + return results::ReadGroupDerivedStrandednessResult { read_group: rg_name.to_string(), succeeded: false, strandedness: "Inconclusive".to_string(), @@ -468,7 +281,7 @@ fn predict_strandedness(rg_name: &str, counts: &Counts) -> ReadGroupDerivedStran reverse_pct: 0.0, }; } - let mut result = ReadGroupDerivedStrandednessResult::new( + let mut result = results::ReadGroupDerivedStrandednessResult::new( rg_name.to_string(), false, "Inconclusive".to_string(), @@ -500,8 +313,8 @@ pub fn predict( exons: &HashMap<&str, Lapper>, all_counts: &mut AllReadGroupsCounts, params: &StrandednessParams, - metrics: &mut RecordTracker, -) -> Result { + metrics: &mut results::RecordTracker, +) -> Result { let mut rng = rand::thread_rng(); let mut num_tested_genes: usize = 0; // Local to this attempt let genes_remaining = gene_records.len(); @@ -578,7 +391,7 @@ pub fn predict( } let overall_result = predict_strandedness("overall", &overall_counts); - let final_result = DerivedStrandednessResult::new( + let final_result = results::DerivedStrandednessResult::new( overall_result.succeeded, overall_result.strandedness, overall_result.forward, @@ -650,7 +463,7 @@ mod tests { counts: HashMap::new(), found_rgs: HashSet::new(), }; - let mut read_metrics = ReadRecordMetrics::default(); + let mut read_metrics = results::ReadRecordMetrics::default(); let counts_key = Arc::new("rg1".to_string()); let rg_tag = sam::record::data::field::Value::String("rg1".to_string()); diff --git a/src/derive/strandedness/results.rs b/src/derive/strandedness/results.rs new file mode 100644 index 0000000..abe9d99 --- /dev/null +++ b/src/derive/strandedness/results.rs @@ -0,0 +1,193 @@ +//! Results structs for the strandedness subcommand. + +use serde::Serialize; + +/// General read record metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize, Debug)] +pub struct ReadRecordMetrics { + /// The number of records that have been filtered because of their flags. + /// (i.e. they were qc_fail, duplicates, secondary, or supplementary) + /// These conditions can be toggled on/off with CL flags + pub filtered_by_flags: usize, + + /// The number of records that have been filtered because + /// they failed the MAPQ filter. + pub low_mapq: usize, + + /// The number of records whose MAPQ couldn't be parsed and were thus discarded. + pub missing_mapq: usize, + + /// The number of records determined to be Paired-End. + pub paired_end_reads: usize, + + /// The number of records determined to be Single-End. + pub single_end_reads: usize, +} + +/// General gene metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize, Debug)] +pub struct GeneRecordMetrics { + /// The total number of genes found in the GFF. + pub total: usize, + + /// The number of genes that were found to be protein coding. + /// If --all-genes is set this will not be tallied. + pub protein_coding: usize, + + /// The number of genes tested. + pub tested: usize, + + /// The number of genes which were discarded due to having + /// an unknown/invalid strand OR with exons on both strands. + pub bad_strands: usize, + + /// The number of genes which were discarded due to not having + /// enough reads. + pub not_enough_reads: usize, +} + +/// General exon metrics that are tallied as a part of the +/// strandedness subcommand. +#[derive(Clone, Default, Serialize, Debug)] +pub struct ExonRecordMetrics { + /// The total number of exons found in the GFF. + pub total: usize, + + /// The number of exons discarded due to having an unknown/invalid strand. + pub bad_strand: usize, +} + +/// Struct for managing record tracking. +#[derive(Clone, Default, Debug)] +pub struct RecordTracker { + /// Read metrics. + pub reads: ReadRecordMetrics, + + /// Gene metrics. + pub genes: GeneRecordMetrics, + + /// Exon metrics. + pub exons: ExonRecordMetrics, +} + +/// Struct holding the per read group results for an `ngs derive strandedness` +/// subcommand call. +#[derive(Debug, Serialize)] +pub struct ReadGroupDerivedStrandednessResult { + /// Name of the read group. + pub read_group: String, + + /// Whether or not strandedness was determined for this read group. + pub succeeded: bool, + + /// The strandedness of this read group or "Inconclusive". + pub strandedness: String, + + /// The total number of reads in this read group. + pub total: usize, + + /// The number of reads that are evidence of Forward Strandedness. + pub forward: usize, + + /// The number of reads that are evidence of Reverse Strandedness. + pub reverse: usize, + + /// The percent of evidence for Forward Strandedness. + pub forward_pct: f64, + + /// The percent of evidence for Reverse Strandedness. + pub reverse_pct: f64, +} + +impl ReadGroupDerivedStrandednessResult { + /// Creates a new [`ReadGroupDerivedStrandednessResult`]. + pub fn new( + read_group: String, + succeeded: bool, + strandedness: String, + forward: usize, + reverse: usize, + ) -> Self { + ReadGroupDerivedStrandednessResult { + read_group, + succeeded, + strandedness, + total: forward + reverse, + forward, + reverse, + forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, + reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, + } + } +} + +/// Struct holding the final results for an `ngs derive strandedness` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedStrandednessResult { + /// Whether or not the `ngs derive strandedness` subcommand succeeded. + pub succeeded: bool, + + /// The strandedness of this read group or "Inconclusive". + pub strandedness: String, + + /// The total number of reads. + pub total: usize, + + /// The number of reads that are evidence of Forward Strandedness. + pub forward: usize, + + /// The number of reads that are evidence of Reverse Strandedness. + pub reverse: usize, + + /// The percent of evidence for Forward Strandedness. + pub forward_pct: f64, + + /// The percent of evidence for Reverse Strandedness. + pub reverse_pct: f64, + + /// Vector of [`ReadGroupDerivedStrandednessResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, + + /// General read record metrics that are tallied as a part of the + /// strandedness subcommand. + pub read_metrics: ReadRecordMetrics, + + /// General gene metrics that are tallied as a part of the + /// strandedness subcommand. + pub gene_metrics: GeneRecordMetrics, + + /// General exon metrics that are tallied as a part of the + /// strandedness subcommand. + pub exon_metrics: ExonRecordMetrics, +} + +impl DerivedStrandednessResult { + /// Creates a new [`DerivedStrandednessResult`]. + pub fn new( + succeeded: bool, + strandedness: String, + forward: usize, + reverse: usize, + read_groups: Vec, + metrics: RecordTracker, + ) -> Self { + DerivedStrandednessResult { + succeeded, + strandedness, + total: forward + reverse, + forward, + reverse, + forward_pct: (forward as f64 / (forward + reverse) as f64) * 100.0, + reverse_pct: (reverse as f64 / (forward + reverse) as f64) * 100.0, + read_groups, + read_metrics: metrics.reads, + gene_metrics: metrics.genes, + exon_metrics: metrics.exons, + } + } +} From d8d9413c2a765925330c3a7f2b7df6b7e240e194 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 11:26:49 -0500 Subject: [PATCH 55/91] fix(strandedness): break when successful --- src/derive/command/strandedness.rs | 7 ++++--- 1 file changed, 4 insertions(+), 3 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 3511dc0..89f7394 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -232,13 +232,14 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { ¶ms, &mut metrics, )?; - - if attempt.succeeded { + let success = attempt.succeeded; + result = Some(attempt); + if success { info!("Strandedness test succeeded."); + break; } else { info!("Strandedness test inconclusive."); } - result = Some(attempt); } let result = result.unwrap(); From b084993bee998c68920fce79cf50cbc60a931d2c Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 11:45:20 -0500 Subject: [PATCH 56/91] docs: junction_annotation to junction-annotation --- src/derive/command/junction_annotation.rs | 8 ++++---- src/derive/junction_annotation.rs | 2 +- src/derive/junction_annotation/compute.rs | 2 +- src/derive/junction_annotation/results.rs | 8 ++++---- 4 files changed, 10 insertions(+), 10 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 2954110..39809f8 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -1,4 +1,4 @@ -//! Functionality relating to the `ngs derive junction_annotation` subcommand itself. +//! Functionality relating to the `ngs derive junction-annotation` subcommand itself. use std::collections::HashMap; use std::path::PathBuf; @@ -17,7 +17,7 @@ use crate::utils::formats; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; -/// Clap arguments for the `ngs derive junction_annotation` subcommand. +/// Clap arguments for the `ngs derive junction-annotation` subcommand. #[derive(Args)] pub struct JunctionAnnotationArgs { /// Source BAM. @@ -60,9 +60,9 @@ pub struct JunctionAnnotationArgs { count_duplicates: bool, } -/// Main function for the `ngs derive junction_annotation` subcommand. +/// Main function for the `ngs derive junction-annotation` subcommand. pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { - info!("Starting derive junction_annotation subcommand."); + info!("Starting derive junction-annotation subcommand."); let mut exons = compute::ExonSets { starts: HashMap::new(), diff --git a/src/derive/junction_annotation.rs b/src/derive/junction_annotation.rs index df1f81a..77e2617 100644 --- a/src/derive/junction_annotation.rs +++ b/src/derive/junction_annotation.rs @@ -1,4 +1,4 @@ -//! Supporting functionality for the `ngs derive junction_annotation` subcommand. +//! Supporting functionality for the `ngs derive junction-annotation` subcommand. pub mod compute; pub mod results; diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 8682a8c..dbfed06 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -261,7 +261,7 @@ fn tally_junctions_and_support(junction_map: &results::JunctionsMap) -> (usize, (junctions, support) } -/// Main function to summarize the results of the junction_annotation subcommand. +/// Main function to summarize the results of the junction-annotation subcommand. pub fn summarize( results: &mut results::JunctionAnnotationResults, params: &JunctionAnnotationParameters, diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 155c0a5..912acc5 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -1,4 +1,4 @@ -//! Results related to the `ngs derive junction_annotation` subcommand. +//! Results related to the `ngs derive junction-annotation` subcommand. use noodles::core::Position; use serde::ser::SerializeStruct; @@ -82,7 +82,7 @@ impl Serialize for JunctionAnnotations { } /// General record metrics that are tallied as a part of the -/// junction_annotation subcommand. +/// junction-annotation subcommand. #[derive(Clone, Default, Serialize)] pub struct RecordMetrics { /// The number of records that have been fully processed. @@ -106,7 +106,7 @@ pub struct RecordMetrics { pub missing_mapq: usize, } -/// Summary statistics for the junction_annotation subcommand. +/// Summary statistics for the junction-annotation subcommand. #[derive(Clone, Default, Serialize)] pub struct SummaryResults { /// The total number of junctions observed in the file. @@ -208,6 +208,6 @@ pub struct JunctionAnnotationResults { /// General record metrics. pub records: RecordMetrics, - /// Summary statistics for the junction_annotation subcommand. + /// Summary statistics for the junction-annotation subcommand. pub summary: SummaryResults, } From 7767da43d63ce96eaba33c852b20cb0c992bafd3 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 9 Feb 2024 17:25:10 -0500 Subject: [PATCH 57/91] [WIP]: switch min_mapq to a proper MappingQuality --- src/derive/command/junction_annotation.rs | 4 +- src/derive/command/strandedness.rs | 24 +-- src/derive/junction_annotation/compute.rs | 178 +++++++++++++++++----- src/derive/junction_annotation/results.rs | 6 +- src/derive/strandedness/compute.rs | 62 ++++---- src/derive/strandedness/results.rs | 8 +- src/utils/alignment.rs | 45 +++++- src/utils/args.rs | 14 +- 8 files changed, 243 insertions(+), 98 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 39809f8..811dd1a 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -5,6 +5,7 @@ use std::path::PathBuf; use anyhow::Context; use clap::Args; +use noodles::sam::record::MappingQuality; use num_format::Locale; use num_format::ToFormattedString; use tracing::debug; @@ -45,7 +46,7 @@ pub struct JunctionAnnotationArgs { /// Set to 0 to disable this filter and allow reads _without_ /// a mapping quality to be considered. #[arg(short, long, value_name = "U8", default_value = "30")] - min_mapq: u8, + min_mapq: Option, /// Do not count supplementary alignments. #[arg(long)] @@ -95,6 +96,7 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { debug!("Done reading GFF."); + // (1.5) Initialize variables (including opening the BAM). let mut counter = RecordCounter::new(); let mut results = JunctionAnnotationResults::default(); let params = compute::JunctionAnnotationParameters { diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 89f7394..3fe1c6a 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -10,6 +10,7 @@ use anyhow::Context; use clap::Args; use noodles::bam; use noodles::gff; +use noodles::sam::record::MappingQuality; use rust_lapper::{Interval, Lapper}; use tracing::debug; use tracing::info; @@ -17,6 +18,7 @@ use tracing::info; use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; use crate::derive::strandedness::results; +use crate::utils::args::parse_min_mapq; use crate::utils::formats; /// Clap arguments for the `ngs derive strandedness` subcommand. @@ -45,10 +47,10 @@ pub struct DeriveStrandednessArgs { num_genes: usize, /// Minimum mapping quality for a record to be considered. - /// Set to 0 to disable this filter and allow reads _without_ - /// a mapping quality to be considered. - #[arg(long, value_name = "U8", default_value = "30")] - min_mapq: u8, + /// Default is to ignore MAPQ values (allowing MAPQs to be considered). + /// Specify any u8 value to enable this filter. + #[arg(long, value_name = "U8", default_value = "30", value_parser = parse_min_mapq)] + min_mapq: Option, /// Consider all genes, not just protein coding genes. #[arg(long)] @@ -88,7 +90,7 @@ pub struct DeriveStrandednessArgs { pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { info!("Starting derive strandedness subcommand."); - // (1) Parse the GFF file and collect all gene features. + // (1) Parse the GFF file and collect all gene and exon features. debug!("Reading all records in GFF."); let mut gff = formats::gff::open(&args.features_gff) .with_context(|| format!("opening GFF file: {}", args.features_gff.display()))?; @@ -130,9 +132,6 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { exon_records.push(record); } } - - debug!("Tabulating GFF gene and exon features."); - if gene_records.is_empty() { bail!("No gene records matched criteria. Check your GFF file and `--gene-feature-name` and `--all-genes` options."); } @@ -145,6 +144,9 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { exon_records.len() ); + // (2) Parse exon features into proper data structure. + debug!("Tabulating GFF exon features."); + let mut exon_intervals: HashMap<&str, Vec>> = HashMap::new(); for record in &exon_records { @@ -180,6 +182,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { debug!("Done reading GFF."); + // (3) Initialize variables (including opening the BAM). let mut reader = File::open(&args.src) .map(bam::Reader::new) .with_context(|| format!("opening BAM file: {}", args.src.display()))?; @@ -219,8 +222,9 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { exons: exon_metrics, reads: results::ReadRecordMetrics::default(), }; - let mut result: Option = None; + + // (4) Run the strandedness test. for try_num in 1..=args.max_tries { info!("Starting try {} of {}", try_num, args.max_tries); @@ -247,7 +251,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { info!("Strandedness test failed after {} tries.", args.max_tries); } - // (4) Print the output to stdout as JSON (more support for different output + // (5) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); print!("{}", output); diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index dbfed06..6d71acd 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -5,11 +5,13 @@ use anyhow::Ok; use noodles::core::Position; use noodles::sam::alignment::Record; use noodles::sam::record::cigar::op::Kind; +use noodles::sam::record::MappingQuality; use noodles::sam::Header; use std::collections::HashMap; use std::collections::HashSet; use crate::derive::junction_annotation::results; +use crate::utils::alignment::filter_by_mapq; /// Struct to hold starts and ends of exons. pub struct ExonSets<'a> { @@ -30,7 +32,7 @@ pub struct JunctionAnnotationParameters { /// Minumum mapping quality for a record to be considered. /// 0 if MAPQ shouldn't be considered. - pub min_mapq: u8, + pub min_mapq: Option, /// Do not count supplementary alignments. pub no_supplementary: bool, @@ -65,6 +67,27 @@ fn increment_junction_map( ); } +/// Function to filter out records based on their flags. +fn filter_by_flags(record: &Record, params: &JunctionAnnotationParameters) -> bool { + let flags = record.flags(); + if flags.is_unmapped() + || (params.no_supplementary && flags.is_supplementary()) + || (!params.count_secondary && flags.is_secondary()) + || (!params.count_duplicates && flags.is_duplicate()) + { + return true; + } + false +} + +/// Function to filter out records that don't have introns. +fn filter_by_cigar(record: &Record) -> bool { + !record + .cigar() + .iter() + .any(|op| matches!(op.kind(), Kind::Skip)) +} + /// Main function to annotate junctions one record at a time. pub fn process( record: &Record, @@ -79,42 +102,24 @@ pub fn process( _ => bail!("Could not parse read name"), }; - // (2) Parse the flags so we can see if the read should be ignored. - let flags = record.flags(); - - if flags.is_unmapped() - || (params.no_supplementary && flags.is_supplementary()) - || (!params.count_secondary && flags.is_secondary()) - || (!params.count_duplicates && flags.is_duplicate()) - { + // (2) Filter by record flags. + if filter_by_flags(record, params) { results.records.filtered_by_flags += 1; return Ok(()); } - // (3) Parse the CIGAR string from the record. + // (3) Filter by CIGAR. // We only care about reads with introns, so if there are no introns // we can skip this read. - let cigar = record.cigar(); - if !cigar.iter().any(|op| matches!(op.kind(), Kind::Skip)) { + if filter_by_cigar(record) { results.records.not_spliced += 1; return Ok(()); } - // (4) If the user is filtering by MAPQ, check if this read passes. - // Log if the read is filtered out for a too low MAPQ or a missing MAPQ. - if params.min_mapq > 0 { - match record.mapping_quality() { - Some(mapq) => { - if mapq.get() < params.min_mapq { - results.records.low_mapq += 1; - return Ok(()); - } - } - None => { - results.records.missing_mapq += 1; - return Ok(()); - } - } + // (4) Filter by MAPQ + if filter_by_mapq(record, params.min_mapq) { + results.records.bad_mapq += 1; + return Ok(()); } // (5) Parse the reference sequence from the record. @@ -144,7 +149,7 @@ pub fn process( // (8) Find introns let cur_pos = start; - for op in cigar.iter() { + for op in record.cigar().iter() { match op.kind() { // This is an intron. Kind::Skip => { @@ -355,6 +360,103 @@ mod tests { use noodles::sam::record::ReadName; use std::num::NonZeroUsize; + #[test] + fn test_filter_by_flags() { + // Setup + let mut record = Record::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + + // Test that records are filtered out correctly + record.flags_mut().set(0x4.into(), true); + assert!(filter_by_flags(&record, ¶ms)); + record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x800.into(), true); + assert!(!filter_by_flags(&record, ¶ms)); + record.flags_mut().set(0x800.into(), false); + record.flags_mut().set(0x100.into(), true); + assert!(filter_by_flags(&record, ¶ms)); + record.flags_mut().set(0x100.into(), false); + record.flags_mut().set(0x400.into(), true); + assert!(filter_by_flags(&record, ¶ms)); + record.flags_mut().set(0x400.into(), false); + assert!(!filter_by_flags(&record, ¶ms)); + } + + #[test] + fn test_filter_by_cigar() { + // Setup + let mut record = Record::default(); + + // Test that records are filtered out correctly + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + assert!(!filter_by_cigar(&record)); + *record.cigar_mut() = "10M".parse().unwrap(); + assert!(filter_by_cigar(&record)); + } + + #[test] + fn test_filter_junction_map() { + // Setup + let mut junction_map = results::JunctionsMap::default(); + junction_map.insert( + "sq1".to_string(), + HashMap::from([ + ((Position::new(1).unwrap(), Position::new(11).unwrap()), 1), + ((Position::new(1).unwrap(), Position::new(5).unwrap()), 1), + ]), + ); + junction_map.insert( + "sq2".to_string(), + HashMap::from([((Position::new(1).unwrap(), Position::new(11).unwrap()), 2)]), + ); + let min_intron_length = 10; + let min_read_support = 2; + let mut metrics = results::SummaryResults::default(); + + // Test that junctions are filtered out correctly + filter_junction_map( + &mut junction_map, + min_intron_length, + min_read_support, + &mut metrics, + ); + assert_eq!(junction_map.len(), 1); + assert_eq!(junction_map.get("sq1"), None); + assert_eq!(junction_map.get("sq2").unwrap().len(), 1); + assert_eq!(metrics.intron_too_short, 1); + assert_eq!(metrics.junctions_with_not_enough_read_support, 2); + assert_eq!(metrics.total_rejected_junctions, 2); + } + + #[test] + fn test_tally_junctions_and_support() { + // Setup + let mut junction_map = results::JunctionsMap::default(); + junction_map.insert( + "sq1".to_string(), + HashMap::from([ + ((Position::new(1).unwrap(), Position::new(11).unwrap()), 1), + ((Position::new(1).unwrap(), Position::new(5).unwrap()), 1), + ]), + ); + junction_map.insert( + "sq2".to_string(), + HashMap::from([((Position::new(1).unwrap(), Position::new(11).unwrap()), 2)]), + ); + + // Test that junctions are tallied correctly + let (juncs, support) = tally_junctions_and_support(&junction_map); + assert_eq!(juncs, 3); + assert_eq!(support, 4); + } + #[test] fn test_process_and_summarize() { // Setup @@ -362,7 +464,7 @@ mod tests { let params = JunctionAnnotationParameters { min_intron_length: 10, min_read_support: 2, - min_mapq: 30, + min_mapq: Some(MappingQuality::new(30).unwrap()), no_supplementary: false, count_secondary: false, count_duplicates: false, @@ -413,8 +515,7 @@ mod tests { assert_eq!(results.records.processed, 1); assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // Test that unmapped gets ignored let mut record = Record::default(); @@ -429,8 +530,7 @@ mod tests { assert_eq!(results.records.processed, 1); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // Test partial novel junction let mut record = Record::default(); @@ -445,8 +545,7 @@ mod tests { assert_eq!(results.records.processed, 2); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // Test partial novel junction (again for more read support) let mut record = Record::default(); @@ -461,8 +560,7 @@ mod tests { assert_eq!(results.records.processed, 3); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // Test that supplementary alignments get counted let mut record = Record::default(); @@ -478,8 +576,7 @@ mod tests { assert_eq!(results.records.processed, 4); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // Test that secondary alignments don't get counted let mut record = Record::default(); @@ -495,8 +592,7 @@ mod tests { assert_eq!(results.records.processed, 4); assert_eq!(results.records.filtered_by_flags, 2); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.low_mapq, 0); - assert_eq!(results.records.missing_mapq, 0); + assert_eq!(results.records.bad_mapq, 0); // TODO: Below tests are not working as expected. Need to fix them. // Test complete novel junction diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 912acc5..a3c945e 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -100,10 +100,8 @@ pub struct RecordMetrics { /// The number of records with junctions that have been ignored because /// they failed the MAPQ filter. - pub low_mapq: usize, - - /// The number of records whose MAPQ couldn't be parsed and were thus ignored. - pub missing_mapq: usize, + /// This could either mean the MAPQ was too low or it was missing. + pub bad_mapq: usize, } /// Summary statistics for the junction-annotation subcommand. diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 53b471c..a939065 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -5,6 +5,7 @@ use noodles::core::Region; use noodles::gff; use noodles::sam; use noodles::sam::record::data::field::Tag; +use noodles::sam::record::MappingQuality; use rand::Rng; use rust_lapper::Lapper; use std::collections::HashMap; @@ -12,6 +13,7 @@ use std::collections::HashSet; use std::sync::Arc; use crate::derive::strandedness::results; +use crate::utils::alignment::filter_by_mapq; use crate::utils::read_groups::{validate_read_group_info, UNKNOWN_READ_GROUP}; const STRANDED_THRESHOLD: f64 = 80.0; @@ -27,6 +29,7 @@ pub struct Counts { reverse: usize, } +/// Struct for tracking possible strand orientations. #[derive(Clone, Copy, Debug)] enum Strand { Forward, @@ -55,6 +58,7 @@ impl TryFrom for Strand { } } +/// Struct for tracking the order of segments in a record. #[derive(Clone, Copy, Debug)] enum SegmentOrder { First, @@ -113,7 +117,7 @@ pub struct StrandednessParams { /// Minumum mapping quality for a record to be considered. /// 0 if MAPQ shouldn't be considered. - pub min_mapq: u8, + pub min_mapq: Option, /// Allow qc failed reads to be counted. pub count_qc_failed: bool, @@ -128,6 +132,7 @@ pub struct StrandednessParams { pub count_duplicates: bool, } +/// Function to disqualify a gene based on its strand and exons. fn disqualify_gene( gene: &gff::Record, exons: &HashMap<&str, Lapper>, @@ -155,6 +160,21 @@ fn disqualify_gene( true } +/// Function to filter out records based on their flags. +fn filter_by_flags(record: &sam::alignment::Record, params: &StrandednessParams) -> bool { + let flags = record.flags(); + if (!params.count_qc_failed && flags.is_qc_fail()) + || (params.no_supplementary && flags.is_supplementary()) + || (!params.count_secondary && flags.is_secondary()) + || (!params.count_duplicates && flags.is_duplicate()) + { + return true; + } + false +} + +/// Function to query the BAM file and filter the records based on the +/// parameters provided. fn query_and_filter( parsed_bam: &mut ParsedBAMFile, gene: &gff::Record, @@ -174,31 +194,16 @@ fn query_and_filter( for read in query { let read = read.unwrap(); - // (1) Parse the flags so we can see if the read should be discarded. - let flags = read.flags(); - if (!params.count_qc_failed && flags.is_qc_fail()) - || (params.no_supplementary && flags.is_supplementary()) - || (!params.count_secondary && flags.is_secondary()) - || (!params.count_duplicates && flags.is_duplicate()) - { + // (1) Filter by flags. + if filter_by_flags(&read, params) { read_metrics.filtered_by_flags += 1; continue; } - // (2) If the user is filtering by MAPQ, check if this read passes. - if params.min_mapq > 0 { - match read.mapping_quality() { - Some(mapq) => { - if mapq.get() < params.min_mapq { - read_metrics.low_mapq += 1; - continue; - } - } - None => { - read_metrics.missing_mapq += 1; - continue; - } - } + // (2) Filter by MAPQ. + if filter_by_mapq(&read, params.min_mapq) { + read_metrics.bad_mapq += 1; + continue; } filtered_reads.push(read); @@ -211,6 +216,7 @@ fn query_and_filter( filtered_reads } +/// Function to classify a read based on its strand and the strand of the gene. fn classify_read( read: &sam::alignment::Record, gene_strand: &Strand, @@ -475,8 +481,7 @@ mod tests { assert_eq!(read_metrics.paired_end_reads, 0); assert_eq!(read_metrics.single_end_reads, 1); assert_eq!(read_metrics.filtered_by_flags, 0); - assert_eq!(read_metrics.low_mapq, 0); - assert_eq!(read_metrics.missing_mapq, 0); + assert_eq!(read_metrics.bad_mapq, 0); let counts = all_counts.counts.get(&counts_key).unwrap(); assert_eq!(counts.forward, 1); assert_eq!(counts.reverse, 0); @@ -490,8 +495,7 @@ mod tests { assert_eq!(read_metrics.paired_end_reads, 1); assert_eq!(read_metrics.single_end_reads, 1); assert_eq!(read_metrics.filtered_by_flags, 0); - assert_eq!(read_metrics.low_mapq, 0); - assert_eq!(read_metrics.missing_mapq, 0); + assert_eq!(read_metrics.bad_mapq, 0); let counts = all_counts.counts.get(&counts_key).unwrap(); assert_eq!(counts.forward, 2); assert_eq!(counts.reverse, 0); @@ -505,8 +509,7 @@ mod tests { assert_eq!(read_metrics.paired_end_reads, 2); assert_eq!(read_metrics.single_end_reads, 1); assert_eq!(read_metrics.filtered_by_flags, 0); - assert_eq!(read_metrics.low_mapq, 0); - assert_eq!(read_metrics.missing_mapq, 0); + assert_eq!(read_metrics.bad_mapq, 0); let counts = all_counts.counts.get(&counts_key).unwrap(); assert_eq!(counts.forward, 3); assert_eq!(counts.reverse, 0); @@ -520,8 +523,7 @@ mod tests { assert_eq!(read_metrics.paired_end_reads, 3); assert_eq!(read_metrics.single_end_reads, 1); assert_eq!(read_metrics.filtered_by_flags, 0); - assert_eq!(read_metrics.low_mapq, 0); - assert_eq!(read_metrics.missing_mapq, 0); + assert_eq!(read_metrics.bad_mapq, 0); let counts = all_counts.counts.get(&counts_key).unwrap(); assert_eq!(counts.forward, 3); assert_eq!(counts.reverse, 1); diff --git a/src/derive/strandedness/results.rs b/src/derive/strandedness/results.rs index abe9d99..3b94963 100644 --- a/src/derive/strandedness/results.rs +++ b/src/derive/strandedness/results.rs @@ -11,12 +11,8 @@ pub struct ReadRecordMetrics { /// These conditions can be toggled on/off with CL flags pub filtered_by_flags: usize, - /// The number of records that have been filtered because - /// they failed the MAPQ filter. - pub low_mapq: usize, - - /// The number of records whose MAPQ couldn't be parsed and were thus discarded. - pub missing_mapq: usize, + /// The number of records that have been ignored because they failed the MAPQ filter. + pub bad_mapq: usize, /// The number of records determined to be Paired-End. pub paired_end_reads: usize, diff --git a/src/utils/alignment.rs b/src/utils/alignment.rs index 1a731f1..c3cd230 100644 --- a/src/utils/alignment.rs +++ b/src/utils/alignment.rs @@ -1,11 +1,25 @@ //! Utilities related to alignment of sequences. use anyhow::bail; -use noodles::sam::record::{cigar::op::Kind, sequence::Base, Cigar}; +use noodles::sam::record::{cigar::op::Kind, sequence::Base, Cigar, MappingQuality}; use super::cigar::consumes_reference; use super::cigar::consumes_sequence; +/// Filter an alignment record by its mapping quality. `true` means "filter the record" and `false` means "do not filter the record". +pub fn filter_by_mapq( + record: &noodles::sam::alignment::Record, + min_mapq: Option, +) -> bool { + match min_mapq { + Some(min_mapq) => match record.mapping_quality() { + Some(mapq) => mapq.get() < min_mapq.get(), + None => false, + }, + None => false, + } +} + /// Turns a condensed Cigar representation into a flattened representation. For /// example, 10M will become a Vec of length 10 comprised completely of /// Kind::MATCH. This utility is useful for generating a representation of a @@ -127,10 +141,37 @@ impl<'a> ReferenceRecordStepThrough<'a> { #[cfg(test)] mod tests { - use noodles::sam::record::{Cigar, Sequence}; + use noodles::sam::record::{Cigar, MappingQuality, Sequence}; use super::ReferenceRecordStepThrough; + #[test] + pub fn it_filters_by_mapq() -> anyhow::Result<()> { + let mut record = noodles::sam::alignment::Record::default(); + assert!(super::filter_by_mapq( + &record, + Some(MappingQuality::new(0).unwrap()) + )); // Get filtered because MAPQ is missing + assert!(!super::filter_by_mapq(&record, None)); // Do not get filtered because filter is disabled + + record + .mapping_quality_mut() + .replace(MappingQuality::new(10).unwrap()); + assert!(!super::filter_by_mapq( + &record, + Some(MappingQuality::new(0).unwrap()) + )); // Do not get filtered because MAPQ is present + assert!(!super::filter_by_mapq( + &record, + Some(MappingQuality::new(1).unwrap()) + )); // Do not get filtered because MAPQ is greater than 1 + assert!(super::filter_by_mapq( + &record, + Some(MappingQuality::new(11).unwrap()) + )); // Do get filtered because MAPQ is less than 11 + Ok(()) + } + #[test] pub fn it_correctly_returns_zero_edits_when_sequences_are_identical() -> anyhow::Result<()> { let reference = "ACTG".parse::()?; diff --git a/src/utils/args.rs b/src/utils/args.rs index 4151cb7..be482ff 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -2,7 +2,7 @@ use std::fmt::Display; -use noodles::bgzf::writer::CompressionLevel; +use noodles::{bgzf::writer::CompressionLevel, sam::record::MappingQuality}; use tracing::debug; //===================// @@ -74,9 +74,9 @@ impl From for CompressionLevel { } } -//==============// -// Float Parser // -//==============// +//=============// +// Arg Parsers // +//=============// /// Utility method to parse command line floats and ensure they are /// within the range [MIN, MAX]. @@ -90,3 +90,9 @@ pub fn arg_in_range(arg: f32, range: std::ops::RangeInclusive) -> anyhow::R ), } } + +/// Utility method to parse command line integers and ensure they are +/// within the range [0, 255) and return them as MappingQualities. +pub fn parse_min_mapq(s: &str) -> Result, std::num::ParseIntError> { + s.parse().map(MappingQuality::new) +} From 64318f338496c3d21068c658e72cb89aa1a092ae Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 09:14:42 -0500 Subject: [PATCH 58/91] fix: rework MappingQuality argument to work --- src/derive/command/junction_annotation.rs | 11 ++++++++--- src/derive/command/strandedness.rs | 14 +++++++++----- src/utils/args.rs | 7 ++++++- 3 files changed, 23 insertions(+), 9 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 811dd1a..578f827 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -43,9 +43,14 @@ pub struct JunctionAnnotationArgs { min_read_support: usize, /// Minumum mapping quality for a record to be considered. - /// Set to 0 to disable this filter and allow reads _without_ - /// a mapping quality to be considered. - #[arg(short, long, value_name = "U8", default_value = "30")] + /// Default behavior is to ignore MAPQ values, + /// which allows reads with _missing_ MAPQs to be considered. + /// Specify any u8 value (lower than 255) to enable this filter. + /// Some aligners erroneously use 255 as the score for a uniquely mapped read; + /// however, 255 is reserved by the spec for a missing MAPQ value. + /// Therefore BAMs produced by aligners using 255 erroneously + /// are not compatible with setting this option. + #[arg(short, long, value_name = "U8")] min_mapq: Option, /// Do not count supplementary alignments. diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 3fe1c6a..304c0d7 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -18,7 +18,6 @@ use tracing::info; use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; use crate::derive::strandedness::results; -use crate::utils::args::parse_min_mapq; use crate::utils::formats; /// Clap arguments for the `ngs derive strandedness` subcommand. @@ -46,10 +45,15 @@ pub struct DeriveStrandednessArgs { #[arg(short = 'n', long, value_name = "USIZE", default_value = "1000")] num_genes: usize, - /// Minimum mapping quality for a record to be considered. - /// Default is to ignore MAPQ values (allowing MAPQs to be considered). - /// Specify any u8 value to enable this filter. - #[arg(long, value_name = "U8", default_value = "30", value_parser = parse_min_mapq)] + /// Minumum mapping quality for a record to be considered. + /// Default behavior is to ignore MAPQ values, + /// which allows reads with _missing_ MAPQs to be considered. + /// Specify any u8 value (lower than 255) to enable this filter. + /// Some aligners erroneously use 255 as the score for a uniquely mapped read; + /// however, 255 is reserved by the spec for a missing MAPQ value. + /// Therefore BAMs produced by aligners using 255 erroneously + /// are not compatible with setting this option. + #[arg(short, long, value_name = "U8")] min_mapq: Option, /// Consider all genes, not just protein coding genes. diff --git a/src/utils/args.rs b/src/utils/args.rs index be482ff..4aa452f 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -91,8 +91,13 @@ pub fn arg_in_range(arg: f32, range: std::ops::RangeInclusive) -> anyhow::R } } +// TODO dead code, not used. Doesn't work as written. /// Utility method to parse command line integers and ensure they are /// within the range [0, 255) and return them as MappingQualities. pub fn parse_min_mapq(s: &str) -> Result, std::num::ParseIntError> { - s.parse().map(MappingQuality::new) + let value = s.parse()?; + match value { + 0..=254 => Ok(Some(MappingQuality::new(value).unwrap())), + 255 => Ok(None), + } } From 2280e1585d588bfe4c94a8f54b4a099b0d5d1275 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 09:15:35 -0500 Subject: [PATCH 59/91] style: f32 -> f64 and code cleanup --- src/derive/command/endedness.rs | 75 ++++----------------------------- src/derive/command/readlen.rs | 4 +- src/derive/endedness/compute.rs | 71 ++++++++++++------------------- src/derive/readlen/compute.rs | 12 +++--- src/generate/command.rs | 4 +- src/utils/args.rs | 2 +- 6 files changed, 45 insertions(+), 123 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 31a9504..7334e2a 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -19,9 +19,7 @@ use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; -use crate::utils::read_groups::{ - get_read_group, validate_read_group_info, ReadGroupPtr, UNKNOWN_READ_GROUP, -}; +use crate::utils::read_groups::{get_read_group, validate_read_group_info, ReadGroupPtr}; /// Clap arguments for the `ngs derive endedness` subcommand. #[derive(Args)] @@ -37,8 +35,8 @@ pub struct DeriveEndednessArgs { /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate /// if the whole file is being processed. - #[arg(long, value_name = "F32", default_value = "0.0")] - paired_deviance: f32, + #[arg(long, value_name = "F64", default_value = "0.0")] + paired_deviance: f64, /// Calculate and output Reads-Per-Template. This will produce a more /// sophisticated estimate for endedness, but uses substantially more memory. @@ -62,8 +60,6 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut found_rgs = HashSet::new(); let mut ordering_flags: HashMap = HashMap::new(); - // TODO change - ordering_flags.insert(Arc::clone(&UNKNOWN_READ_GROUP), OrderingFlagsCounts::new()); // only used if args.calc_rpt is true let mut read_names: HashMap> = HashMap::new(); @@ -113,70 +109,15 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } if !record.flags().is_segmented() { - ordering_flags - .entry(read_group) - .and_modify(|e| { - e.unsegmented += 1; - }) - .or_insert(OrderingFlagsCounts { - unsegmented: 1, - first: 0, - last: 0, - both: 0, - neither: 0, - }); + ordering_flags.entry(read_group).or_default().unsegmented += 1; } else if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags - .entry(read_group) - .and_modify(|e| { - e.first += 1; - }) - .or_insert(OrderingFlagsCounts { - unsegmented: 0, - first: 1, - last: 0, - both: 0, - neither: 0, - }); + ordering_flags.entry(read_group).or_default().first += 1; } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags - .entry(read_group) - .and_modify(|e| { - e.last += 1; - }) - .or_insert(OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 1, - both: 0, - neither: 0, - }); + ordering_flags.entry(read_group).or_default().last += 1; } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags - .entry(read_group) - .and_modify(|e| { - e.both += 1; - }) - .or_insert(OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 0, - both: 1, - neither: 0, - }); + ordering_flags.entry(read_group).or_default().both += 1; } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags - .entry(read_group) - .and_modify(|e| { - e.neither += 1; - }) - .or_insert(OrderingFlagsCounts { - unsegmented: 0, - first: 0, - last: 0, - both: 0, - neither: 1, - }); + ordering_flags.entry(read_group).or_default().neither += 1; } else { unreachable!(); } diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 52bf1f0..668741a 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -28,8 +28,8 @@ pub struct DeriveReadlenArgs { num_records: Option, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. - #[arg(short, long, value_name = "F32", default_value = "0.7")] - majority_vote_cutoff: f32, + #[arg(short, long, value_name = "F64", default_value = "0.7")] + majority_vote_cutoff: f64, } /// Main function for the `ngs derive readlen` subcommand. diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 2b7aad6..0d1043e 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -6,7 +6,7 @@ use std::collections::HashSet; use std::sync::Arc; use tracing::warn; -use crate::utils::read_groups::{ReadGroupPtr, UNKNOWN_READ_GROUP}; +use crate::utils::read_groups::ReadGroupPtr; /// Struct holding the ordering flags for a single read group. #[derive(Debug, Clone)] @@ -240,8 +240,8 @@ fn calculate_reads_per_template( fn predict_endedness( read_group_name: String, rg_ordering_flags: &OrderingFlagsCounts, - paired_deviance: f32, - reads_per_template: Option<&f64>, + paired_deviance: f64, + reads_per_template: Option, round_rpt: bool, ) -> ReadGroupDerivedEndednessResult { let unsegmented = rg_ordering_flags.unsegmented; @@ -262,7 +262,7 @@ fn predict_endedness( false, "Unknown".to_string(), rg_ordering_flags.clone(), - reads_per_template.copied(), + reads_per_template, ); } @@ -271,14 +271,14 @@ fn predict_endedness( false, "Unknown".to_string(), rg_ordering_flags.clone(), - reads_per_template.copied(), + reads_per_template, ); // only unsegmented present if unsegmented > 0 && first == 0 && last == 0 && both == 0 && neither == 0 { match reads_per_template { Some(rpt) => { - if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { + if rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { result.succeeded = true; result.endedness = String::from("Single-End"); } @@ -306,18 +306,18 @@ fn predict_endedness( } // only both present if first == 0 && last == 0 && both > 0 && neither == 0 { - match reads_per_template { - Some(rpt) => { - if *rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { - result.succeeded = true; - result.endedness = String::from("Single-End"); - } - } - None => { - result.succeeded = true; - result.endedness = String::from("Single-End"); - } - } + // match reads_per_template { + // Some(rpt) => { + // if rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { + // result.succeeded = true; + // result.endedness = String::from("Single-End"); + // } + // } + // None => { + // result.succeeded = true; + // result.endedness = String::from("Single-End"); + // } + // } return result; } // only neither present @@ -336,13 +336,13 @@ fn predict_endedness( // both and neither are now guarenteed to be 0 // We only need to check first and last - let first_frac = first as f32 / (first + last) as f32; + let first_frac = first as f64 / (first + last) as f64; let lower_limit = 0.5 - paired_deviance; let upper_limit = 0.5 + paired_deviance; if (first == last) || (lower_limit <= first_frac && first_frac <= upper_limit) { match reads_per_template { Some(rpt) => { - if *rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { + if rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { result.succeeded = true; result.endedness = String::from("Paired-End"); } @@ -362,30 +362,19 @@ fn predict_endedness( pub fn predict( ordering_flags: HashMap, read_names: HashMap>, - paired_deviance: f32, + paired_deviance: f64, round_rpt: bool, ) -> DerivedEndednessResult { let mut rg_rpts: HashMap = HashMap::new(); - let mut overall_rpt: f64 = 0.0; + let mut overall_rpt: Option = None; if !read_names.is_empty() { - overall_rpt = calculate_reads_per_template(read_names, &mut rg_rpts); + overall_rpt = Some(calculate_reads_per_template(read_names, &mut rg_rpts)); } let mut overall_flags = OrderingFlagsCounts::new(); let mut rg_results = Vec::new(); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - // TODO consider refactor to make this unneccessary - if (*read_group == *UNKNOWN_READ_GROUP) - && (rg_ordering_flags.unsegmented == 0 - && rg_ordering_flags.first == 0 - && rg_ordering_flags.last == 0 - && rg_ordering_flags.both == 0 - && rg_ordering_flags.neither == 0) - { - continue; - } - // TODO can make prettier? overall_flags.unsegmented += rg_ordering_flags.unsegmented; overall_flags.first += rg_ordering_flags.first; @@ -397,7 +386,7 @@ pub fn predict( read_group.to_string(), rg_ordering_flags, paired_deviance, - rg_rpts.get(read_group), + rg_rpts.get(read_group).copied(), round_rpt, ); rg_results.push(result); @@ -407,11 +396,7 @@ pub fn predict( "overall".to_string(), &overall_flags, paired_deviance, - if overall_rpt == 0.0 { - None - } else { - Some(&overall_rpt) - }, + overall_rpt, round_rpt, ); @@ -419,11 +404,7 @@ pub fn predict( overall_result.succeeded, overall_result.endedness, overall_flags, - if overall_rpt == 0.0 { - None - } else { - Some(overall_rpt) - }, + overall_rpt, rg_results, ) } diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 9f3bbf7..28b6f97 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -14,11 +14,11 @@ pub struct DerivedReadlenResult { /// The concsensus read length, if available. pub consensus_read_length: Option, - /// The majority vote percentage of the consensus read length, if available. - pub majority_pct_detected: f32, + /// The majority vote percentage of the consensus read length. + pub majority_pct_detected: f64, /// Status of the evidence that supports (or does not support) this - /// read length, if available. + /// read length. pub evidence: Vec<(usize, usize)>, } @@ -27,7 +27,7 @@ impl DerivedReadlenResult { pub fn new( succeeded: bool, consensus_read_length: Option, - majority_pct_detected: f32, + majority_pct_detected: f64, evidence: Vec<(usize, usize)>, ) -> Self { DerivedReadlenResult { @@ -45,7 +45,7 @@ impl DerivedReadlenResult { pub fn predict( read_lengths: HashMap, num_samples: usize, - majority_vote_cutoff: f32, + majority_vote_cutoff: f64, ) -> Result { if num_samples == 0 { bail!("No read lengths were detected in the file."); @@ -59,7 +59,7 @@ pub fn predict( let max_count = read_lengths[0].1; let consensus_read_length = max_read_length; - let majority_detected = max_count as f32 / num_samples as f32; + let majority_detected = max_count as f64 / num_samples as f64; let mut result = DerivedReadlenResult::new(false, None, majority_detected * 100.0, read_lengths); diff --git a/src/generate/command.rs b/src/generate/command.rs index 167e9fd..f106d30 100644 --- a/src/generate/command.rs +++ b/src/generate/command.rs @@ -30,8 +30,8 @@ pub struct GenerateArgs { reference_providers: Vec, /// The error rate for the sequencer as a fraction between [0.0, 1.0] (per base). - #[arg(short, long, value_name = "F32", default_value = "0.0001")] - error_rate: f32, + #[arg(short, long, value_name = "F64", default_value = "0.0001")] + error_rate: f64, /// Specifies the number of records to generate. #[arg(short, long, value_name = "USIZE", conflicts_with = "coverage")] diff --git a/src/utils/args.rs b/src/utils/args.rs index 4aa452f..094819a 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -80,7 +80,7 @@ impl From for CompressionLevel { /// Utility method to parse command line floats and ensure they are /// within the range [MIN, MAX]. -pub fn arg_in_range(arg: f32, range: std::ops::RangeInclusive) -> anyhow::Result { +pub fn arg_in_range(arg: f64, range: std::ops::RangeInclusive) -> anyhow::Result { match range.contains(&arg) { true => Ok(arg), false => anyhow::bail!( From 8f90308c4aaad2f25ebe7507edf66bf9c934a66a Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 15:27:54 -0500 Subject: [PATCH 60/91] feat: add a log_every param to RecordCounter --- src/convert/bam.rs | 4 +-- src/convert/cram.rs | 4 +-- src/convert/sam.rs | 4 +-- src/derive/command/encoding.rs | 2 +- src/derive/command/endedness.rs | 2 +- src/derive/command/junction_annotation.rs | 2 +- src/derive/command/readlen.rs | 2 +- src/derive/strandedness/compute.rs | 3 ++ src/index/bam.rs | 2 +- src/qc/command.rs | 4 +-- src/utils/display.rs | 35 +++++++++++++++++------ 11 files changed, 42 insertions(+), 22 deletions(-) diff --git a/src/convert/bam.rs b/src/convert/bam.rs index 59068dd..153c52e 100644 --- a/src/convert/bam.rs +++ b/src/convert/bam.rs @@ -45,7 +45,7 @@ pub async fn to_sam_async( .await .with_context(|| "writing SAM header")?; - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut record = Record::default(); // (4) Write each record in the BAM file to the SAM file. @@ -131,7 +131,7 @@ pub async fn to_cram_async( .await .with_context(|| "writing CRAM file header")?; - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut record = Record::default(); // (6) Write each record in the BAM file to the CRAM file. diff --git a/src/convert/cram.rs b/src/convert/cram.rs index f9045e9..6679cc1 100644 --- a/src/convert/cram.rs +++ b/src/convert/cram.rs @@ -52,7 +52,7 @@ pub async fn to_sam_async( .with_context(|| "writing SAM header")?; // (5) Write each record in the CRAM file to the SAM file. - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut records = reader.records(&repository, &header.parsed); while let Some(record) = records @@ -125,7 +125,7 @@ pub async fn to_bam_async( .with_context(|| "writing BAM reference sequences")?; // (6) Write each record in the CRAM file to the BAM file. - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut records = reader.records(&repository, &header.parsed); while let Some(record) = records diff --git a/src/convert/sam.rs b/src/convert/sam.rs index 697e52b..ea0bddb 100644 --- a/src/convert/sam.rs +++ b/src/convert/sam.rs @@ -58,7 +58,7 @@ pub async fn to_bam_async( .await .with_context(|| "writing BAM reference sequences")?; - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut record = Record::default(); // (5) Write each record in the BAM file to the SAM file. @@ -151,7 +151,7 @@ pub async fn to_cram_async( .await .with_context(|| "writing CRAM file header")?; - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut record = Record::default(); // (6) Write each record in the SAM file to the CRAM file. diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index 31a2965..a81c618 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -45,7 +45,7 @@ pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { // (1) Collect quality scores from reads within the // file. Support for sampling only a portion of the reads is provided. let num_records = NumberOfRecords::from(args.num_records); - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); for result in reader.lazy_records() { let record = result?; diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 7334e2a..247f332 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -71,7 +71,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (1) Collect ordering flags (and QNAMEs) from reads within the // file. Support for sampling only a portion of the reads is provided. let num_records = NumberOfRecords::from(args.num_records); - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 578f827..5b29b64 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -102,7 +102,7 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { debug!("Done reading GFF."); // (1.5) Initialize variables (including opening the BAM). - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); let mut results = JunctionAnnotationResults::default(); let params = compute::JunctionAnnotationParameters { min_intron_length: args.min_intron_length, diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 668741a..903f56a 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -49,7 +49,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. let num_records = NumberOfRecords::from(args.num_records); - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index a939065..f1e0abb 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -14,6 +14,7 @@ use std::sync::Arc; use crate::derive::strandedness::results; use crate::utils::alignment::filter_by_mapq; +use crate::utils::display::RecordCounter; use crate::utils::read_groups::{validate_read_group_info, UNKNOWN_READ_GROUP}; const STRANDED_THRESHOLD: f64 = 80.0; @@ -323,6 +324,7 @@ pub fn predict( ) -> Result { let mut rng = rand::thread_rng(); let mut num_tested_genes: usize = 0; // Local to this attempt + let mut counter = RecordCounter::new(Some(1_000)); let genes_remaining = gene_records.len(); let max_iters = if params.max_iterations_per_try > genes_remaining { @@ -346,6 +348,7 @@ pub fn predict( } let cur_gene = gene_records.swap_remove(rng.gen_range(0..gene_records.len())); + counter.inc(); if disqualify_gene(&cur_gene, exons) { metrics.genes.bad_strands += 1; diff --git a/src/index/bam.rs b/src/index/bam.rs index d9e3b4a..c663ac1 100644 --- a/src/index/bam.rs +++ b/src/index/bam.rs @@ -69,7 +69,7 @@ pub fn index(src: PathBuf) -> anyhow::Result<()> { let mut builder = bai::Index::builder(); let mut start_position = reader.virtual_position(); - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); loop { match reader.read_record(&header.parsed, &mut record) { diff --git a/src/qc/command.rs b/src/qc/command.rs index 0f95bac..d628c00 100644 --- a/src/qc/command.rs +++ b/src/qc/command.rs @@ -300,7 +300,7 @@ fn app( //====================================================================// info!("Starting first pass for QC stats."); - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; @@ -351,7 +351,7 @@ fn app( let index = bai::read(src.with_extension("bam.bai")).with_context(|| "reading BAM index")?; - let mut counter = RecordCounter::new(); + let mut counter = RecordCounter::default(); for (name, seq) in header.parsed.reference_sequences() { let start = Position::MIN; diff --git a/src/utils/display.rs b/src/utils/display.rs index 2a58111..7ee4f56 100644 --- a/src/utils/display.rs +++ b/src/utils/display.rs @@ -24,29 +24,46 @@ impl fmt::Display for PercentageFormat { } /// Utility struct used to uniformly count and report the number of records processed. -#[derive(Default)] -pub struct RecordCounter(usize); +pub struct RecordCounter { + /// The number of records processed. + count: usize, + + /// The number of records to log every. + log_every: usize, +} + +impl Default for RecordCounter { + fn default() -> Self { + RecordCounter { + count: 0, + log_every: 1_000_000, + } + } +} impl RecordCounter { /// Creates a new `RecordCounter`. - pub fn new() -> Self { - Self::default() + pub fn new(log_every: Option) -> Self { + RecordCounter { + count: 0, + log_every: log_every.unwrap_or(1_000_000), + } } /// Gets the current number of records counted via a copy. pub fn get(&self) -> usize { - self.0 + self.count } /// Increments the counter and reports the number of records processed (if /// appropriate). pub fn inc(&mut self) { - self.0 += 1; + self.count += 1; - if self.0 % 1_000_000 == 0 { + if self.count % self.log_every == 0 { info!( " [*] Processed {} records.", - self.0.to_formatted_string(&Locale::en), + self.count.to_formatted_string(&Locale::en), ); } } @@ -57,7 +74,7 @@ impl RecordCounter { /// (if it exists, otherwise it loops forever). pub fn time_to_break(&self, limit: &NumberOfRecords) -> bool { match limit { - NumberOfRecords::Some(v) => self.0 >= *v, + NumberOfRecords::Some(v) => self.count >= *v, NumberOfRecords::All => false, } } From 85d9f193531cbe4ae5ffac5ac5798fbef8a858bb Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 15:31:43 -0500 Subject: [PATCH 61/91] style: code clean up --- src/derive/command/strandedness.rs | 39 +++++++++++++++++++++--------- src/derive/strandedness/compute.rs | 35 +++++++++++++-------------- src/derive/strandedness/results.rs | 15 +++++++++--- 3 files changed, 56 insertions(+), 33 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 304c0d7..deb9d93 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -41,7 +41,12 @@ pub struct DeriveStrandednessArgs { #[arg(short = 'm', long, value_name = "USIZE", default_value = "10")] min_reads_per_gene: usize, - /// How many genes to sample. + /// How many genes to use as evidence in strandendess classification per try. + /// This does not count genes which fail filtering + /// due to `--min-reads-per-gene` or are discarded + /// due to problematic Strand information in the GFF. + /// Problematic Strand information is caused by contradictions between + /// gene entries and overlapping exon entries. #[arg(short = 'n', long, value_name = "USIZE", default_value = "1000")] num_genes: usize, @@ -53,7 +58,7 @@ pub struct DeriveStrandednessArgs { /// however, 255 is reserved by the spec for a missing MAPQ value. /// Therefore BAMs produced by aligners using 255 erroneously /// are not compatible with setting this option. - #[arg(short, long, value_name = "U8")] + #[arg(long, value_name = "U8")] min_mapq: Option, /// Consider all genes, not just protein coding genes. @@ -84,10 +89,10 @@ pub struct DeriveStrandednessArgs { #[arg(long)] count_qc_failed: bool, - /// At most, search this many times for genes that satisfy our search criteria. - /// Default is 10 * --num-genes. + /// At most, evaluate this many genes + /// per try. Default is 10 * --num-genes. #[arg(long, value_name = "USIZE")] - max_iterations_per_try: Option, + max_genes_per_try: Option, } /// Main function for the `ngs derive strandedness` subcommand. @@ -106,7 +111,9 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { for result in gff.records() { let record = result.unwrap(); if record.ty() == args.gene_feature_name { - // If --all-genes is set, keep the record. + gene_metrics.total += 1; + + // If --all-genes is set, don't check the gene type or biotype. // Otherwise, check the gene type or biotype and keep the record if it's protein coding. // If the record does not have a gene type or biotype, discard it. let mut keep_record = false; @@ -127,10 +134,20 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { } } } - gene_metrics.total += 1; - if keep_record { - gene_records.push(record); + if !keep_record { + continue; } + + // Make sure the gene record has a valid strand. + let gene_strand = record.strand(); + if gene_strand != gff::record::Strand::Forward + && gene_strand != gff::record::Strand::Reverse + { + gene_metrics.bad_strand += 1; + continue; + } + + gene_records.push(record); } else if record.ty() == args.exon_feature_name { exon_metrics.total += 1; exon_records.push(record); @@ -204,11 +221,11 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { index, }; - let max_iterations_per_try = args.max_iterations_per_try.unwrap_or(args.num_genes * 10); + let max_genes_per_try = args.max_genes_per_try.unwrap_or(args.num_genes * 10); let params = compute::StrandednessParams { num_genes: args.num_genes, - max_iterations_per_try, + max_genes_per_try, min_reads_per_gene: args.min_reads_per_gene, min_mapq: args.min_mapq, count_qc_failed: args.count_qc_failed, diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index f1e0abb..d191769 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -110,7 +110,7 @@ pub struct StrandednessParams { pub num_genes: usize, /// The maximum number of iterations to try before giving up. - pub max_iterations_per_try: usize, + pub max_genes_per_try: usize, /// Minimum number of reads mapped to a gene to be considered /// for evidence of strandedness. @@ -138,10 +138,8 @@ fn disqualify_gene( gene: &gff::Record, exons: &HashMap<&str, Lapper>, ) -> bool { + // gene_strand guaranteed to be Forward or Reverse by initialization code. let gene_strand = gene.strand(); - if gene_strand != gff::record::Strand::Forward && gene_strand != gff::record::Strand::Reverse { - return true; - } let mut all_on_same_strand = true; let mut at_least_one_exon = false; @@ -323,26 +321,26 @@ pub fn predict( metrics: &mut results::RecordTracker, ) -> Result { let mut rng = rand::thread_rng(); - let mut num_tested_genes: usize = 0; // Local to this attempt + let mut num_genes_considered: usize = 0; // Local to this attempt let mut counter = RecordCounter::new(Some(1_000)); let genes_remaining = gene_records.len(); - let max_iters = if params.max_iterations_per_try > genes_remaining { + let max_iters = if params.max_genes_per_try > genes_remaining { tracing::warn!( "The number of genes remaining ({}) is less than the maximum iterations per try ({}).", genes_remaining, - params.max_iterations_per_try, + params.max_genes_per_try, ); genes_remaining } else { - params.max_iterations_per_try + params.max_genes_per_try }; for _ in 0..max_iters { - if num_tested_genes >= params.num_genes { + if num_genes_considered >= params.num_genes { tracing::info!( - "Reached the maximum number of genes ({}) for this try.", - num_tested_genes, + "Reached the maximum number of considered genes ({}) for this try.", + num_genes_considered, ); break; } @@ -351,7 +349,7 @@ pub fn predict( counter.inc(); if disqualify_gene(&cur_gene, exons) { - metrics.genes.bad_strands += 1; + metrics.genes.mixed_strands += 1; // Tracked across attempts continue; } let cur_gene_strand = Strand::try_from(cur_gene.strand()).unwrap(); @@ -363,21 +361,22 @@ pub fn predict( classify_read(&read, &cur_gene_strand, all_counts, &mut metrics.reads); } if enough_reads { - num_tested_genes += 1; + num_genes_considered += 1; } else { - metrics.genes.not_enough_reads += 1; + metrics.genes.not_enough_reads += 1; // Tracked across attempts } } - if num_tested_genes < params.num_genes { + if num_genes_considered < params.num_genes { tracing::warn!( - "Reached the maximum number of iterations ({}) before testing the requested amount of genes ({}) for this try. Only tested {} genes.", + "Reached the maximum number of iterations ({}) before considering the requested amount of genes ({}) for this try. Only considering {} genes.", max_iters, params.num_genes, - num_tested_genes, + num_genes_considered, ); } - metrics.genes.tested += num_tested_genes; // Add to any other attempts + metrics.genes.considered += num_genes_considered; // Add to any other attempts + metrics.genes.evaluated += counter.get(); // Add to any other attempts // TODO: Should this be done in derive()? Will re-run for each attempt. // Might cause false positives? diff --git a/src/derive/strandedness/results.rs b/src/derive/strandedness/results.rs index 3b94963..7bd294b 100644 --- a/src/derive/strandedness/results.rs +++ b/src/derive/strandedness/results.rs @@ -32,16 +32,23 @@ pub struct GeneRecordMetrics { /// If --all-genes is set this will not be tallied. pub protein_coding: usize, - /// The number of genes tested. - pub tested: usize, + /// The number of genes which were discarded due to having + /// an unknown or missing strand. + pub bad_strand: usize, + + /// The number of genes randomly selected for evaluation. + pub evaluated: usize, /// The number of genes which were discarded due to having - /// an unknown/invalid strand OR with exons on both strands. - pub bad_strands: usize, + /// mixed strands (the gene has exons on both strands). + pub mixed_strands: usize, /// The number of genes which were discarded due to not having /// enough reads. pub not_enough_reads: usize, + + /// The number of genes considered for strandedness evidence. + pub considered: usize, } /// General exon metrics that are tallied as a part of the From 54dc2175d0032c11322a5f24aa83d5635482f912 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 16:56:55 -0500 Subject: [PATCH 62/91] style: use custom Strand enum more --- src/derive/command/strandedness.rs | 6 +++--- src/derive/strandedness/compute.rs | 32 +++++++++++++++--------------- 2 files changed, 19 insertions(+), 19 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index deb9d93..192e396 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -168,8 +168,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { // (2) Parse exon features into proper data structure. debug!("Tabulating GFF exon features."); - let mut exon_intervals: HashMap<&str, Vec>> = - HashMap::new(); + let mut exon_intervals: HashMap<&str, Vec>> = HashMap::new(); for record in &exon_records { let seq_name = record.reference_sequence_name(); let start: usize = record.start().into(); @@ -180,6 +179,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { exon_metrics.bad_strand += 1; continue; } + let strand = compute::Strand::try_from(strand).unwrap(); // above check guarantees safety exon_intervals.entry(seq_name).or_default().push(Interval { start, @@ -196,7 +196,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { exon_metrics.bad_strand ); - let mut exons: HashMap<&str, Lapper> = HashMap::new(); + let mut exons: HashMap<&str, Lapper> = HashMap::new(); for (seq_name, intervals) in exon_intervals { exons.insert(seq_name, Lapper::new(intervals)); } diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index d191769..eaef4d1 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -30,10 +30,13 @@ pub struct Counts { reverse: usize, } -/// Struct for tracking possible strand orientations. -#[derive(Clone, Copy, Debug)] -enum Strand { +/// Struct for possible (valid) strand orientations. +#[derive(Clone, Copy, Debug, PartialEq, Eq)] +pub enum Strand { + /// Forward strand. Forward, + + /// Reverse strand. Reverse, } @@ -134,12 +137,9 @@ pub struct StrandednessParams { } /// Function to disqualify a gene based on its strand and exons. -fn disqualify_gene( - gene: &gff::Record, - exons: &HashMap<&str, Lapper>, -) -> bool { +fn disqualify_gene(gene: &gff::Record, exons: &HashMap<&str, Lapper>) -> bool { // gene_strand guaranteed to be Forward or Reverse by initialization code. - let gene_strand = gene.strand(); + let gene_strand = Strand::try_from(gene.strand()).unwrap(); let mut all_on_same_strand = true; let mut at_least_one_exon = false; @@ -315,19 +315,19 @@ fn predict_strandedness( pub fn predict( parsed_bam: &mut ParsedBAMFile, gene_records: &mut Vec, - exons: &HashMap<&str, Lapper>, + exons: &HashMap<&str, Lapper>, all_counts: &mut AllReadGroupsCounts, params: &StrandednessParams, metrics: &mut results::RecordTracker, ) -> Result { let mut rng = rand::thread_rng(); - let mut num_genes_considered: usize = 0; // Local to this attempt + let mut num_genes_considered = 0; // Local to this attempt let mut counter = RecordCounter::new(Some(1_000)); let genes_remaining = gene_records.len(); let max_iters = if params.max_genes_per_try > genes_remaining { tracing::warn!( - "The number of genes remaining ({}) is less than the maximum iterations per try ({}).", + "The number of genes remaining ({}) is less than the --max-genes-per-try ({}).", genes_remaining, params.max_genes_per_try, ); @@ -368,7 +368,7 @@ pub fn predict( } if num_genes_considered < params.num_genes { tracing::warn!( - "Reached the maximum number of iterations ({}) before considering the requested amount of genes ({}) for this try. Only considering {} genes.", + "Evaluated the maximum number of genes ({}) before considering the requested amount of genes ({}) for this try. Only considering {} genes.", max_iters, params.num_genes, num_genes_considered, @@ -425,12 +425,12 @@ mod tests { Interval { start: 1, stop: 10, - val: gff::record::Strand::Forward, + val: Strand::Forward, }, Interval { start: 11, stop: 20, - val: gff::record::Strand::Reverse, + val: Strand::Reverse, }, ]), ); @@ -445,12 +445,12 @@ mod tests { Interval { start: 1, stop: 10, - val: gff::record::Strand::Forward, + val: Strand::Forward, }, Interval { start: 11, stop: 20, - val: gff::record::Strand::Forward, + val: Strand::Forward, }, ]), ); From 725b936f5f3f75c8ffc035c0d27a3a44b0d830b5 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 18:47:55 -0500 Subject: [PATCH 63/91] fix: junction-annotation works again --- src/derive/command/junction_annotation.rs | 3 ++- src/derive/endedness/compute.rs | 32 ++++++++++++++++++----- src/derive/junction_annotation/compute.rs | 11 ++++---- src/derive/strandedness/compute.rs | 25 +++++++++++++++--- 4 files changed, 56 insertions(+), 15 deletions(-) diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index 5b29b64..a39b899 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -88,8 +88,9 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { } exon_records.push(record); } + debug!("Read {} exon records.", exon_records.len()); - debug!("Tabulating GFF exon features."); + debug!("Processing GFF exon features."); for record in &exon_records { let seq_name = record.reference_sequence_name(); let start = record.start(); diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 0d1043e..823d9bb 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -3,6 +3,7 @@ use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; +use std::ops::{Add, AddAssign}; use std::sync::Arc; use tracing::warn; @@ -45,6 +46,30 @@ impl Default for OrderingFlagsCounts { } } +impl Add for OrderingFlagsCounts { + type Output = Self; + + fn add(self, other: Self) -> Self { + OrderingFlagsCounts { + unsegmented: self.unsegmented + other.unsegmented, + first: self.first + other.first, + last: self.last + other.last, + both: self.both + other.both, + neither: self.neither + other.neither, + } + } +} + +impl AddAssign for OrderingFlagsCounts { + fn add_assign(&mut self, other: Self) { + self.unsegmented += other.unsegmented; + self.first += other.first; + self.last += other.last; + self.both += other.both; + self.neither += other.neither; + } +} + /// Struct holding the per read group results for an `ngs derive endedness` /// subcommand call. #[derive(Debug, Serialize)] @@ -375,12 +400,7 @@ pub fn predict( let mut rg_results = Vec::new(); for (read_group, rg_ordering_flags) in ordering_flags.iter() { - // TODO can make prettier? - overall_flags.unsegmented += rg_ordering_flags.unsegmented; - overall_flags.first += rg_ordering_flags.first; - overall_flags.last += rg_ordering_flags.last; - overall_flags.both += rg_ordering_flags.both; - overall_flags.neither += rg_ordering_flags.neither; + overall_flags += rg_ordering_flags.clone(); let result = predict_endedness( read_group.to_string(), diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 6d71acd..3a67b9b 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -31,7 +31,8 @@ pub struct JunctionAnnotationParameters { pub min_read_support: usize, /// Minumum mapping quality for a record to be considered. - /// 0 if MAPQ shouldn't be considered. + /// `None` means no filtering by MAPQ. This also allows + /// for records _without_ a MAPQ to be counted. pub min_mapq: Option, /// Do not count supplementary alignments. @@ -148,7 +149,7 @@ pub fn process( }; // (8) Find introns - let cur_pos = start; + let mut cur_pos = start; for op in record.cigar().iter() { match op.kind() { // This is an intron. @@ -158,7 +159,7 @@ pub fn process( let intron_start = cur_pos; // Update cur_pos to the end of the intron. - cur_pos.checked_add(op.len()); + cur_pos = cur_pos.checked_add(op.len()).unwrap(); let intron_end = cur_pos; let junction: results::Junction = (intron_start, intron_end); @@ -216,9 +217,9 @@ pub fn process( junction, ) } - // Operations (beside Skip which is handled above) that increment the reference position. + // Operations that increment the reference position (beside Skip which is handled above). Kind::Match | Kind::Deletion | Kind::SequenceMatch | Kind::SequenceMismatch => { - cur_pos.checked_add(op.len()); + cur_pos = cur_pos.checked_add(op.len()).unwrap(); } // Operations that do not increment the reference position. _ => {} diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index eaef4d1..d2d05c6 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -10,6 +10,7 @@ use rand::Rng; use rust_lapper::Lapper; use std::collections::HashMap; use std::collections::HashSet; +use std::ops::{Add, AddAssign}; use std::sync::Arc; use crate::derive::strandedness::results; @@ -30,6 +31,24 @@ pub struct Counts { reverse: usize, } +impl Add for Counts { + type Output = Self; + + fn add(self, other: Self) -> Self { + Self { + forward: self.forward + other.forward, + reverse: self.reverse + other.reverse, + } + } +} + +impl AddAssign for Counts { + fn add_assign(&mut self, other: Self) { + self.forward += other.forward; + self.reverse += other.reverse; + } +} + /// Struct for possible (valid) strand orientations. #[derive(Clone, Copy, Debug, PartialEq, Eq)] pub enum Strand { @@ -120,7 +139,8 @@ pub struct StrandednessParams { pub min_reads_per_gene: usize, /// Minumum mapping quality for a record to be considered. - /// 0 if MAPQ shouldn't be considered. + /// `None` means no filtering by MAPQ. This also allows + /// for records _without_ a MAPQ to be counted. pub min_mapq: Option, /// Allow qc failed reads to be counted. @@ -391,8 +411,7 @@ pub fn predict( let mut overall_counts = Counts::default(); let mut rg_results = Vec::new(); for (rg, counts) in &all_counts.counts { - overall_counts.forward += counts.forward; - overall_counts.reverse += counts.reverse; + overall_counts += counts.clone(); let result = predict_strandedness(rg, counts); rg_results.push(result) From 9668a6ddad4d01d9262a240a1d4598bb84d7eff8 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 19:19:30 -0500 Subject: [PATCH 64/91] feat(derive/instrument): behave more like other derive commands --- src/derive/command/instrument.rs | 71 +++++++--------- src/derive/instrument/compute.rs | 134 ++++++++++++++++++------------- 2 files changed, 109 insertions(+), 96 deletions(-) diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 65ac0a0..26c102f 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -1,15 +1,17 @@ //! Functionality relating to the `ngs derive instrument` subcommand itself. use anyhow::bail; +use clap::Args; +use num_format::Locale; +use num_format::ToFormattedString; use std::collections::HashSet; use std::path::PathBuf; -use std::thread; - -use clap::Args; use tracing::info; use crate::derive::instrument::compute; use crate::derive::instrument::reads::IlluminaReadName; +use crate::utils::args::NumberOfRecords; +use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; @@ -23,36 +25,16 @@ pub struct DeriveInstrumentArgs { /// Only examine the first n records in the file. #[arg(short, long, value_name = "USIZE")] num_records: Option, - - /// Use a specific number of threads. - #[arg(short, long, value_name = "USIZE")] - threads: Option, -} - -/// Entrypoint for the `ngs derive instrument` subcommand. -pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { - let first_n_reads: Option = args.num_records; - let threads = match args.threads { - Some(t) => t, - None => thread::available_parallelism().map(usize::from)?, - }; - - info!( - "Starting derive instrument subcommand with {} threads.", - threads - ); - - let rt = tokio::runtime::Builder::new_multi_thread() - .worker_threads(threads) - .build()?; - - rt.block_on(app(args.src, first_n_reads)) } /// Main function for the `ngs derive instrument` subcommand. -async fn app(src: PathBuf, first_n_reads: Option) -> anyhow::Result<()> { +pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { + let src = args.src; let mut instrument_names = HashSet::new(); let mut flowcell_names = HashSet::new(); + let mut metrics = compute::RecordMetrics::default(); + + info!("Starting derive instrument subcommand."); let ParsedBAMFile { mut reader, header, .. @@ -60,12 +42,8 @@ async fn app(src: PathBuf, first_n_reads: Option) -> anyhow::Result<()> { // (1) Collect instrument names and flowcell names from reads within the // file. Support for sampling only a portion of the reads is provided. - let mut samples = 0; - let mut sample_max = 0; - - if let Some(s) = first_n_reads { - sample_max = s; - } + let num_records = NumberOfRecords::from(args.num_records); + let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; @@ -76,8 +54,10 @@ async fn app(src: PathBuf, first_n_reads: Option) -> anyhow::Result<()> { match name.parse::() { Ok(read) => { instrument_names.insert(read.instrument_name); + metrics.found_instrument_name += 1; if let Some(fc) = read.flowcell { flowcell_names.insert(fc); + metrics.found_flowcell_name += 1; } } Err(_) => { @@ -87,24 +67,33 @@ async fn app(src: PathBuf, first_n_reads: Option) -> anyhow::Result<()> { ); } } + } else { + metrics.bad_read_name += 1; } - if sample_max > 0 { - samples += 1; - if samples > sample_max { - break; - } + counter.inc(); + if counter.time_to_break(&num_records) { + break; } } + info!( + "Processed {} records.", + counter.get().to_formatted_string(&Locale::en) + ); + metrics.total_records = counter.get(); + metrics.unique_instrument_names = instrument_names.len(); + metrics.unique_flowcell_names = flowcell_names.len(); + // (2) Derive the predict instrument results based on these detected // instrument names and flowcell names. - let result = compute::predict(instrument_names, flowcell_names); + let mut result = compute::predict(instrument_names, flowcell_names); + result.records = metrics; // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); + println!("{}", output); Ok(()) } diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 48d0106..9bfd387 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -5,7 +5,7 @@ use std::collections::HashSet; use regex::Regex; use serde::Serialize; -use tracing::info; +use tracing::debug; use super::flowcells; use super::instruments; @@ -42,6 +42,31 @@ impl InstrumentDetectionResults { } } +/// Metrics related to how read records were processed. +#[derive(Debug, Default, Serialize)] +pub struct RecordMetrics { + /// The total number of records that were processed. + pub total_records: usize, + + /// The total number of records that couldn't be parsed + /// due to a missing or invalid read name. + pub bad_read_name: usize, + + /// The total number of records that contained a valid + /// instrument name in their read name. + pub found_instrument_name: usize, + + /// The total number of records that contained a valid + /// flowcell name in their read name. + pub found_flowcell_name: usize, + + /// The number of unique instrument names that were detected. + pub unique_instrument_names: usize, + + /// The number of unique flowcell names that were detected. + pub unique_flowcell_names: usize, +} + /// Struct holding the final results for an `ngs derive instrument` subcommand /// call. #[derive(Debug, Serialize)] @@ -57,11 +82,14 @@ pub struct DerivedInstrumentResult { pub confidence: String, /// Status of the evidence that supports (or lack thereof) these predicted - /// instruments, if available. + /// instruments, if available. pub evidence: Option, /// A general comment field, if available. pub comment: Option, + + /// Metrics related to how read records were processed. + pub records: RecordMetrics, } impl DerivedInstrumentResult { @@ -72,6 +100,7 @@ impl DerivedInstrumentResult { confidence: String, evidence: Option, comment: Option, + records: RecordMetrics, ) -> Self { DerivedInstrumentResult { succeeded, @@ -79,6 +108,20 @@ impl DerivedInstrumentResult { confidence, evidence, comment, + records, + } + } +} + +impl Default for DerivedInstrumentResult { + fn default() -> Self { + DerivedInstrumentResult { + succeeded: false, + instruments: None, + confidence: "unknown".to_string(), + evidence: None, + comment: None, + records: RecordMetrics::default(), } } } @@ -114,7 +157,7 @@ pub fn possible_instruments_for_query( } } - info!(" [*] {}, Possible Instruments: {:?}", query, result); + debug!(" [*] {}, Possible Instruments: {:?}", query, result); result } @@ -161,44 +204,34 @@ pub fn resolve_instrument_prediction( let possible_instruments_by_iid = iid_results.possible_instruments.unwrap_or_default(); let possible_instruments_by_fcid = fcid_results.possible_instruments.unwrap_or_default(); + let mut result = DerivedInstrumentResult::default(); + // (1) If the set of possible instruments as determined by the instrument id // is empty _and_ we have seen at least one machine, then the only possible // scenario is there are conflicting instrument ids. if possible_instruments_by_iid.is_empty() && iid_results.detected_at_least_one_machine { - return DerivedInstrumentResult::new( - false, - None, - "unknown".to_string(), - Some("instrument id".to_string()), - Some( - "multiple instruments were detected in this file via the instrument id".to_string(), - ), + result.evidence = Some("instrument id".to_string()); + result.comment = Some( + "multiple instruments were detected in this file via the instrument id".to_string(), ); + return result; } // (2) If the set of possible instruments as determined by the flowcell id // is empty _and_ we have seen at least one machine, then the only possible // scenario is there are conflicting flowcell ids. if possible_instruments_by_fcid.is_empty() && fcid_results.detected_at_least_one_machine { - return DerivedInstrumentResult::new( - false, - None, - "unknown".to_string(), - Some("flowcell id".to_string()), - Some("multiple instruments were detected in this file via the flowcell id".to_string()), - ); + result.evidence = Some("flowcell id".to_string()); + result.comment = + Some("multiple instruments were detected in this file via the flowcell id".to_string()); + return result; } // (3) if neither result turns up anything, then we can simply say that the // machine was not able to be detected. if possible_instruments_by_iid.is_empty() && possible_instruments_by_fcid.is_empty() { - return DerivedInstrumentResult::new( - false, - None, - "unknown".to_string(), - None, - Some("no matching instruments were found".to_string()), - ); + result.comment = Some("no matching instruments were found".to_string()); + return result; } // (4) If both aren't empty and iid_results _is_ empty, then the fcid @@ -211,13 +244,11 @@ pub fn resolve_instrument_prediction( _ => "low", }; - return DerivedInstrumentResult::new( - true, - Some(instruments), - confidence.to_string(), - Some("flowcell id".to_string()), - None, - ); + result.succeeded = true; + result.instruments = Some(instruments); + result.confidence = confidence.to_string(); + result.evidence = Some("flowcell id".to_string()); + return result; } // (5) Same as the block above, except now we are evaluating the opposite @@ -229,13 +260,11 @@ pub fn resolve_instrument_prediction( _ => "low", }; - return DerivedInstrumentResult::new( - true, - Some(instruments), - confidence.to_string(), - Some("instrument id".to_string()), - None, - ); + result.succeeded = true; + result.instruments = Some(instruments); + result.confidence = confidence.to_string(); + result.evidence = Some("instrument id".to_string()); + return result; } let overlapping_instruments: HashSet = possible_instruments_by_fcid @@ -244,26 +273,21 @@ pub fn resolve_instrument_prediction( .collect(); if overlapping_instruments.is_empty() { - return DerivedInstrumentResult::new( - false, - None, - "high".to_string(), - Some("instrument and flowcell id".to_string()), - Some( - "Case needs triaging, results from instrument id and \ + result.confidence = "high".to_string(); + result.evidence = Some("instrument and flowcell id".to_string()); + result.comment = Some( + "Case needs triaging, results from instrument id and \ flowcell id are mutually exclusive." - .to_string(), - ), + .to_string(), ); + return result; } - DerivedInstrumentResult::new( - true, - Some(overlapping_instruments), - "high".to_string(), - Some("instrument and flowcell id".to_string()), - None, - ) + result.succeeded = true; + result.instruments = Some(overlapping_instruments); + result.confidence = "high".to_string(); + result.evidence = Some("instrument and flowcell id".to_string()); + result } /// Main method to evaluate the detected instrument names and flowcell names and From f569b3898982d27e73c21c2c5addbeae7dac02bc Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 10 Feb 2024 19:20:21 -0500 Subject: [PATCH 65/91] fix(derive): print!(output) -> println!(output) --- src/derive/command/encoding.rs | 2 +- src/derive/command/endedness.rs | 2 +- src/derive/command/junction_annotation.rs | 2 +- src/derive/command/readlen.rs | 2 +- src/derive/command/strandedness.rs | 2 +- 5 files changed, 5 insertions(+), 5 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index a81c618..a85d30c 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -72,7 +72,7 @@ pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); + println!("{}", output); Ok(()) } diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 247f332..7532e06 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -145,7 +145,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); + println!("{}", output); anyhow::Ok(()) } diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index a39b899..c265867 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -136,7 +136,7 @@ pub fn derive(args: JunctionAnnotationArgs) -> anyhow::Result<()> { // (4) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&results).unwrap(); - print!("{}", output); + println!("{}", output); anyhow::Ok(()) } diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 903f56a..ccaa1a6 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -74,7 +74,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); + println!("{}", output); anyhow::Ok(()) } diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 192e396..7bbd3c2 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -275,7 +275,7 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { // (5) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); - print!("{}", output); + println!("{}", output); anyhow::Ok(()) } From 6a23773cdbd69fc426c5e9171001452522bb36a0 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 11 Feb 2024 09:53:45 -0500 Subject: [PATCH 66/91] fix(derive/strandedness): move RG validation out of compute --- src/derive/command/strandedness.rs | 19 +++++++++++++++++-- src/derive/strandedness/compute.rs | 14 ++------------ 2 files changed, 19 insertions(+), 14 deletions(-) diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index 7bbd3c2..bc10479 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -19,6 +19,7 @@ use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; use crate::derive::strandedness::results; use crate::utils::formats; +use crate::utils::read_groups::validate_read_group_info; /// Clap arguments for the `ngs derive strandedness` subcommand. #[derive(Args)] @@ -266,12 +267,26 @@ pub fn derive(args: DeriveStrandednessArgs) -> anyhow::Result<()> { info!("Strandedness test inconclusive."); } } - let result = result.unwrap(); + let mut result = result.unwrap(); if !result.succeeded { - info!("Strandedness test failed after {} tries.", args.max_tries); + info!( + "Strandedness test still failed after {} tries.", + args.max_tries + ); } + let rgs_in_header_not_found = + validate_read_group_info(&all_counts.found_rgs, &parsed_bam.header); + let mut empty_rg_results = Vec::new(); + for rg in rgs_in_header_not_found { + empty_rg_results.push(compute::predict_strandedness( + &rg, + &compute::Counts::default(), + )); + } + result.read_groups.extend(empty_rg_results); + // (5) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index d2d05c6..a880467 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -16,7 +16,7 @@ use std::sync::Arc; use crate::derive::strandedness::results; use crate::utils::alignment::filter_by_mapq; use crate::utils::display::RecordCounter; -use crate::utils::read_groups::{validate_read_group_info, UNKNOWN_READ_GROUP}; +use crate::utils::read_groups::UNKNOWN_READ_GROUP; const STRANDED_THRESHOLD: f64 = 80.0; const UNSTRANDED_THRESHOLD: f64 = 40.0; @@ -290,7 +290,7 @@ fn classify_read( } /// Method to predict the strandedness of a read group. -fn predict_strandedness( +pub fn predict_strandedness( rg_name: &str, counts: &Counts, ) -> results::ReadGroupDerivedStrandednessResult { @@ -398,16 +398,6 @@ pub fn predict( metrics.genes.considered += num_genes_considered; // Add to any other attempts metrics.genes.evaluated += counter.get(); // Add to any other attempts - // TODO: Should this be done in derive()? Will re-run for each attempt. - // Might cause false positives? - let rgs_in_header_not_found = - validate_read_group_info(&all_counts.found_rgs, &parsed_bam.header); - for rg in rgs_in_header_not_found { - all_counts - .counts - .insert(Arc::new(rg.to_string()), Counts::default()); - } - let mut overall_counts = Counts::default(); let mut rg_results = Vec::new(); for (rg, counts) in &all_counts.counts { From 9be385006d0d8d79c97721889224a99b4ff48e1b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 11 Feb 2024 09:54:30 -0500 Subject: [PATCH 67/91] tests(derive/endedness): fix the broken tests --- src/derive/endedness/compute.rs | 567 ++++++++++++++++---------------- 1 file changed, 281 insertions(+), 286 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 823d9bb..2b63d26 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -429,289 +429,284 @@ pub fn predict( ) } -// #[cfg(test)] -// mod tests { -// use super::*; - -// // TODO add tests for unsegmented reads -// #[test] -// fn test_predict_endedness() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict_endedness( -// "overall".to_string(), -// ordering_flags.get(&Arc::clone(&OVERALL)).unwrap(), -// 0.0, -// None, -// false, -// ); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// } - -// #[test] -// fn test_derive_endedness_from_all_zero_counts() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); -// let result = predict_endedness( -// String::from("rg1"), -// ordering_flags.get(&Arc::new(String::from("rg1"))).unwrap(), -// 0.0, -// None, -// false, -// ); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// } - -// #[test] -// fn test_derive_endedness_from_only_first() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 1, -// last: 0, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_last() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 0, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_both() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 0, -// last: 0, -// both: 1, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Single-End"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 1); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_only_neither() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 0, -// last: 0, -// both: 0, -// neither: 1, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 0); -// assert_eq!(result.last, 0); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 1); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last() { -// let mut ordering_flags: HashMap = HashMap::new(); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 1, -// last: 1, -// both: 0, -// neither: 0, -// }, -// ); -// let result = predict(ordering_flags, HashMap::new(), 0.0, false); -// assert!(result.succeeded); -// assert_eq!(result.endedness, "Paired-End"); -// assert_eq!(result.first, 1); -// assert_eq!(result.last, 1); -// assert_eq!(result.both, 0); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, None); -// assert_eq!(result.read_groups.len(), 0); -// } - -// #[test] -// fn test_calculate_reads_per_template() { -// let mut read_names: HashMap> = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let results = calculate_reads_per_template(read_names); -// assert_eq!(results.len(), 3); -// assert_eq!(results.get(&Arc::new("overall".to_string())).unwrap(), &2.2); -// assert_eq!(results.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); -// assert_eq!(results.get(&Arc::clone(&rg_single)).unwrap(), &1.0); -// } - -// #[test] -// fn test_derive_endedness_from_first_and_last_with_rpt() { -// let mut ordering_flags: HashMap = HashMap::new(); -// let rg_paired = Arc::new("rg_paired".to_string()); -// let rg_single = Arc::new("rg_single".to_string()); -// ordering_flags.insert( -// Arc::clone(&OVERALL), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 8, -// last: 8, -// both: 2, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_paired), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 8, -// last: 8, -// both: 0, -// neither: 0, -// }, -// ); -// ordering_flags.insert( -// Arc::clone(&rg_single), -// OrderingFlagsCounts { -// unsegmented: 0, -// first: 0, -// last: 0, -// both: 2, -// neither: 0, -// }, -// ); -// let mut read_names: HashMap> = HashMap::new(); -// read_names.insert( -// "read1".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read2".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); -// read_names.insert( -// "read4".to_string(), -// vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], -// ); -// read_names.insert( -// "read5".to_string(), -// vec![ -// Arc::clone(&rg_paired), -// Arc::clone(&rg_paired), -// Arc::clone(&rg_single), -// ], -// ); -// let result = predict(ordering_flags, read_names, 0.0, false); -// assert!(!result.succeeded); -// assert_eq!(result.endedness, "Unknown"); -// assert_eq!(result.first, 8); -// assert_eq!(result.last, 8); -// assert_eq!(result.both, 2); -// assert_eq!(result.neither, 0); -// assert_eq!(result.rpt, Some(2.2)); -// assert_eq!(result.read_groups.len(), 2); -// // We can't know which read group will be first in the vector. -// // But both should succeed. -// assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); -// } -// } +#[cfg(test)] +mod tests { + use super::*; + + // TODO add tests for unsegmented reads + #[test] + fn test_predict_endedness() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict_endedness( + "overall".to_string(), + ordering_flags + .get(&Arc::new("overall".to_string())) + .unwrap(), + 0.0, + None, + false, + ); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_all_zero_counts() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); + let result = predict_endedness( + String::from("rg1"), + ordering_flags.get(&Arc::new(String::from("rg1"))).unwrap(), + 0.0, + None, + false, + ); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_derive_endedness_from_only_first() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 1, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 1); + } + + #[test] + fn test_derive_endedness_from_only_last() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 0, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 1); + } + + #[test] + fn test_derive_endedness_from_only_both() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 0, + last: 0, + both: 1, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 1); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 1); + } + + #[test] + fn test_derive_endedness_from_only_neither() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 0, + last: 0, + both: 0, + neither: 1, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 1); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 1); + } + + #[test] + fn test_derive_endedness_from_first_and_last() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 0, + first: 1, + last: 1, + both: 0, + neither: 0, + }, + ); + let result = predict(ordering_flags, HashMap::new(), 0.0, false); + assert!(result.succeeded); + assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.first, 1); + assert_eq!(result.last, 1); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + assert_eq!(result.read_groups.len(), 1); + } + + #[test] + fn test_calculate_reads_per_template() { + let mut read_names: HashMap> = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let mut rg_rpts: HashMap = HashMap::new(); + let overall_rpt = calculate_reads_per_template(read_names, &mut rg_rpts); + assert_eq!(rg_rpts.len(), 2); + assert_eq!(overall_rpt, 2.2); + assert_eq!(rg_rpts.get(&Arc::clone(&rg_paired)).unwrap(), &2.0); + assert_eq!(rg_rpts.get(&Arc::clone(&rg_single)).unwrap(), &1.0); + } + + #[test] + fn test_derive_endedness_from_first_and_last_with_rpt() { + let mut ordering_flags: HashMap = HashMap::new(); + let rg_paired = Arc::new("rg_paired".to_string()); + let rg_single = Arc::new("rg_single".to_string()); + ordering_flags.insert( + Arc::clone(&rg_paired), + OrderingFlagsCounts { + unsegmented: 0, + first: 8, + last: 8, + both: 0, + neither: 0, + }, + ); + ordering_flags.insert( + Arc::clone(&rg_single), + OrderingFlagsCounts { + unsegmented: 2, + first: 0, + last: 0, + both: 0, + neither: 0, + }, + ); + let mut read_names: HashMap> = HashMap::new(); + read_names.insert( + "read1".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read2".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + read_names.insert("read3".to_string(), vec![Arc::clone(&rg_single)]); + read_names.insert( + "read4".to_string(), + vec![Arc::clone(&rg_paired), Arc::clone(&rg_paired)], + ); + read_names.insert( + "read5".to_string(), + vec![ + Arc::clone(&rg_paired), + Arc::clone(&rg_paired), + Arc::clone(&rg_single), + ], + ); + let result = predict(ordering_flags, read_names, 0.0, false); + assert!(!result.succeeded); + assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.unsegmented, 2); + assert_eq!(result.first, 8); + assert_eq!(result.last, 8); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, Some(2.2)); + assert_eq!(result.read_groups.len(), 2); + // We can't know which read group will be first in the vector. + // But both should succeed. + print!("{:?}", result.read_groups); + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); + } +} From 2eac8d742fb828fbbc9a3de5e041a26642cd30dc Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 11 Feb 2024 09:55:14 -0500 Subject: [PATCH 68/91] style(derive/junction-annotation): group reported junctions by contig --- src/derive/junction_annotation/results.rs | 17 ++++++++++++----- 1 file changed, 12 insertions(+), 5 deletions(-) diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index a3c945e..ced00da 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -41,35 +41,42 @@ pub struct JunctionAnnotations { pub unannotated_reference: JunctionsMap, } -// TODO: This is a temporary implementation. It should be replaced with something better. impl Serialize for JunctionAnnotations { fn serialize(&self, serializer: S) -> Result { let mut known = Vec::new(); for (ref_name, junctions) in &self.known { + let mut junctions_vec = Vec::new(); for ((start, end), count) in junctions { - known.push((ref_name, start.get(), end.get(), count)); + junctions_vec.push((start.get(), end.get(), count)); } + known.push((ref_name.clone(), junctions_vec)); } let mut partial_novel = Vec::new(); for (ref_name, junctions) in &self.partial_novel { + let mut junctions_vec = Vec::new(); for ((start, end), count) in junctions { - partial_novel.push((ref_name, start.get(), end.get(), count)); + junctions_vec.push((start.get(), end.get(), count)); } + partial_novel.push((ref_name.clone(), junctions_vec)); } let mut complete_novel = Vec::new(); for (ref_name, junctions) in &self.complete_novel { + let mut junctions_vec = Vec::new(); for ((start, end), count) in junctions { - complete_novel.push((ref_name, start.get(), end.get(), count)); + junctions_vec.push((start.get(), end.get(), count)); } + complete_novel.push((ref_name.clone(), junctions_vec)); } let mut unannotated_reference = Vec::new(); for (ref_name, junctions) in &self.unannotated_reference { + let mut junctions_vec = Vec::new(); for ((start, end), count) in junctions { - unannotated_reference.push((ref_name, start.get(), end.get(), count)); + junctions_vec.push((start.get(), end.get(), count)); } + unannotated_reference.push((ref_name.clone(), junctions_vec)); } let mut s = serializer.serialize_struct("JunctionAnnotations", 4)?; From b67bac8e503c8459bcb4e9a786299d7d8359a05b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 11 Feb 2024 13:09:40 -0500 Subject: [PATCH 69/91] fix: lots of code cleanup --- src/derive/command/endedness.rs | 34 +++-- src/derive/command/readlen.rs | 2 +- src/derive/endedness.rs | 1 + src/derive/endedness/compute.rs | 160 +++------------------- src/derive/endedness/results.rs | 119 ++++++++++++++++ src/derive/junction_annotation/compute.rs | 57 ++++---- src/derive/junction_annotation/results.rs | 1 + src/derive/readlen/compute.rs | 25 ++-- src/derive/strandedness/compute.rs | 71 +++++----- src/utils/read_groups.rs | 4 +- 10 files changed, 238 insertions(+), 236 deletions(-) create mode 100644 src/derive/endedness/results.rs diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 7532e06..a35b390 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -44,7 +44,7 @@ pub struct DeriveEndednessArgs { calc_rpt: bool, /// Round RPT to the nearest INT before comparing to expected values. - /// Appropriate if using `-n` > 0. + /// Appropriate if using `-n` > 0. Unrounded value is reported in output. #[arg(long, default_value = "false")] round_rpt: bool, } @@ -108,18 +108,26 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } } - if !record.flags().is_segmented() { - ordering_flags.entry(read_group).or_default().unsegmented += 1; - } else if record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(read_group).or_default().first += 1; - } else if !record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(read_group).or_default().last += 1; - } else if record.flags().is_first_segment() && record.flags().is_last_segment() { - ordering_flags.entry(read_group).or_default().both += 1; - } else if !record.flags().is_first_segment() && !record.flags().is_last_segment() { - ordering_flags.entry(read_group).or_default().neither += 1; - } else { - unreachable!(); + match ( + record.flags().is_segmented(), + record.flags().is_first_segment(), + record.flags().is_last_segment(), + ) { + (false, _, _) => { + ordering_flags.entry(read_group).or_default().unsegmented += 1; + } + (true, true, false) => { + ordering_flags.entry(read_group).or_default().first += 1; + } + (true, false, true) => { + ordering_flags.entry(read_group).or_default().last += 1; + } + (true, true, true) => { + ordering_flags.entry(read_group).or_default().both += 1; + } + (true, false, false) => { + ordering_flags.entry(read_group).or_default().neither += 1; + } } counter.inc(); diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index ccaa1a6..e6b0c51 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -55,7 +55,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let record = result?; let len = record.sequence().len(); - read_lengths.entry(len).and_modify(|e| *e += 1).or_insert(1); + *read_lengths.entry(len).or_default() += 1; counter.inc(); if counter.time_to_break(&num_records) { diff --git a/src/derive/endedness.rs b/src/derive/endedness.rs index bf321e0..6f8d5cc 100644 --- a/src/derive/endedness.rs +++ b/src/derive/endedness.rs @@ -1,3 +1,4 @@ //! Supporting functionality for the `ngs derive endedness` subcommand. pub mod compute; +pub mod results; diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 2b63d26..265fc2d 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,16 +1,16 @@ //! Module holding the logic for computing the endedness of a BAM. -use serde::Serialize; use std::collections::HashMap; use std::collections::HashSet; use std::ops::{Add, AddAssign}; use std::sync::Arc; use tracing::warn; +use crate::derive::endedness::results; use crate::utils::read_groups::ReadGroupPtr; /// Struct holding the ordering flags for a single read group. -#[derive(Debug, Clone)] +#[derive(Debug, Clone, Default)] pub struct OrderingFlagsCounts { /// The number of reads without 0x1 set. pub unsegmented: usize, @@ -40,12 +40,6 @@ impl OrderingFlagsCounts { } } -impl Default for OrderingFlagsCounts { - fn default() -> Self { - Self::new() - } -} - impl Add for OrderingFlagsCounts { type Output = Self; @@ -70,120 +64,6 @@ impl AddAssign for OrderingFlagsCounts { } } -/// Struct holding the per read group results for an `ngs derive endedness` -/// subcommand call. -#[derive(Debug, Serialize)] -pub struct ReadGroupDerivedEndednessResult { - /// Name of the read group. - pub read_group: String, - - /// Whether or not an endedness was determined for this read group. - pub succeeded: bool, - - /// The endedness of this read group or "Unknown". - pub endedness: String, - - /// The number of reads without 0x1 set. - pub unsegmented: usize, - - /// The f+l- read count. - pub first: usize, - - /// The f-l+ read count. - pub last: usize, - - /// The f+l+ read count. - pub both: usize, - - /// The f-l- read count. - pub neither: usize, - - /// The reads per template (RPT). - /// Only available if `args.calc_rpt` is true. - pub rpt: Option, -} - -impl ReadGroupDerivedEndednessResult { - /// Creates a new [`ReadGroupDerivedEndednessResult`]. - fn new( - read_group: String, - succeeded: bool, - endedness: String, - counts: OrderingFlagsCounts, - rpt: Option, - ) -> Self { - ReadGroupDerivedEndednessResult { - read_group, - succeeded, - endedness, - unsegmented: counts.unsegmented, - first: counts.first, - last: counts.last, - both: counts.both, - neither: counts.neither, - rpt, - } - } -} - -/// Struct holding the final results for an `ngs derive endedness` subcommand -/// call. -#[derive(Debug, Serialize)] -pub struct DerivedEndednessResult { - /// Whether or not the `ngs derive endedness` subcommand succeeded. - pub succeeded: bool, - - /// The overall endedness of the file or "Unknown". - pub endedness: String, - - /// The number of reads without 0x1 set. - pub unsegmented: usize, - - /// The overall f+l- read count. - pub first: usize, - - /// The overall f-l+ read count. - pub last: usize, - - /// The overall f+l+ read count. - pub both: usize, - - /// The overall f-l- read count. - pub neither: usize, - - /// The overall reads per template (RPT). - /// Only available if `args.calc_rpt` is true. - pub rpt: Option, - - /// Vector of [`ReadGroupDerivedEndednessResult`]s. - /// One for each read group in the BAM, - /// and potentially one for any reads with an unknown read group. - pub read_groups: Vec, -} - -impl DerivedEndednessResult { - /// Creates a new [`DerivedEndednessResult`]. - pub fn new( - succeeded: bool, - endedness: String, - counts: OrderingFlagsCounts, - rpt: Option, - read_groups: Vec, - ) -> Self { - DerivedEndednessResult { - succeeded, - endedness, - unsegmented: counts.unsegmented, - first: counts.first, - last: counts.last, - both: counts.both, - neither: counts.neither, - rpt, - read_groups, - } - } -} - /// Calculate the reads per template overall and for each read group. fn calculate_reads_per_template( read_names: HashMap>, @@ -204,17 +84,16 @@ fn calculate_reads_per_template( let read_group_set: HashSet = read_groups.iter().cloned().collect(); if read_group_set.len() == 1 { + // All found read groups assigned to this QNAME are the same. + // We assume this means all the reads came from the same template. let read_group = Arc::clone(read_group_set.iter().next().unwrap()); - read_group_reads - .entry(Arc::clone(&read_group)) - .and_modify(|e| *e += num_reads) - .or_insert(num_reads); - read_group_templates - .entry(read_group) - .and_modify(|e| *e += 1) - .or_insert(1); + *read_group_reads.entry(Arc::clone(&read_group)).or_default() += num_reads; + *read_group_templates.entry(read_group).or_default() += 1; } else { + // The QNAME is in multiple read groups. + // We assume this means the reads came from multiple templates. + // More specifically, we assume that exactly one template will originate from each read group. warning_count += 1; match warning_count { 1..=100 => { @@ -230,16 +109,10 @@ fn calculate_reads_per_template( } for read_group in read_groups { - read_group_reads - .entry(Arc::clone(read_group)) - .and_modify(|e| *e += 1) - .or_insert(1); + *read_group_reads.entry(Arc::clone(read_group)).or_default() += 1; } for read_group in read_group_set { - read_group_templates - .entry(read_group) - .and_modify(|e| *e += 1) - .or_insert(1); + *read_group_templates.entry(read_group).or_default() += 1; } } } @@ -268,7 +141,7 @@ fn predict_endedness( paired_deviance: f64, reads_per_template: Option, round_rpt: bool, -) -> ReadGroupDerivedEndednessResult { +) -> results::ReadGroupDerivedEndednessResult { let unsegmented = rg_ordering_flags.unsegmented; let first = rg_ordering_flags.first; let last = rg_ordering_flags.last; @@ -282,7 +155,7 @@ fn predict_endedness( "No reads were detected in this read group: {}", read_group_name ); - return ReadGroupDerivedEndednessResult::new( + return results::ReadGroupDerivedEndednessResult::new( read_group_name, false, "Unknown".to_string(), @@ -291,7 +164,7 @@ fn predict_endedness( ); } - let mut result = ReadGroupDerivedEndednessResult::new( + let mut result = results::ReadGroupDerivedEndednessResult::new( read_group_name, false, "Unknown".to_string(), @@ -331,6 +204,7 @@ fn predict_endedness( } // only both present if first == 0 && last == 0 && both > 0 && neither == 0 { + // Prior logic (before addition of unsegmented checks) left as comment for posterity // match reads_per_template { // Some(rpt) => { // if rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { @@ -389,7 +263,7 @@ pub fn predict( read_names: HashMap>, paired_deviance: f64, round_rpt: bool, -) -> DerivedEndednessResult { +) -> results::DerivedEndednessResult { let mut rg_rpts: HashMap = HashMap::new(); let mut overall_rpt: Option = None; if !read_names.is_empty() { @@ -420,7 +294,7 @@ pub fn predict( round_rpt, ); - DerivedEndednessResult::new( + results::DerivedEndednessResult::new( overall_result.succeeded, overall_result.endedness, overall_flags, diff --git a/src/derive/endedness/results.rs b/src/derive/endedness/results.rs new file mode 100644 index 0000000..aef11c9 --- /dev/null +++ b/src/derive/endedness/results.rs @@ -0,0 +1,119 @@ +//! Module holding the results structs for the `ngs derive endedness` subcommand. + +use serde::Serialize; + +use crate::derive::endedness::compute::OrderingFlagsCounts; + +/// Struct holding the per read group results for an `ngs derive endedness` +/// subcommand call. +#[derive(Debug, Serialize)] +pub struct ReadGroupDerivedEndednessResult { + /// Name of the read group. + pub read_group: String, + + /// Whether or not an endedness was determined for this read group. + pub succeeded: bool, + + /// The endedness of this read group or "Unknown". + pub endedness: String, + + /// The number of reads without 0x1 set. + pub unsegmented: usize, + + /// The f+l- read count. + pub first: usize, + + /// The f-l+ read count. + pub last: usize, + + /// The f+l+ read count. + pub both: usize, + + /// The f-l- read count. + pub neither: usize, + + /// The reads per template (RPT). + /// Only available if `args.calc_rpt` is true. + pub rpt: Option, +} + +impl ReadGroupDerivedEndednessResult { + /// Creates a new [`ReadGroupDerivedEndednessResult`]. + pub fn new( + read_group: String, + succeeded: bool, + endedness: String, + counts: OrderingFlagsCounts, + rpt: Option, + ) -> Self { + ReadGroupDerivedEndednessResult { + read_group, + succeeded, + endedness, + unsegmented: counts.unsegmented, + first: counts.first, + last: counts.last, + both: counts.both, + neither: counts.neither, + rpt, + } + } +} + +/// Struct holding the final results for an `ngs derive endedness` subcommand +/// call. +#[derive(Debug, Serialize)] +pub struct DerivedEndednessResult { + /// Whether or not the `ngs derive endedness` subcommand succeeded. + pub succeeded: bool, + + /// The overall endedness of the file or "Unknown". + pub endedness: String, + + /// The number of reads without 0x1 set. + pub unsegmented: usize, + + /// The overall f+l- read count. + pub first: usize, + + /// The overall f-l+ read count. + pub last: usize, + + /// The overall f+l+ read count. + pub both: usize, + + /// The overall f-l- read count. + pub neither: usize, + + /// The overall reads per template (RPT). + /// Only available if `args.calc_rpt` is true. + pub rpt: Option, + + /// Vector of [`ReadGroupDerivedEndednessResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, +} + +impl DerivedEndednessResult { + /// Creates a new [`DerivedEndednessResult`]. + pub fn new( + succeeded: bool, + endedness: String, + counts: OrderingFlagsCounts, + rpt: Option, + read_groups: Vec, + ) -> Self { + DerivedEndednessResult { + succeeded, + endedness, + unsegmented: counts.unsegmented, + first: counts.first, + last: counts.last, + both: counts.both, + neither: counts.neither, + rpt, + read_groups, + } + } +} diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 3a67b9b..069a5dc 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -142,7 +142,7 @@ pub fn process( } // (7) Calculate the start position of this read. This will - // later be used to find the position of any introns. + // be used to find the position of any introns. let start = match record.alignment_start() { Some(s) => s, _ => bail!("Could not parse record's start position."), @@ -155,6 +155,7 @@ pub fn process( // This is an intron. Kind::Skip => { // Check that `op.len() >= params.min_intron_length` later, + // once all reads supporting short junctions have been collected // for better metric reporting. let intron_start = cur_pos; @@ -175,14 +176,10 @@ pub fn process( continue; } - let exon_starts = match exons.starts.get(seq_name) { - Some(starts) => starts, - _ => bail!("Could not find exon starts for contig: {}", seq_name), - }; - let exon_ends = match exons.ends.get(seq_name) { - Some(ends) => ends, - _ => bail!("Could not find exon ends for contig: {}", seq_name), - }; + // The following unwraps are safe because we checked that the reference + // sequence is annotated above. + let exon_starts = exons.starts.get(seq_name).unwrap(); + let exon_ends = exons.ends.get(seq_name).unwrap(); let mut intron_start_known = false; let mut intron_end_known = false; @@ -299,27 +296,29 @@ pub fn summarize( ); // Tally up observed junctions and spliced reads. - let mut juncs; - let mut support; - (juncs, support) = tally_junctions_and_support(&results.junction_annotations.known); - results.summary.known_junctions = juncs; - results.summary.known_junctions_read_support = support; - (juncs, support) = tally_junctions_and_support(&results.junction_annotations.partial_novel); - results.summary.partial_novel_junctions = juncs; - results.summary.partial_novel_junctions_read_support = support; - (juncs, support) = tally_junctions_and_support(&results.junction_annotations.complete_novel); - results.summary.complete_novel_junctions = juncs; - results.summary.complete_novel_junctions_read_support = support; - (juncs, support) = - tally_junctions_and_support(&results.junction_annotations.unannotated_reference); - results.summary.unannotated_reference_junctions = juncs; - results.summary.unannotated_reference_junctions_read_support = support; - - // Tally up total junctions and spliced reads. + ( + results.summary.known_junctions, + results.summary.known_junctions_read_support, + ) = tally_junctions_and_support(&results.junction_annotations.known); + ( + results.summary.partial_novel_junctions, + results.summary.partial_novel_junctions_read_support, + ) = tally_junctions_and_support(&results.junction_annotations.partial_novel); + ( + results.summary.complete_novel_junctions, + results.summary.complete_novel_junctions_read_support, + ) = tally_junctions_and_support(&results.junction_annotations.complete_novel); + ( + results.summary.unannotated_reference_junctions, + results.summary.unannotated_reference_junctions_read_support, + ) = tally_junctions_and_support(&results.junction_annotations.unannotated_reference); + + // Tally up total junctions. results.summary.total_junctions = results.summary.known_junctions + results.summary.partial_novel_junctions + results.summary.complete_novel_junctions + results.summary.unannotated_reference_junctions; + // Tally up total read support. results.summary.total_junctions_read_support = results.summary.known_junctions_read_support + results.summary.partial_novel_junctions_read_support + results.summary.complete_novel_junctions_read_support @@ -327,7 +326,7 @@ pub fn summarize( // Calculate percentages. let total_junctions = results.summary.total_junctions as f64 - - results.summary.unannotated_reference_junctions as f64; + - results.summary.unannotated_reference_junctions as f64; // exclude unannotated junctions from percentages results.summary.known_junctions_percent = results.summary.known_junctions as f64 / total_junctions * 100.0; results.summary.partial_novel_junctions_percent = @@ -336,15 +335,19 @@ pub fn summarize( results.summary.complete_novel_junctions as f64 / total_junctions * 100.0; // Calculate average read support. + // Total results.summary.average_junction_read_support = results.summary.total_junctions_read_support as f64 / results.summary.total_junctions as f64; + // Known results.summary.average_known_junction_read_support = results.summary.known_junctions_read_support as f64 / results.summary.known_junctions as f64; + // Partial Novel results.summary.average_partial_novel_junction_read_support = results.summary.partial_novel_junctions_read_support as f64 / results.summary.partial_novel_junctions as f64; + // Complete Novel results.summary.average_complete_novel_junction_read_support = results.summary.complete_novel_junctions_read_support as f64 / results.summary.complete_novel_junctions as f64; diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index ced00da..29daf20 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -41,6 +41,7 @@ pub struct JunctionAnnotations { pub unannotated_reference: JunctionsMap, } +// TODO should contigs be sorted? impl Serialize for JunctionAnnotations { fn serialize(&self, serializer: S) -> Result { let mut known = Vec::new(); diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 28b6f97..2867dbb 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -17,8 +17,8 @@ pub struct DerivedReadlenResult { /// The majority vote percentage of the consensus read length. pub majority_pct_detected: f64, - /// Status of the evidence that supports (or does not support) this - /// read length. + /// Status of the evidence that supports (or does not support) the + /// consensus read length. pub evidence: Vec<(usize, usize)>, } @@ -61,15 +61,20 @@ pub fn predict( let consensus_read_length = max_read_length; let majority_detected = max_count as f64 / num_samples as f64; - let mut result = - DerivedReadlenResult::new(false, None, majority_detected * 100.0, read_lengths); - - if majority_detected >= majority_vote_cutoff { - result.succeeded = true; - result.consensus_read_length = Some(consensus_read_length); + match majority_detected >= majority_vote_cutoff { + true => anyhow::Ok(DerivedReadlenResult::new( + true, + Some(consensus_read_length), + majority_detected * 100.0, + read_lengths, + )), + false => anyhow::Ok(DerivedReadlenResult::new( + false, + None, + majority_detected * 100.0, + read_lengths, + )), } - - anyhow::Ok(result) } #[cfg(test)] diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index a880467..f2cb477 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -1,22 +1,16 @@ //! Module holding the logic for computing the strandedness. -use noodles::bam; -use noodles::core::Region; -use noodles::gff; -use noodles::sam; -use noodles::sam::record::data::field::Tag; use noodles::sam::record::MappingQuality; +use noodles::{bam, core::Region, gff, sam}; use rand::Rng; use rust_lapper::Lapper; -use std::collections::HashMap; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; use std::ops::{Add, AddAssign}; use std::sync::Arc; use crate::derive::strandedness::results; -use crate::utils::alignment::filter_by_mapq; -use crate::utils::display::RecordCounter; -use crate::utils::read_groups::UNKNOWN_READ_GROUP; +use crate::utils::read_groups; +use crate::utils::{alignment::filter_by_mapq, display::RecordCounter}; const STRANDED_THRESHOLD: f64 = 80.0; const UNSTRANDED_THRESHOLD: f64 = 40.0; @@ -49,7 +43,7 @@ impl AddAssign for Counts { } } -/// Struct for possible (valid) strand orientations. +/// Struct for valid strand orientations. #[derive(Clone, Copy, Debug, PartialEq, Eq)] pub enum Strand { /// Forward strand. @@ -84,7 +78,10 @@ impl TryFrom for Strand { /// Struct for tracking the order of segments in a record. #[derive(Clone, Copy, Debug)] enum SegmentOrder { + /// The first segment in a record. First, + + /// The last segment in a record. Last, } @@ -105,6 +102,8 @@ impl TryFrom for SegmentOrder { } /// Struct holding the parsed BAM file and its index. +/// TODO this code is repeated. Should be in a common module. +/// Will be moved to utils::formats in a future PR. pub struct ParsedBAMFile { /// The BAM reader. pub reader: bam::Reader>, @@ -131,7 +130,7 @@ pub struct StrandednessParams { /// The number of genes to test for strandedness. pub num_genes: usize, - /// The maximum number of iterations to try before giving up. + /// The maximum number of genes to try before giving up. pub max_genes_per_try: usize, /// Minimum number of reads mapped to a gene to be considered @@ -139,7 +138,7 @@ pub struct StrandednessParams { pub min_reads_per_gene: usize, /// Minumum mapping quality for a record to be considered. - /// `None` means no filtering by MAPQ. This also allows + /// `None` means no filtering by MAPQ. This allows /// for records _without_ a MAPQ to be counted. pub min_mapq: Option, @@ -156,22 +155,23 @@ pub struct StrandednessParams { pub count_duplicates: bool, } -/// Function to disqualify a gene based on its strand and exons. +/// Function to disqualify a gene based on its strand and its exons' strand. fn disqualify_gene(gene: &gff::Record, exons: &HashMap<&str, Lapper>) -> bool { // gene_strand guaranteed to be Forward or Reverse by initialization code. let gene_strand = Strand::try_from(gene.strand()).unwrap(); let mut all_on_same_strand = true; - let mut at_least_one_exon = false; - - if let Some(intervals) = exons.get(gene.reference_sequence_name()) { - for exon in intervals.find(gene.start().into(), gene.end().into()) { - at_least_one_exon = true; - if exon.val != gene_strand { - all_on_same_strand = false; - break; - } - } - } + + let at_least_one_exon = match exons.get(gene.reference_sequence_name()) { + Some(intervals) => intervals + .find(gene.start().into(), gene.end().into()) + .any(|exon| { + if exon.val != gene_strand { + all_on_same_strand = false; + } + true + }), + None => false, + }; if all_on_same_strand && at_least_one_exon { return false; @@ -242,16 +242,7 @@ fn classify_read( all_counts: &mut AllReadGroupsCounts, read_metrics: &mut results::ReadRecordMetrics, ) { - let read_group = match read.data().get(Tag::ReadGroup) { - Some(rg) => { - let rg = rg.to_string(); - if !all_counts.found_rgs.contains(&rg) { - all_counts.found_rgs.insert(Arc::new(rg.clone())); - } - Arc::clone(all_counts.found_rgs.get(&rg).unwrap()) - } - None => Arc::clone(&UNKNOWN_READ_GROUP), - }; + let read_group = read_groups::get_read_group(read, Some(&mut all_counts.found_rgs)); let rg_counts = all_counts.counts.entry(read_group).or_default(); @@ -324,7 +315,7 @@ pub fn predict_strandedness( { result.succeeded = true; result.strandedness = "Unstranded".to_string(); - } + } // else Inconclusive result } @@ -342,7 +333,7 @@ pub fn predict( ) -> Result { let mut rng = rand::thread_rng(); let mut num_genes_considered = 0; // Local to this attempt - let mut counter = RecordCounter::new(Some(1_000)); + let mut counter = RecordCounter::new(Some(1_000)); // Also local to this attempt let genes_remaining = gene_records.len(); let max_iters = if params.max_genes_per_try > genes_remaining { @@ -366,12 +357,13 @@ pub fn predict( } let cur_gene = gene_records.swap_remove(rng.gen_range(0..gene_records.len())); - counter.inc(); + counter.inc(); // Technically this is off-by-one, as the gene hasn't been processed yet. if disqualify_gene(&cur_gene, exons) { metrics.genes.mixed_strands += 1; // Tracked across attempts continue; } + // gene_strand guaranteed to be Forward or Reverse by initialization code. let cur_gene_strand = Strand::try_from(cur_gene.strand()).unwrap(); let mut enough_reads = false; @@ -388,7 +380,7 @@ pub fn predict( } if num_genes_considered < params.num_genes { tracing::warn!( - "Evaluated the maximum number of genes ({}) before considering the requested amount of genes ({}) for this try. Only considering {} genes.", + "Evaluated the maximum number of genes ({}) before considering the requested amount of genes ({}) for this try. Only considering an additional {} genes this try.", max_iters, params.num_genes, num_genes_considered, @@ -423,6 +415,7 @@ pub fn predict( #[cfg(test)] mod tests { use super::*; + use noodles::sam::record::data::field::Tag; use rust_lapper::Interval; #[test] diff --git a/src/utils/read_groups.rs b/src/utils/read_groups.rs index 7cd0b57..86caf5d 100644 --- a/src/utils/read_groups.rs +++ b/src/utils/read_groups.rs @@ -11,10 +11,8 @@ use tracing::warn; /// Type alias for a read group pointer. pub type ReadGroupPtr = Arc; -// Strings used to index into the HashMaps used to store the Read Group ordering flags. -// Lazy statics are used to save memory. lazy_static! { - /// String used to index into th HashMaps used to store the "unknown_read_group" ordering flags. + /// String used to represent an unknown read group. Wrapped in an Arc to prevent redundant memory usage. pub static ref UNKNOWN_READ_GROUP: ReadGroupPtr = Arc::new(String::from("unknown_read_group")); } From e1473a6d4108143b43560986ba74d67ea9bdaa88 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sun, 11 Feb 2024 16:53:47 -0500 Subject: [PATCH 70/91] tests(derive): all derive commands have tests --- src/derive/encoding/compute.rs | 247 +++++++++++++++++ src/derive/endedness/compute.rs | 49 +++- src/derive/junction_annotation/compute.rs | 319 +++++++++++----------- src/derive/junction_annotation/results.rs | 2 +- src/utils/alignment.rs | 16 +- 5 files changed, 447 insertions(+), 186 deletions(-) diff --git a/src/derive/encoding/compute.rs b/src/derive/encoding/compute.rs index eb60962..421ce79 100644 --- a/src/derive/encoding/compute.rs +++ b/src/derive/encoding/compute.rs @@ -77,3 +77,250 @@ pub fn predict(score_set: HashSet) -> Result = HashSet::new(); + score_set.insert(40); + score_set.insert(41); + score_set.insert(42); + score_set.insert(43); + score_set.insert(44); + score_set.insert(45); + score_set.insert(46); + score_set.insert(47); + score_set.insert(48); + score_set.insert(49); + score_set.insert(50); + score_set.insert(51); + score_set.insert(52); + score_set.insert(53); + score_set.insert(54); + score_set.insert(55); + score_set.insert(56); + score_set.insert(57); + score_set.insert(58); + score_set.insert(59); + score_set.insert(60); + score_set.insert(61); + score_set.insert(62); + score_set.insert(63); + score_set.insert(64); + score_set.insert(65); + score_set.insert(66); + score_set.insert(67); + score_set.insert(68); + score_set.insert(69); + score_set.insert(70); + score_set.insert(71); + score_set.insert(72); + score_set.insert(73); + score_set.insert(74); + score_set.insert(75); + score_set.insert(76); + score_set.insert(77); + score_set.insert(78); + score_set.insert(79); + score_set.insert(80); + score_set.insert(81); + score_set.insert(82); + score_set.insert(83); + score_set.insert(84); + score_set.insert(85); + score_set.insert(86); + score_set.insert(87); + score_set.insert(88); + score_set.insert(89); + score_set.insert(90); + score_set.insert(91); + score_set.insert(92); + score_set.insert(93); + + let result = predict(score_set).unwrap(); + assert!(result.succeeded); + assert_eq!(result.encoding, Some("Illumina 1.3".to_string())); + assert_eq!(result.observed_min, 40); + assert_eq!(result.observed_max, 93); + } + + #[test] + fn test_predict_illumina_1_0() { + let mut score_set: HashSet = HashSet::new(); + score_set.insert(26); + score_set.insert(27); + score_set.insert(28); + score_set.insert(29); + score_set.insert(30); + score_set.insert(31); + score_set.insert(32); + score_set.insert(33); + score_set.insert(34); + score_set.insert(35); + score_set.insert(36); + score_set.insert(37); + score_set.insert(38); + score_set.insert(39); + score_set.insert(40); + score_set.insert(41); + score_set.insert(42); + score_set.insert(43); + score_set.insert(44); + score_set.insert(45); + score_set.insert(46); + score_set.insert(47); + score_set.insert(48); + score_set.insert(49); + score_set.insert(50); + score_set.insert(51); + score_set.insert(52); + score_set.insert(53); + score_set.insert(54); + score_set.insert(55); + score_set.insert(56); + score_set.insert(57); + score_set.insert(58); + score_set.insert(59); + score_set.insert(60); + score_set.insert(61); + score_set.insert(62); + score_set.insert(63); + score_set.insert(64); + score_set.insert(65); + score_set.insert(66); + score_set.insert(67); + score_set.insert(68); + score_set.insert(69); + score_set.insert(70); + score_set.insert(71); + score_set.insert(72); + score_set.insert(73); + score_set.insert(74); + score_set.insert(75); + score_set.insert(76); + score_set.insert(77); + score_set.insert(78); + score_set.insert(79); + score_set.insert(80); + score_set.insert(81); + score_set.insert(82); + score_set.insert(83); + score_set.insert(84); + score_set.insert(85); + score_set.insert(86); + score_set.insert(87); + score_set.insert(88); + score_set.insert(89); + score_set.insert(90); + score_set.insert(91); + score_set.insert(92); + score_set.insert(93); + + let result = predict(score_set).unwrap(); + assert!(result.succeeded); + assert_eq!(result.encoding, Some("Illumina 1.0".to_string())); + assert_eq!(result.observed_min, 26); + assert_eq!(result.observed_max, 93); + } + + #[test] + fn test_predict_sanger() { + let mut score_set: HashSet = HashSet::new(); + score_set.insert(0); + score_set.insert(1); + score_set.insert(2); + score_set.insert(3); + score_set.insert(4); + score_set.insert(5); + score_set.insert(6); + score_set.insert(7); + score_set.insert(8); + score_set.insert(9); + score_set.insert(10); + score_set.insert(11); + score_set.insert(12); + score_set.insert(13); + score_set.insert(14); + score_set.insert(15); + score_set.insert(16); + score_set.insert(17); + score_set.insert(18); + score_set.insert(19); + score_set.insert(20); + score_set.insert(21); + score_set.insert(22); + score_set.insert(23); + score_set.insert(24); + score_set.insert(25); + score_set.insert(26); + score_set.insert(27); + score_set.insert(28); + score_set.insert(29); + score_set.insert(30); + score_set.insert(31); + score_set.insert(32); + score_set.insert(33); + score_set.insert(34); + score_set.insert(35); + score_set.insert(36); + score_set.insert(37); + score_set.insert(38); + score_set.insert(39); + score_set.insert(40); + score_set.insert(41); + score_set.insert(42); + score_set.insert(43); + score_set.insert(44); + score_set.insert(45); + score_set.insert(46); + score_set.insert(47); + score_set.insert(48); + score_set.insert(49); + score_set.insert(50); + score_set.insert(51); + score_set.insert(52); + score_set.insert(53); + score_set.insert(54); + score_set.insert(55); + score_set.insert(56); + score_set.insert(57); + score_set.insert(58); + score_set.insert(59); + score_set.insert(60); + score_set.insert(61); + score_set.insert(62); + score_set.insert(63); + score_set.insert(64); + score_set.insert(65); + score_set.insert(66); + score_set.insert(67); + score_set.insert(68); + + let result = predict(score_set).unwrap(); + assert!(result.succeeded); + assert_eq!(result.encoding, Some("Sanger/Illumina 1.8".to_string())); + assert_eq!(result.observed_min, 0); + assert_eq!(result.observed_max, 68); + } + + #[test] + fn test_predict_fail() { + let score_set: HashSet = HashSet::new(); + let result = predict(score_set); + assert!(result.is_err()); + } + + #[test] + fn test_predict_too_high_max_score() { + let mut score_set: HashSet = HashSet::new(); + score_set.insert(94); + let result = predict(score_set).unwrap(); + assert!(!result.succeeded); + assert_eq!(result.encoding, None); + assert_eq!(result.observed_min, 94); + assert_eq!(result.observed_max, 94); + } +} diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 265fc2d..4179d2e 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -307,9 +307,8 @@ pub fn predict( mod tests { use super::*; - // TODO add tests for unsegmented reads #[test] - fn test_predict_endedness() { + fn test_predict_endedness_from_first_and_last() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -340,7 +339,38 @@ mod tests { } #[test] - fn test_derive_endedness_from_all_zero_counts() { + fn test_predict_endedness_from_unsegmented() { + let mut ordering_flags: HashMap = HashMap::new(); + ordering_flags.insert( + Arc::new("overall".to_string()), + OrderingFlagsCounts { + unsegmented: 1, + first: 0, + last: 0, + both: 0, + neither: 0, + }, + ); + let result = predict_endedness( + "overall".to_string(), + ordering_flags + .get(&Arc::new("overall".to_string())) + .unwrap(), + 0.0, + None, + false, + ); + assert!(result.succeeded); + assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.first, 0); + assert_eq!(result.last, 0); + assert_eq!(result.both, 0); + assert_eq!(result.neither, 0); + assert_eq!(result.rpt, None); + } + + #[test] + fn test_predict_endedness_from_all_zero_counts() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert(Arc::new(String::from("rg1")), OrderingFlagsCounts::new()); let result = predict_endedness( @@ -360,7 +390,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_only_first() { + fn test_predict_from_only_first() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -384,7 +414,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_only_last() { + fn test_predict_from_only_last() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -408,7 +438,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_only_both() { + fn test_predict_from_only_both() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -432,7 +462,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_only_neither() { + fn test_predict_from_only_neither() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -456,7 +486,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_first_and_last() { + fn test_predict_from_first_and_last() { let mut ordering_flags: HashMap = HashMap::new(); ordering_flags.insert( Arc::new("overall".to_string()), @@ -518,7 +548,7 @@ mod tests { } #[test] - fn test_derive_endedness_from_first_and_last_with_rpt() { + fn test_predict_with_rpt_complex() { let mut ordering_flags: HashMap = HashMap::new(); let rg_paired = Arc::new("rg_paired".to_string()); let rg_single = Arc::new("rg_single".to_string()); @@ -580,7 +610,6 @@ mod tests { assert_eq!(result.read_groups.len(), 2); // We can't know which read group will be first in the vector. // But both should succeed. - print!("{:?}", result.read_groups); assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); } } diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 069a5dc..428daac 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -598,172 +598,157 @@ mod tests { assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); - // TODO: Below tests are not working as expected. Need to fix them. - // Test complete novel junction - // let mut record = Record::default(); - // let r6_name: ReadName = "complete1".parse().unwrap(); - // *record.read_name_mut() = Some(r6_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 6); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 0); - // assert_eq!(results.records.low_mapq, 0); - // assert_eq!(results.records.missing_mapq, 0); - - // // Test complete novel junction (again for more read support) - // let mut record = Record::default(); - // let r6_name: ReadName = "complete2".parse().unwrap(); - // *record.read_name_mut() = Some(r6_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 7); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 0); - // assert_eq!(results.records.low_mapq, 0); - // assert_eq!(results.records.missing_mapq, 0); - - // // Test fails MAPQ filter - // let mut record = Record::default(); - // let r7_name: ReadName = "low_mapq".parse().unwrap(); - // *record.read_name_mut() = Some(r7_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(20); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 6); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 0); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 0); - - // // Test missing MAPQ - // let mut record = Record::default(); - // let r8_name: ReadName = "missing_mapq".parse().unwrap(); - // *record.read_name_mut() = Some(r8_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(255); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 6); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 0); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 1); - - // // Test that intron is too short - // let mut record = Record::default(); - // let r9_name: ReadName = "short".parse().unwrap(); - // *record.read_name_mut() = Some(r9_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "5M5N5M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 0); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 1); - - // // Test that that reads not spliced are ignored - // let mut record = Record::default(); - // let r10_name: ReadName = "not_spliced".parse().unwrap(); - // *record.read_name_mut() = Some(r10_name); - // *record.reference_sequence_id_mut() = Some(0); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 7); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 1); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 1); - - // // Test unannoted reference - // let mut record = Record::default(); - // let r11_name: ReadName = "unannotated1".parse().unwrap(); - // *record.read_name_mut() = Some(r11_name); - // *record.reference_sequence_id_mut() = Some(1); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 8); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 1); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 1); - - // // Test unannoted reference (again for more read support) - // let mut record = Record::default(); - // let r11_name: ReadName = "unannotated2".parse().unwrap(); - // *record.read_name_mut() = Some(r11_name); - // *record.reference_sequence_id_mut() = Some(1); - // *record.alignment_start_mut() = Position::new(1); - // *record.cigar_mut() = "10M10N10M".parse().unwrap(); - // *record.mapping_quality_mut() = MappingQuality::new(60); - // record.flags_mut().set(0x4.into(), false); - // process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - // assert_eq!(results.records.processed, 9); - // assert_eq!(results.records.filtered_by_flags, 2); - // assert_eq!(results.records.not_spliced, 1); - // assert_eq!(results.records.low_mapq, 1); - // assert_eq!(results.records.missing_mapq, 1); - - // // Test summarize - // summarize(&mut results, ¶ms); - - // assert_eq!(results.summary.total_rejected_junctions, 1); - // assert_eq!(results.summary.intron_too_short, 1); - // assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); - // assert_eq!(results.summary.known_junctions, 1); - // assert_eq!(results.summary.known_junctions_read_support, 2); - // assert_eq!(results.summary.partial_novel_junctions, 1); - // assert_eq!(results.summary.partial_novel_junctions_read_support, 2); - // assert_eq!(results.summary.complete_novel_junctions, 1); - // assert_eq!(results.summary.complete_novel_junctions_read_support, 2); - // assert_eq!(results.summary.unannotated_reference_junctions, 1); - // assert_eq!( - // results.summary.unannotated_reference_junctions_read_support, - // 2 - // ); - // assert_eq!(results.summary.total_junctions, 4); - // assert_eq!(results.summary.total_junctions_read_support, 8); - // assert_eq!(results.summary.known_junctions_percent, 33.33333333333333); - // assert_eq!( - // results.summary.partial_novel_junctions_percent, - // 33.33333333333333 - // ); - // assert_eq!( - // results.summary.complete_novel_junctions_percent, - // 33.33333333333333 - // ); - // assert_eq!(results.summary.average_junction_read_support, 2.0); - // assert_eq!(results.summary.average_known_junction_read_support, 2.0); - // assert_eq!( - // results.summary.average_partial_novel_junction_read_support, - // 2.0 - // ); - // assert_eq!( - // results.summary.average_complete_novel_junction_read_support, - // 2.0 - // ); + // Test complete novel junction with 2 junctions + let mut record = Record::default(); + let r6_name: ReadName = "complete_twice1".parse().unwrap(); + *record.read_name_mut() = Some(r6_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(200); + *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 5); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 0); + + // Test complete novel junction with 2 junctions (again for more read support) + let mut record = Record::default(); + let r6_name: ReadName = "complete_twice2".parse().unwrap(); + *record.read_name_mut() = Some(r6_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(200); + *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 0); + + // Test fails MAPQ filter + let mut record = Record::default(); + let r7_name: ReadName = "low_mapq".parse().unwrap(); + *record.read_name_mut() = Some(r7_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(20); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 1); + + // Test missing MAPQ + let mut record = Record::default(); + let r8_name: ReadName = "bad_mapq".parse().unwrap(); + *record.read_name_mut() = Some(r8_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = None; + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 6); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 2); + + // Test that intron is too short + let mut record = Record::default(); + let r9_name: ReadName = "short".parse().unwrap(); + *record.read_name_mut() = Some(r9_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "5M5N5M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 2); + + // Test that that reads not spliced are ignored + let mut record = Record::default(); + let r10_name: ReadName = "not_spliced".parse().unwrap(); + *record.read_name_mut() = Some(r10_name); + *record.reference_sequence_id_mut() = Some(0); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 7); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.bad_mapq, 2); + + // Test unannoted reference + let mut record = Record::default(); + let r11_name: ReadName = "unannotated1".parse().unwrap(); + *record.read_name_mut() = Some(r11_name); + *record.reference_sequence_id_mut() = Some(1); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 8); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.bad_mapq, 2); + + // Test unannoted reference (again for more read support) + let mut record = Record::default(); + let r11_name: ReadName = "unannotated2".parse().unwrap(); + *record.read_name_mut() = Some(r11_name); + *record.reference_sequence_id_mut() = Some(1); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); + process(&record, &exons, &header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 9); + assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.not_spliced, 1); + assert_eq!(results.records.bad_mapq, 2); + + // Test summarize + summarize(&mut results, ¶ms); + + assert_eq!(results.summary.total_rejected_junctions, 1); + assert_eq!(results.summary.intron_too_short, 1); + assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); + assert_eq!(results.summary.known_junctions, 1); + assert_eq!(results.summary.known_junctions_read_support, 2); + assert_eq!(results.summary.partial_novel_junctions, 1); + assert_eq!(results.summary.partial_novel_junctions_read_support, 2); + assert_eq!(results.summary.complete_novel_junctions, 2); + assert_eq!(results.summary.complete_novel_junctions_read_support, 4); + assert_eq!(results.summary.unannotated_reference_junctions, 1); + assert_eq!( + results.summary.unannotated_reference_junctions_read_support, + 2 + ); + assert_eq!(results.summary.total_junctions, 5); + assert_eq!(results.summary.total_junctions_read_support, 10); + assert_eq!(results.summary.known_junctions_percent, 25.0); + assert_eq!(results.summary.partial_novel_junctions_percent, 25.0); + assert_eq!(results.summary.complete_novel_junctions_percent, 50.0); + assert_eq!(results.summary.average_junction_read_support, 2.0); + assert_eq!(results.summary.average_known_junction_read_support, 2.0); + assert_eq!( + results.summary.average_partial_novel_junction_read_support, + 2.0 + ); + assert_eq!( + results.summary.average_complete_novel_junction_read_support, + 2.0 + ); } } diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 29daf20..edd1dd3 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -113,7 +113,7 @@ pub struct RecordMetrics { } /// Summary statistics for the junction-annotation subcommand. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Default, Debug, Serialize)] pub struct SummaryResults { /// The total number of junctions observed in the file. pub total_junctions: usize, diff --git a/src/utils/alignment.rs b/src/utils/alignment.rs index c3cd230..678114f 100644 --- a/src/utils/alignment.rs +++ b/src/utils/alignment.rs @@ -14,9 +14,9 @@ pub fn filter_by_mapq( match min_mapq { Some(min_mapq) => match record.mapping_quality() { Some(mapq) => mapq.get() < min_mapq.get(), - None => false, + None => true, // Filter if no MAPQ is present }, - None => false, + None => false, // Do not filter if no min MAPQ is specified } } @@ -143,29 +143,29 @@ impl<'a> ReferenceRecordStepThrough<'a> { mod tests { use noodles::sam::record::{Cigar, MappingQuality, Sequence}; - use super::ReferenceRecordStepThrough; + use super::*; #[test] pub fn it_filters_by_mapq() -> anyhow::Result<()> { let mut record = noodles::sam::alignment::Record::default(); - assert!(super::filter_by_mapq( + assert!(filter_by_mapq( &record, Some(MappingQuality::new(0).unwrap()) )); // Get filtered because MAPQ is missing - assert!(!super::filter_by_mapq(&record, None)); // Do not get filtered because filter is disabled + assert!(!filter_by_mapq(&record, None)); // Do not get filtered because filter is disabled record .mapping_quality_mut() .replace(MappingQuality::new(10).unwrap()); - assert!(!super::filter_by_mapq( + assert!(!filter_by_mapq( &record, Some(MappingQuality::new(0).unwrap()) )); // Do not get filtered because MAPQ is present - assert!(!super::filter_by_mapq( + assert!(!filter_by_mapq( &record, Some(MappingQuality::new(1).unwrap()) )); // Do not get filtered because MAPQ is greater than 1 - assert!(super::filter_by_mapq( + assert!(filter_by_mapq( &record, Some(MappingQuality::new(11).unwrap()) )); // Do get filtered because MAPQ is less than 11 From 42d09dc4c544abe18f673611fc5643e9f5e5226b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 08:57:15 -0500 Subject: [PATCH 71/91] fix: consistently return Options for String results --- src/derive/encoding/compute.rs | 2 +- src/derive/endedness/compute.rs | 30 +++++++++++++++--------------- src/derive/endedness/results.rs | 12 ++++++------ src/derive/instrument/compute.rs | 2 +- src/derive/readlen/compute.rs | 2 +- src/derive/strandedness/compute.rs | 20 ++++++++++---------- src/derive/strandedness/results.rs | 12 ++++++------ 7 files changed, 40 insertions(+), 40 deletions(-) diff --git a/src/derive/encoding/compute.rs b/src/derive/encoding/compute.rs index 421ce79..8063fb7 100644 --- a/src/derive/encoding/compute.rs +++ b/src/derive/encoding/compute.rs @@ -16,7 +16,7 @@ pub struct DerivedEncodingResult { /// Whether or not the `ngs derive encoding` subcommand succeeded. pub succeeded: bool, - /// The detected quality score encoding, if available. + /// The detected quality score encoding, if derivable. pub encoding: Option, /// The minimum quality score observed. diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 4179d2e..d70633c 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -158,7 +158,7 @@ fn predict_endedness( return results::ReadGroupDerivedEndednessResult::new( read_group_name, false, - "Unknown".to_string(), + None, rg_ordering_flags.clone(), reads_per_template, ); @@ -167,7 +167,7 @@ fn predict_endedness( let mut result = results::ReadGroupDerivedEndednessResult::new( read_group_name, false, - "Unknown".to_string(), + None, rg_ordering_flags.clone(), reads_per_template, ); @@ -178,12 +178,12 @@ fn predict_endedness( Some(rpt) => { if rpt == 1.0 || (round_rpt && rpt.round() as usize == 1) { result.succeeded = true; - result.endedness = String::from("Single-End"); + result.endedness = Some(String::from("Single-End")); } } None => { result.succeeded = true; - result.endedness = String::from("Single-End"); + result.endedness = Some(String::from("Single-End")); } } return result; @@ -243,12 +243,12 @@ fn predict_endedness( Some(rpt) => { if rpt == 2.0 || (round_rpt && rpt.round() as usize == 2) { result.succeeded = true; - result.endedness = String::from("Paired-End"); + result.endedness = Some(String::from("Paired-End")); } } None => { result.succeeded = true; - result.endedness = String::from("Paired-End"); + result.endedness = Some(String::from("Paired-End")); } } } @@ -330,7 +330,7 @@ mod tests { false, ); assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.endedness, Some("Paired-End".to_string())); assert_eq!(result.first, 1); assert_eq!(result.last, 1); assert_eq!(result.both, 0); @@ -361,7 +361,7 @@ mod tests { false, ); assert!(result.succeeded); - assert_eq!(result.endedness, "Single-End"); + assert_eq!(result.endedness, Some("Single-End".to_string())); assert_eq!(result.first, 0); assert_eq!(result.last, 0); assert_eq!(result.both, 0); @@ -381,7 +381,7 @@ mod tests { false, ); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.first, 0); assert_eq!(result.last, 0); assert_eq!(result.both, 0); @@ -404,7 +404,7 @@ mod tests { ); let result = predict(ordering_flags, HashMap::new(), 0.0, false); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.first, 1); assert_eq!(result.last, 0); assert_eq!(result.both, 0); @@ -428,7 +428,7 @@ mod tests { ); let result = predict(ordering_flags, HashMap::new(), 0.0, false); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.first, 0); assert_eq!(result.last, 1); assert_eq!(result.both, 0); @@ -452,7 +452,7 @@ mod tests { ); let result = predict(ordering_flags, HashMap::new(), 0.0, false); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.first, 0); assert_eq!(result.last, 0); assert_eq!(result.both, 1); @@ -476,7 +476,7 @@ mod tests { ); let result = predict(ordering_flags, HashMap::new(), 0.0, false); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.first, 0); assert_eq!(result.last, 0); assert_eq!(result.both, 0); @@ -500,7 +500,7 @@ mod tests { ); let result = predict(ordering_flags, HashMap::new(), 0.0, false); assert!(result.succeeded); - assert_eq!(result.endedness, "Paired-End"); + assert_eq!(result.endedness, Some("Paired-End".to_string())); assert_eq!(result.first, 1); assert_eq!(result.last, 1); assert_eq!(result.both, 0); @@ -600,7 +600,7 @@ mod tests { ); let result = predict(ordering_flags, read_names, 0.0, false); assert!(!result.succeeded); - assert_eq!(result.endedness, "Unknown"); + assert_eq!(result.endedness, None); assert_eq!(result.unsegmented, 2); assert_eq!(result.first, 8); assert_eq!(result.last, 8); diff --git a/src/derive/endedness/results.rs b/src/derive/endedness/results.rs index aef11c9..43cddd4 100644 --- a/src/derive/endedness/results.rs +++ b/src/derive/endedness/results.rs @@ -14,8 +14,8 @@ pub struct ReadGroupDerivedEndednessResult { /// Whether or not an endedness was determined for this read group. pub succeeded: bool, - /// The endedness of this read group or "Unknown". - pub endedness: String, + /// The endedness of this read group, if derivable. + pub endedness: Option, /// The number of reads without 0x1 set. pub unsegmented: usize, @@ -42,7 +42,7 @@ impl ReadGroupDerivedEndednessResult { pub fn new( read_group: String, succeeded: bool, - endedness: String, + endedness: Option, counts: OrderingFlagsCounts, rpt: Option, ) -> Self { @@ -67,8 +67,8 @@ pub struct DerivedEndednessResult { /// Whether or not the `ngs derive endedness` subcommand succeeded. pub succeeded: bool, - /// The overall endedness of the file or "Unknown". - pub endedness: String, + /// The overall endedness, if derivable. + pub endedness: Option, /// The number of reads without 0x1 set. pub unsegmented: usize, @@ -99,7 +99,7 @@ impl DerivedEndednessResult { /// Creates a new [`DerivedEndednessResult`]. pub fn new( succeeded: bool, - endedness: String, + endedness: Option, counts: OrderingFlagsCounts, rpt: Option, read_groups: Vec, diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 9bfd387..59ce1ed 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -75,7 +75,7 @@ pub struct DerivedInstrumentResult { pub succeeded: bool, /// The possible instruments detected by `ngs derive instrument`, if - /// available. + /// derivable. pub instruments: Option>, /// The level of confidence that the tool has concerning these results. diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 2867dbb..97a3227 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -11,7 +11,7 @@ pub struct DerivedReadlenResult { /// Whether or not the `ngs derive readlen` subcommand succeeded. pub succeeded: bool, - /// The concsensus read length, if available. + /// The consensus read length, if derivable. pub consensus_read_length: Option, /// The majority vote percentage of the consensus read length. diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index f2cb477..6fffda5 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -289,7 +289,7 @@ pub fn predict_strandedness( return results::ReadGroupDerivedStrandednessResult { read_group: rg_name.to_string(), succeeded: false, - strandedness: "Inconclusive".to_string(), + strandedness: None, total: 0, forward: 0, reverse: 0, @@ -300,22 +300,22 @@ pub fn predict_strandedness( let mut result = results::ReadGroupDerivedStrandednessResult::new( rg_name.to_string(), false, - "Inconclusive".to_string(), + None, counts.forward, counts.reverse, ); if result.forward_pct > STRANDED_THRESHOLD { result.succeeded = true; - result.strandedness = "Forward".to_string(); + result.strandedness = Some("Forward".to_string()); } else if result.reverse_pct > STRANDED_THRESHOLD { result.succeeded = true; - result.strandedness = "Reverse".to_string(); + result.strandedness = Some("Reverse".to_string()); } else if result.forward_pct > UNSTRANDED_THRESHOLD && result.reverse_pct > UNSTRANDED_THRESHOLD { result.succeeded = true; - result.strandedness = "Unstranded".to_string(); - } // else Inconclusive + result.strandedness = Some("Unstranded".to_string()); + } // else did not succeed result } @@ -541,7 +541,7 @@ mod tests { }; let result = predict_strandedness("rg1", &counts); assert!(result.succeeded); - assert_eq!(result.strandedness, "Reverse"); + assert_eq!(result.strandedness, Some("Reverse".to_string())); assert_eq!(result.forward, 10); assert_eq!(result.reverse, 90); assert_eq!(result.forward_pct, 10.0); @@ -553,7 +553,7 @@ mod tests { }; let result = predict_strandedness("rg1", &counts); assert!(result.succeeded); - assert_eq!(result.strandedness, "Unstranded"); + assert_eq!(result.strandedness, Some("Unstranded".to_string())); assert_eq!(result.forward, 50); assert_eq!(result.reverse, 50); assert_eq!(result.forward_pct, 50.0); @@ -565,7 +565,7 @@ mod tests { }; let result = predict_strandedness("rg1", &counts); assert!(result.succeeded); - assert_eq!(result.strandedness, "Forward"); + assert_eq!(result.strandedness, Some("Forward".to_string())); assert_eq!(result.forward, 90); assert_eq!(result.reverse, 10); assert_eq!(result.forward_pct, 90.0); @@ -577,7 +577,7 @@ mod tests { }; let result = predict_strandedness("rg1", &counts); assert!(!result.succeeded); - assert_eq!(result.strandedness, "Inconclusive"); + assert_eq!(result.strandedness, None); assert_eq!(result.forward, 0); assert_eq!(result.reverse, 0); assert_eq!(result.forward_pct, 0.0); diff --git a/src/derive/strandedness/results.rs b/src/derive/strandedness/results.rs index 7bd294b..3082ba8 100644 --- a/src/derive/strandedness/results.rs +++ b/src/derive/strandedness/results.rs @@ -85,8 +85,8 @@ pub struct ReadGroupDerivedStrandednessResult { /// Whether or not strandedness was determined for this read group. pub succeeded: bool, - /// The strandedness of this read group or "Inconclusive". - pub strandedness: String, + /// The strandedness of this read group, if derivable. + pub strandedness: Option, /// The total number of reads in this read group. pub total: usize, @@ -109,7 +109,7 @@ impl ReadGroupDerivedStrandednessResult { pub fn new( read_group: String, succeeded: bool, - strandedness: String, + strandedness: Option, forward: usize, reverse: usize, ) -> Self { @@ -133,8 +133,8 @@ pub struct DerivedStrandednessResult { /// Whether or not the `ngs derive strandedness` subcommand succeeded. pub succeeded: bool, - /// The strandedness of this read group or "Inconclusive". - pub strandedness: String, + /// The strandedness of this read group, if derivable. + pub strandedness: Option, /// The total number of reads. pub total: usize, @@ -173,7 +173,7 @@ impl DerivedStrandednessResult { /// Creates a new [`DerivedStrandednessResult`]. pub fn new( succeeded: bool, - strandedness: String, + strandedness: Option, forward: usize, reverse: usize, read_groups: Vec, From 55b3bc7481a591b2989fd21f3b8d17cb3a173258 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 09:11:39 -0500 Subject: [PATCH 72/91] style: prettify imports --- src/derive/command/encoding.rs | 11 ++++------- src/derive/command/endedness.rs | 14 +++++--------- src/derive/command/instrument.rs | 3 +-- src/derive/command/junction_annotation.rs | 11 ++++------- src/derive/command/readlen.rs | 8 +++----- src/derive/command/strandedness.rs | 20 +++++++------------- src/derive/endedness/compute.rs | 3 +-- src/derive/instrument/compute.rs | 7 ++----- src/derive/instrument/flowcells.rs | 3 +-- src/derive/instrument/instruments.rs | 3 +-- src/derive/junction_annotation/compute.rs | 6 ++---- src/derive/junction_annotation/results.rs | 3 +-- src/derive/strandedness/compute.rs | 6 ++++-- 13 files changed, 36 insertions(+), 62 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index a85d30c..5b3ee52 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -1,15 +1,12 @@ //! Functionality relating to the `ngs derive encoding` subcommand itself. +use anyhow::{Context, Ok}; +use clap::Args; +use noodles::bam; +use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; use std::io::BufReader; use std::path::PathBuf; - -use anyhow::Context; -use anyhow::Ok; -use clap::Args; -use noodles::bam; -use num_format::Locale; -use num_format::ToFormattedString; use tracing::info; use crate::derive::encoding::compute; diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index a35b390..5a457e0 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -1,16 +1,12 @@ //! Functionality relating to the `ngs derive endedness` subcommand itself. -use std::collections::HashMap; -use std::collections::HashSet; -use std::path::PathBuf; -use std::sync::Arc; - use anyhow::Context; use clap::Args; -use num_format::Locale; -use num_format::ToFormattedString; -use tracing::info; -use tracing::trace; +use num_format::{Locale, ToFormattedString}; +use std::collections::{HashMap, HashSet}; +use std::path::PathBuf; +use std::sync::Arc; +use tracing::{info, trace}; use crate::derive::endedness::compute; use crate::derive::endedness::compute::OrderingFlagsCounts; diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 26c102f..93690ba 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -2,8 +2,7 @@ use anyhow::bail; use clap::Args; -use num_format::Locale; -use num_format::ToFormattedString; +use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; use std::path::PathBuf; use tracing::info; diff --git a/src/derive/command/junction_annotation.rs b/src/derive/command/junction_annotation.rs index c265867..def6281 100644 --- a/src/derive/command/junction_annotation.rs +++ b/src/derive/command/junction_annotation.rs @@ -1,15 +1,12 @@ //! Functionality relating to the `ngs derive junction-annotation` subcommand itself. -use std::collections::HashMap; -use std::path::PathBuf; - use anyhow::Context; use clap::Args; use noodles::sam::record::MappingQuality; -use num_format::Locale; -use num_format::ToFormattedString; -use tracing::debug; -use tracing::info; +use num_format::{Locale, ToFormattedString}; +use std::collections::HashMap; +use std::path::PathBuf; +use tracing::{debug, info}; use crate::derive::junction_annotation::compute; use crate::derive::junction_annotation::results::JunctionAnnotationResults; diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index e6b0c51..f5c2246 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -1,12 +1,10 @@ //! Functionality relating to the `ngs derive readlen` subcommand itself. -use std::collections::HashMap; -use std::path::PathBuf; - use anyhow::Context; use clap::Args; -use num_format::Locale; -use num_format::ToFormattedString; +use num_format::{Locale, ToFormattedString}; +use std::collections::HashMap; +use std::path::PathBuf; use tracing::info; use crate::derive::readlen::compute; diff --git a/src/derive/command/strandedness.rs b/src/derive/command/strandedness.rs index bc10479..5ce71d7 100644 --- a/src/derive/command/strandedness.rs +++ b/src/derive/command/strandedness.rs @@ -1,23 +1,17 @@ //! Functionality relating to the `ngs derive strandedness` subcommand itself. -use std::collections::HashMap; -use std::collections::HashSet; -use std::fs::File; -use std::path::PathBuf; - -use anyhow::bail; -use anyhow::Context; +use anyhow::{bail, Context}; use clap::Args; -use noodles::bam; -use noodles::gff; use noodles::sam::record::MappingQuality; +use noodles::{bam, gff}; use rust_lapper::{Interval, Lapper}; -use tracing::debug; -use tracing::info; +use std::collections::{HashMap, HashSet}; +use std::fs::File; +use std::path::PathBuf; +use tracing::{debug, info}; -use crate::derive::strandedness::compute; use crate::derive::strandedness::compute::ParsedBAMFile; -use crate::derive::strandedness::results; +use crate::derive::strandedness::{compute, results}; use crate::utils::formats; use crate::utils::read_groups::validate_read_group_info; diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index d70633c..388b6a2 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -1,7 +1,6 @@ //! Module holding the logic for computing the endedness of a BAM. -use std::collections::HashMap; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; use std::ops::{Add, AddAssign}; use std::sync::Arc; use tracing::warn; diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 59ce1ed..8af7b9e 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -1,14 +1,11 @@ //! Combines the flowcell and instrument checks into a single workflow. -use std::collections::HashMap; -use std::collections::HashSet; - use regex::Regex; use serde::Serialize; +use std::collections::{HashMap, HashSet}; use tracing::debug; -use super::flowcells; -use super::instruments; +use crate::derive::instrument::{flowcells, instruments}; /// Generalized struct for holding instrument detection results. #[derive(Debug, Default, Serialize)] diff --git a/src/derive/instrument/flowcells.rs b/src/derive/instrument/flowcells.rs index 4bb2389..b339cfa 100644 --- a/src/derive/instrument/flowcells.rs +++ b/src/derive/instrument/flowcells.rs @@ -1,7 +1,6 @@ //! Knowledge about which flowcells map to which machine types. -use std::collections::HashMap; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; /// Encapsulates the knowledge we currently have on which flowcell patterns map /// to which machine types as a [`HashMap`]. diff --git a/src/derive/instrument/instruments.rs b/src/derive/instrument/instruments.rs index e10b114..a1df971 100644 --- a/src/derive/instrument/instruments.rs +++ b/src/derive/instrument/instruments.rs @@ -1,7 +1,6 @@ //! Knowledge about which instrument ids map to which machine types. -use std::collections::HashMap; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; /// Encapsulates the knowledge we currently have on which instrument name patterns map /// to which machine types as a [`HashMap`]. diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 428daac..3c6b5ee 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -1,14 +1,12 @@ //! Module holding the logic for annotating junctions. -use anyhow::bail; -use anyhow::Ok; +use anyhow::{bail, Ok}; use noodles::core::Position; use noodles::sam::alignment::Record; use noodles::sam::record::cigar::op::Kind; use noodles::sam::record::MappingQuality; use noodles::sam::Header; -use std::collections::HashMap; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; use crate::derive::junction_annotation::results; use crate::utils::alignment::filter_by_mapq; diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index edd1dd3..9e363cc 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -2,8 +2,7 @@ use noodles::core::Position; use serde::ser::SerializeStruct; -use serde::Serialize; -use serde::Serializer; +use serde::{Serialize, Serializer}; use std::collections::HashMap; /// A junction is a tuple of (start, end) coordinates. diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 6fffda5..7133adc 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -1,7 +1,8 @@ //! Module holding the logic for computing the strandedness. +use noodles::core::Region; use noodles::sam::record::MappingQuality; -use noodles::{bam, core::Region, gff, sam}; +use noodles::{bam, gff, sam}; use rand::Rng; use rust_lapper::Lapper; use std::collections::{HashMap, HashSet}; @@ -9,8 +10,9 @@ use std::ops::{Add, AddAssign}; use std::sync::Arc; use crate::derive::strandedness::results; +use crate::utils::alignment::filter_by_mapq; +use crate::utils::display::RecordCounter; use crate::utils::read_groups; -use crate::utils::{alignment::filter_by_mapq, display::RecordCounter}; const STRANDED_THRESHOLD: f64 = 80.0; const UNSTRANDED_THRESHOLD: f64 = 40.0; From ded9a8906812f17af011dcfdb4769d3795de1997 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 09:29:31 -0500 Subject: [PATCH 73/91] style: wrap optional variable in Option --- src/derive/command/endedness.rs | 7 ++++--- src/derive/endedness/compute.rs | 16 ++++++++-------- 2 files changed, 12 insertions(+), 11 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 5a457e0..8cf78a5 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -58,7 +58,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let mut ordering_flags: HashMap = HashMap::new(); // only used if args.calc_rpt is true - let mut read_names: HashMap> = HashMap::new(); + let mut read_names: Option>> = None; let ParsedBAMFile { mut reader, header, .. @@ -82,17 +82,18 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let read_group = get_read_group(&record, Some(&mut found_rgs)); if args.calc_rpt { + let read_name_map = read_names.get_or_insert_with(HashMap::new); match record.read_name() { Some(rn) => { let rn = rn.to_string(); - let rg_vec = read_names.get_mut(&rn); + let rg_vec = read_name_map.get_mut(&rn); match rg_vec { Some(rg_vec) => { rg_vec.push(Arc::clone(&read_group)); } None => { - read_names.insert(rn, vec![(Arc::clone(&read_group))]); + read_name_map.insert(rn, vec![(Arc::clone(&read_group))]); } } } diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index 388b6a2..afb8ae6 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -259,13 +259,13 @@ fn predict_endedness( /// resulting [`DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( ordering_flags: HashMap, - read_names: HashMap>, + read_names: Option>>, paired_deviance: f64, round_rpt: bool, ) -> results::DerivedEndednessResult { let mut rg_rpts: HashMap = HashMap::new(); let mut overall_rpt: Option = None; - if !read_names.is_empty() { + if let Some(read_names) = read_names { overall_rpt = Some(calculate_reads_per_template(read_names, &mut rg_rpts)); } @@ -401,7 +401,7 @@ mod tests { neither: 0, }, ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); + let result = predict(ordering_flags, None, 0.0, false); assert!(!result.succeeded); assert_eq!(result.endedness, None); assert_eq!(result.first, 1); @@ -425,7 +425,7 @@ mod tests { neither: 0, }, ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); + let result = predict(ordering_flags, None, 0.0, false); assert!(!result.succeeded); assert_eq!(result.endedness, None); assert_eq!(result.first, 0); @@ -449,7 +449,7 @@ mod tests { neither: 0, }, ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); + let result = predict(ordering_flags, None, 0.0, false); assert!(!result.succeeded); assert_eq!(result.endedness, None); assert_eq!(result.first, 0); @@ -473,7 +473,7 @@ mod tests { neither: 1, }, ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); + let result = predict(ordering_flags, None, 0.0, false); assert!(!result.succeeded); assert_eq!(result.endedness, None); assert_eq!(result.first, 0); @@ -497,7 +497,7 @@ mod tests { neither: 0, }, ); - let result = predict(ordering_flags, HashMap::new(), 0.0, false); + let result = predict(ordering_flags, None, 0.0, false); assert!(result.succeeded); assert_eq!(result.endedness, Some("Paired-End".to_string())); assert_eq!(result.first, 1); @@ -597,7 +597,7 @@ mod tests { Arc::clone(&rg_single), ], ); - let result = predict(ordering_flags, read_names, 0.0, false); + let result = predict(ordering_flags, Some(read_names), 0.0, false); assert!(!result.succeeded); assert_eq!(result.endedness, None); assert_eq!(result.unsegmented, 2); From 11dc18dddd16e85caa9e0aa228876cce78b9d230 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 09:44:00 -0500 Subject: [PATCH 74/91] docs: fix intra links --- src/derive/endedness/compute.rs | 2 +- src/derive/strandedness/compute.rs | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/src/derive/endedness/compute.rs b/src/derive/endedness/compute.rs index afb8ae6..9482ac1 100644 --- a/src/derive/endedness/compute.rs +++ b/src/derive/endedness/compute.rs @@ -256,7 +256,7 @@ fn predict_endedness( /// Main method to evaluate the collected ordering flags and /// return a result for the endedness of the file. This may fail, and the -/// resulting [`DerivedEndednessResult`] should be evaluated accordingly. +/// resulting [`results::DerivedEndednessResult`] should be evaluated accordingly. pub fn predict( ordering_flags: HashMap, read_names: Option>>, diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 7133adc..c63035b 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -324,7 +324,7 @@ pub fn predict_strandedness( /// Main method to evaluate the observed strand state and /// return a result for the derived strandedness. This may fail, and the -/// resulting [`DerivedStrandednessResult`] should be evaluated accordingly. +/// resulting [`results::DerivedStrandednessResult`] should be evaluated accordingly. pub fn predict( parsed_bam: &mut ParsedBAMFile, gene_records: &mut Vec, From 4225ec05b5a5b64fcce8ec5359536e9743f714f2 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 10:24:51 -0500 Subject: [PATCH 75/91] tests: fix broken gene test --- src/derive/strandedness/compute.rs | 17 +++++++++++------ 1 file changed, 11 insertions(+), 6 deletions(-) diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index c63035b..9be8f3c 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -166,7 +166,7 @@ fn disqualify_gene(gene: &gff::Record, exons: &HashMap<&str, Lapper intervals .find(gene.start().into(), gene.end().into()) - .any(|exon| { + .all(|exon| { if exon.val != gene_strand { all_on_same_strand = false; } @@ -422,6 +422,7 @@ mod tests { #[test] fn test_disqualify_gene() { + // test mixed strands let mut exons = HashMap::new(); exons.insert( "chr1", @@ -439,9 +440,11 @@ mod tests { ]), ); - let gene = gff::Record::default(); - assert!(disqualify_gene(&gene, &exons)); + let s = "chr1\tNOODLES\tgene\t5\t14\t.\t+\t.\tgene_id=ndls0;gene_name=gene0"; + let record = s.parse::().unwrap(); + assert!(disqualify_gene(&record, &exons)); // disqualified + // test all on same strand let mut exons = HashMap::new(); exons.insert( "chr1", @@ -459,9 +462,11 @@ mod tests { ]), ); - let s = "chr1\tNOODLES\tgene\t8\t13\t.\t+\t.\tgene_id=ndls0;gene_name=gene0"; - let record = s.parse::().unwrap(); - assert!(!disqualify_gene(&record, &exons)); + assert!(!disqualify_gene(&record, &exons)); // accepted + + // test no exons + let exons = HashMap::new(); + assert!(disqualify_gene(&record, &exons)); // disqualified } #[test] From 780d3bc090161749850df1983c7c9b5261bed716 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 12 Feb 2024 16:15:36 -0500 Subject: [PATCH 76/91] feat(derive/instrument): more debug statements --- src/derive/instrument/compute.rs | 9 +++++++++ 1 file changed, 9 insertions(+) diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 8af7b9e..49b46ba 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -297,7 +297,16 @@ pub fn predict( let instruments = instruments::build_instrument_lookup_table(); let flowcells = flowcells::build_flowcell_lookup_table(); + debug!( + "Predicting instruments from instrument names: {:?}", + instrument_names + ); let iid_results = predict_instrument(instrument_names, &instruments); + + debug!( + "Predicting instruments from flowcell names: {:?}", + flowcell_names + ); let fcid_results = predict_instrument(flowcell_names, &flowcells); resolve_instrument_prediction(iid_results, fcid_results) From 09a2be2e841d47344a47b7cf3e0a68ff23362fd1 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 13 Feb 2024 10:30:36 -0500 Subject: [PATCH 77/91] feat(derive/instrument): in output, report found unique names --- src/derive/command/instrument.rs | 4 ++-- src/derive/instrument/compute.rs | 18 ++++++------------ 2 files changed, 8 insertions(+), 14 deletions(-) diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 93690ba..48de551 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -81,8 +81,8 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { counter.get().to_formatted_string(&Locale::en) ); metrics.total_records = counter.get(); - metrics.unique_instrument_names = instrument_names.len(); - metrics.unique_flowcell_names = flowcell_names.len(); + metrics.unique_instrument_names = instrument_names.clone(); + metrics.unique_flowcell_names = flowcell_names.clone(); // (2) Derive the predict instrument results based on these detected // instrument names and flowcell names. diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 49b46ba..e77a1be 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -57,11 +57,11 @@ pub struct RecordMetrics { /// flowcell name in their read name. pub found_flowcell_name: usize, - /// The number of unique instrument names that were detected. - pub unique_instrument_names: usize, + /// The unique instrument names that were detected. + pub unique_instrument_names: HashSet, - /// The number of unique flowcell names that were detected. - pub unique_flowcell_names: usize, + /// The unique flowcell names that were detected. + pub unique_flowcell_names: HashSet, } /// Struct holding the final results for an `ngs derive instrument` subcommand @@ -297,16 +297,10 @@ pub fn predict( let instruments = instruments::build_instrument_lookup_table(); let flowcells = flowcells::build_flowcell_lookup_table(); - debug!( - "Predicting instruments from instrument names: {:?}", - instrument_names - ); + debug!("Predicting instruments from instrument names"); let iid_results = predict_instrument(instrument_names, &instruments); - debug!( - "Predicting instruments from flowcell names: {:?}", - flowcell_names - ); + debug!("Predicting instruments from flowcell names"); let fcid_results = predict_instrument(flowcell_names, &flowcells); resolve_instrument_prediction(iid_results, fcid_results) From 05a53fc7af372a8f4faefd30e79d58af0d518466 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 13 Feb 2024 11:35:48 -0500 Subject: [PATCH 78/91] feat: more info in JSON report. Not complete yet. Open TODOs --- src/derive/instrument/compute.rs | 66 +++++++++++++++++++++++--------- 1 file changed, 48 insertions(+), 18 deletions(-) diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index e77a1be..83b69bb 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -39,6 +39,16 @@ impl InstrumentDetectionResults { } } +/// TODO +#[derive(Debug, Serialize)] +pub struct QueryResult { + /// The query that was used to generate the result. + pub query: String, + + /// The possible instruments that could have generated the query. + pub result: HashSet, +} + /// Metrics related to how read records were processed. #[derive(Debug, Default, Serialize)] pub struct RecordMetrics { @@ -85,6 +95,12 @@ pub struct DerivedInstrumentResult { /// A general comment field, if available. pub comment: Option, + /// TODO + pub instrument_name_queries: Vec, + + /// TODO + pub flowcell_name_queries: Vec, + /// Metrics related to how read records were processed. pub records: RecordMetrics, } @@ -97,6 +113,8 @@ impl DerivedInstrumentResult { confidence: String, evidence: Option, comment: Option, + instrument_name_queries: Vec, + flowcell_name_queries: Vec, records: RecordMetrics, ) -> Self { DerivedInstrumentResult { @@ -105,6 +123,8 @@ impl DerivedInstrumentResult { confidence, evidence, comment, + instrument_name_queries, + flowcell_name_queries, records, } } @@ -118,6 +138,8 @@ impl Default for DerivedInstrumentResult { confidence: "unknown".to_string(), evidence: None, comment: None, + instrument_name_queries: Vec::new(), + flowcell_name_queries: Vec::new(), records: RecordMetrics::default(), } } @@ -143,25 +165,28 @@ impl Default for DerivedInstrumentResult { pub fn possible_instruments_for_query( query: String, lookup_table: &HashMap<&'static str, HashSet<&'static str>>, -) -> HashSet { - let mut result: HashSet = HashSet::new(); +) -> QueryResult { + let mut result_set: HashSet = HashSet::new(); for (pattern, machines) in lookup_table { let re = Regex::new(pattern).unwrap(); if re.is_match(query.as_str()) { let matching_machines: Vec = machines.iter().map(|x| x.to_string()).collect(); - result.extend(matching_machines); + result_set.extend(matching_machines); } } - debug!(" [*] {}, Possible Instruments: {:?}", query, result); - result + debug!(" [*] {}, Possible Instruments: {:?}", query, result_set); + QueryResult { + query, + result: result_set, + } } /// Given a HashSet of unique queries (usually a instrument ID or flowcell ID /// parsed from a read name) that were detected from a SAM/BAM/CRAM file, return /// a HashSet that contains all possible machines that could have generated that -/// list of queries. +/// list of queries and a vec recording the query look-ups that were made. /// /// This is done by iterating through the HashSet of machines that could have /// produced each name and taking the intersection. It is possible, of course, @@ -181,15 +206,17 @@ pub fn possible_instruments_for_query( pub fn predict_instrument( queries: HashSet, lookup_table: &HashMap<&'static str, HashSet<&'static str>>, -) -> InstrumentDetectionResults { +) -> (InstrumentDetectionResults, Vec) { let mut result = InstrumentDetectionResults::default(); + let mut query_results = Vec::new(); for name in queries { let derived = possible_instruments_for_query(name, lookup_table); - result.update_instruments(&derived); + result.update_instruments(&derived.result); + query_results.push(derived); } - result + (result, query_results) } /// Combines evidence from the instrument id detection and flowcell id detection @@ -298,12 +325,15 @@ pub fn predict( let flowcells = flowcells::build_flowcell_lookup_table(); debug!("Predicting instruments from instrument names"); - let iid_results = predict_instrument(instrument_names, &instruments); + let (iid_results, instrument_name_queries) = predict_instrument(instrument_names, &instruments); debug!("Predicting instruments from flowcell names"); - let fcid_results = predict_instrument(flowcell_names, &flowcells); + let (fcid_results, flowcell_name_queries) = predict_instrument(flowcell_names, &flowcells); - resolve_instrument_prediction(iid_results, fcid_results) + let mut final_results = resolve_instrument_prediction(iid_results, fcid_results); + final_results.instrument_name_queries = instrument_name_queries; + final_results.flowcell_name_queries = flowcell_name_queries; + final_results } #[cfg(test)] @@ -314,30 +344,30 @@ mod tests { fn test_derive_instrument_from_invalid_instrument_name() { let instruments = instruments::build_instrument_lookup_table(); let result = possible_instruments_for_query(String::from("NoMatchingName"), &instruments); - assert!(result.is_empty()); + assert!(result.result.is_empty()); } #[test] fn test_derive_instrument_from_valid_instrument_name() { let instruments = instruments::build_instrument_lookup_table(); let result = possible_instruments_for_query(String::from("A00000"), &instruments); - assert_eq!(result.len(), 1); - assert!(result.contains("NovaSeq")); + assert_eq!(result.result.len(), 1); + assert!(result.result.contains("NovaSeq")); } #[test] fn test_derive_instrument_from_invalid_flowcell_name() { let flowcells = flowcells::build_flowcell_lookup_table(); let result = possible_instruments_for_query(String::from("NoMatchingName"), &flowcells); - assert!(result.is_empty()); + assert!(result.result.is_empty()); } #[test] fn test_derive_instrument_from_valid_flowcell_name() { let flowcells = flowcells::build_flowcell_lookup_table(); let result = possible_instruments_for_query(String::from("H00000RXX"), &flowcells); - assert_eq!(result.len(), 1); - assert!(result.contains("NovaSeq")); + assert_eq!(result.result.len(), 1); + assert!(result.result.contains("NovaSeq")); } #[test] From eae95c751cdbdd102196a3961987df0c78638030 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 13 Feb 2024 12:35:05 -0500 Subject: [PATCH 79/91] style(derive/instrument): a bit of code clean up --- src/derive/command/instrument.rs | 2 -- src/derive/instrument/compute.rs | 23 ++++++++++++----------- 2 files changed, 12 insertions(+), 13 deletions(-) diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 48de551..a0776f8 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -81,8 +81,6 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { counter.get().to_formatted_string(&Locale::en) ); metrics.total_records = counter.get(); - metrics.unique_instrument_names = instrument_names.clone(); - metrics.unique_flowcell_names = flowcell_names.clone(); // (2) Derive the predict instrument results based on these detected // instrument names and flowcell names. diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 83b69bb..c875aba 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -49,6 +49,15 @@ pub struct QueryResult { pub result: HashSet, } +/// TODO +pub struct Queries { + /// TODO + pub instrument_name_queries: Vec, + + /// TODO + pub flowcell_name_queries: Vec, +} + /// Metrics related to how read records were processed. #[derive(Debug, Default, Serialize)] pub struct RecordMetrics { @@ -66,12 +75,6 @@ pub struct RecordMetrics { /// The total number of records that contained a valid /// flowcell name in their read name. pub found_flowcell_name: usize, - - /// The unique instrument names that were detected. - pub unique_instrument_names: HashSet, - - /// The unique flowcell names that were detected. - pub unique_flowcell_names: HashSet, } /// Struct holding the final results for an `ngs derive instrument` subcommand @@ -113,8 +116,7 @@ impl DerivedInstrumentResult { confidence: String, evidence: Option, comment: Option, - instrument_name_queries: Vec, - flowcell_name_queries: Vec, + queries: Queries, records: RecordMetrics, ) -> Self { DerivedInstrumentResult { @@ -123,8 +125,8 @@ impl DerivedInstrumentResult { confidence, evidence, comment, - instrument_name_queries, - flowcell_name_queries, + instrument_name_queries: queries.instrument_name_queries, + flowcell_name_queries: queries.flowcell_name_queries, records, } } @@ -176,7 +178,6 @@ pub fn possible_instruments_for_query( } } - debug!(" [*] {}, Possible Instruments: {:?}", query, result_set); QueryResult { query, result: result_set, From 00999c128c67e40565ef04962c5a774530f6f3fd Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Tue, 13 Feb 2024 13:20:45 -0500 Subject: [PATCH 80/91] docs: filling in TODOs --- src/derive/instrument/compute.rs | 20 ++++++++------------ 1 file changed, 8 insertions(+), 12 deletions(-) diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index c875aba..18a029a 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -3,7 +3,6 @@ use regex::Regex; use serde::Serialize; use std::collections::{HashMap, HashSet}; -use tracing::debug; use crate::derive::instrument::{flowcells, instruments}; @@ -39,7 +38,7 @@ impl InstrumentDetectionResults { } } -/// TODO +/// A query for a look-up table and the resulting hits from that table. #[derive(Debug, Serialize)] pub struct QueryResult { /// The query that was used to generate the result. @@ -49,12 +48,13 @@ pub struct QueryResult { pub result: HashSet, } -/// TODO +/// Utility struct for holding the results of look-ups for instrument and +/// flowcell names. pub struct Queries { - /// TODO + /// The results of the instrument name look-ups. pub instrument_name_queries: Vec, - /// TODO + /// The results of the flowcell name look-ups. pub flowcell_name_queries: Vec, } @@ -98,10 +98,10 @@ pub struct DerivedInstrumentResult { /// A general comment field, if available. pub comment: Option, - /// TODO + /// The results of the instrument name look-ups. pub instrument_name_queries: Vec, - /// TODO + /// The results of the flowcell name look-ups. pub flowcell_name_queries: Vec, /// Metrics related to how read records were processed. @@ -177,7 +177,6 @@ pub fn possible_instruments_for_query( result_set.extend(matching_machines); } } - QueryResult { query, result: result_set, @@ -221,7 +220,7 @@ pub fn predict_instrument( } /// Combines evidence from the instrument id detection and flowcell id detection -/// to produce a final [`DerivedInstrumentResult`]. +/// to produce a [`DerivedInstrumentResult`]. pub fn resolve_instrument_prediction( iid_results: InstrumentDetectionResults, fcid_results: InstrumentDetectionResults, @@ -325,10 +324,7 @@ pub fn predict( let instruments = instruments::build_instrument_lookup_table(); let flowcells = flowcells::build_flowcell_lookup_table(); - debug!("Predicting instruments from instrument names"); let (iid_results, instrument_name_queries) = predict_instrument(instrument_names, &instruments); - - debug!("Predicting instruments from flowcell names"); let (fcid_results, flowcell_name_queries) = predict_instrument(flowcell_names, &flowcells); let mut final_results = resolve_instrument_prediction(iid_results, fcid_results); From 2661300c03f2e3c722700b5c7e1d7dbc0b03e072 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 14 Feb 2024 08:38:48 -0500 Subject: [PATCH 81/91] chore: delete dead code --- src/derive/instrument/compute.rs | 34 -------------------------------- 1 file changed, 34 deletions(-) diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 18a029a..c51070d 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -48,16 +48,6 @@ pub struct QueryResult { pub result: HashSet, } -/// Utility struct for holding the results of look-ups for instrument and -/// flowcell names. -pub struct Queries { - /// The results of the instrument name look-ups. - pub instrument_name_queries: Vec, - - /// The results of the flowcell name look-ups. - pub flowcell_name_queries: Vec, -} - /// Metrics related to how read records were processed. #[derive(Debug, Default, Serialize)] pub struct RecordMetrics { @@ -108,30 +98,6 @@ pub struct DerivedInstrumentResult { pub records: RecordMetrics, } -impl DerivedInstrumentResult { - /// Creates a new [`DerivedInstrumentResult`]. - pub fn new( - succeeded: bool, - instruments: Option>, - confidence: String, - evidence: Option, - comment: Option, - queries: Queries, - records: RecordMetrics, - ) -> Self { - DerivedInstrumentResult { - succeeded, - instruments, - confidence, - evidence, - comment, - instrument_name_queries: queries.instrument_name_queries, - flowcell_name_queries: queries.flowcell_name_queries, - records, - } - } -} - impl Default for DerivedInstrumentResult { fn default() -> Self { DerivedInstrumentResult { From bfac8a111210ca543a194e62aba9947958b2883b Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Wed, 14 Feb 2024 10:49:10 -0500 Subject: [PATCH 82/91] tests(derive/junction-annotation): rewrite tests more modular --- src/derive/junction_annotation/compute.rs | 603 +++++++++++++++++----- src/derive/junction_annotation/results.rs | 6 +- 2 files changed, 467 insertions(+), 142 deletions(-) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 3c6b5ee..60bde5b 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -362,6 +362,41 @@ mod tests { use noodles::sam::record::ReadName; use std::num::NonZeroUsize; + fn create_test_exons() -> ExonSets<'static> { + let exon_starts: HashMap<&str, HashSet> = HashMap::from([( + "sq1", + HashSet::from([ + Position::new(1).unwrap(), + Position::new(11).unwrap(), + Position::new(21).unwrap(), + Position::new(31).unwrap(), + Position::new(41).unwrap(), + Position::new(51).unwrap(), + Position::new(61).unwrap(), + Position::new(71).unwrap(), + ]), + )]); + let exon_ends: HashMap<&str, HashSet> = exon_starts + .iter() + .map(|(k, v)| (*k, v.iter().map(|e| e.checked_add(10).unwrap()).collect())) + .collect::>>(); + let exons: ExonSets<'_> = ExonSets { + starts: exon_starts, + ends: exon_ends, + }; + exons + } + + fn create_test_header() -> Header { + Header::builder() + .set_header(Map::::new(Version::new(1, 6))) + .add_reference_sequence( + "sq1".parse().unwrap(), + Map::::new(NonZeroUsize::try_from(800).unwrap()), + ) + .build() + } + #[test] fn test_filter_by_flags() { // Setup @@ -460,7 +495,7 @@ mod tests { } #[test] - fn test_process_and_summarize() { + fn test_process_known_junction() { // Setup let mut results = results::JunctionAnnotationResults::default(); let params = JunctionAnnotationParameters { @@ -471,38 +506,8 @@ mod tests { count_secondary: false, count_duplicates: false, }; - let header = Header::builder() - .set_header(Map::::new(Version::new(1, 6))) - .add_reference_sequence( - "sq1".parse().unwrap(), - Map::::new(NonZeroUsize::try_from(800).unwrap()), - ) - .add_reference_sequence( - "sq1_random".parse().unwrap(), // unannotated - Map::::new(NonZeroUsize::try_from(400).unwrap()), - ) - .build(); - let exon_starts: HashMap<&str, HashSet> = HashMap::from([( - "sq1", - HashSet::from([ - Position::new(1).unwrap(), - Position::new(11).unwrap(), - Position::new(21).unwrap(), - Position::new(31).unwrap(), - Position::new(41).unwrap(), - Position::new(51).unwrap(), - Position::new(61).unwrap(), - Position::new(71).unwrap(), - ]), - )]); - let exon_ends = exon_starts - .iter() - .map(|(k, v)| (*k, v.iter().map(|e| e.checked_add(10).unwrap()).collect())) - .collect::>>(); - let exons = ExonSets { - starts: exon_starts, - ends: exon_ends, - }; + let exons = create_test_exons(); + let header = create_test_header(); // Test known junction let mut record = Record::default(); @@ -518,56 +523,175 @@ mod tests { assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + assert_eq!(results.junction_annotations.known.len(), 1); + assert_eq!( + results.junction_annotations.known.get("sq1").unwrap().len(), + 1 + ); + assert_eq!( + results + .junction_annotations + .known + .get("sq1") + .unwrap() + .get(&(Position::new(11).unwrap(), Position::new(21).unwrap())) + .unwrap(), + &1 + ); + } - // Test that unmapped gets ignored + #[test] + fn test_process_partial_novel_junction() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); + + // Test partial novel junction let mut record = Record::default(); - let r2_name: ReadName = "unmapped".parse().unwrap(); - *record.read_name_mut() = Some(r2_name); + let r1_name: ReadName = "partial1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(255); - record.flags_mut().set(0x4.into(), true); + *record.cigar_mut() = "10M12N10M".parse().unwrap(); + *record.mapping_quality_mut() = MappingQuality::new(60); + record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); assert_eq!(results.records.processed, 1); - assert_eq!(results.records.filtered_by_flags, 1); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + assert_eq!(results.junction_annotations.partial_novel.len(), 1); + assert_eq!( + results + .junction_annotations + .partial_novel + .get("sq1") + .unwrap() + .len(), + 1 + ); + assert_eq!( + results + .junction_annotations + .partial_novel + .get("sq1") + .unwrap() + .get(&(Position::new(11).unwrap(), Position::new(23).unwrap())) + .unwrap(), + &1 + ); + } - // Test partial novel junction + #[test] + fn test_process_complete_novel_junction() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); + + // Test complete novel junction let mut record = Record::default(); - let r3_name: ReadName = "partial1".parse().unwrap(); - *record.read_name_mut() = Some(r3_name); + let r1_name: ReadName = "complete1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M12N10M".parse().unwrap(); + *record.cigar_mut() = "85M14N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 2); - assert_eq!(results.records.filtered_by_flags, 1); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + assert_eq!(results.junction_annotations.complete_novel.len(), 1); + assert_eq!( + results + .junction_annotations + .complete_novel + .get("sq1") + .unwrap() + .len(), + 1 + ); + assert_eq!( + results + .junction_annotations + .complete_novel + .get("sq1") + .unwrap() + .get(&(Position::new(86).unwrap(), Position::new(100).unwrap())) + .unwrap(), + &1 + ); + } + + #[test] + fn test_process_ignores_unmapped() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); - // Test partial novel junction (again for more read support) + // Test that unmapped gets ignored let mut record = Record::default(); - let r3_name: ReadName = "partial2".parse().unwrap(); - *record.read_name_mut() = Some(r3_name); + let r1_name: ReadName = "unmapped".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M12N10M".parse().unwrap(); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); - record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x4.into(), true); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 3); + assert_eq!(results.records.processed, 0); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + } - // Test that supplementary alignments get counted + #[test] + fn test_process_supplementary_toggle() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let mut params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: true, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); + + // Test that supplementary gets ignored let mut record = Record::default(); - let r4_name: ReadName = "supplementary".parse().unwrap(); - *record.read_name_mut() = Some(r4_name); + let r1_name: ReadName = "supplementary1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); @@ -575,174 +699,375 @@ mod tests { record.flags_mut().set(0x4.into(), false); record.flags_mut().set(0x800.into(), true); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 4); + assert_eq!(results.records.processed, 0); assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); - // Test that secondary alignments don't get counted + // Test that supplementary gets processed + params.no_supplementary = false; + let mut record = Record::default(); - let r5_name: ReadName = "secondary".parse().unwrap(); - *record.read_name_mut() = Some(r5_name); + let r2_name = "supplementary2".parse().unwrap(); + *record.read_name_mut() = Some(r2_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); - record.flags_mut().set(0x100.into(), true); + record.flags_mut().set(0x800.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 4); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + } + + #[test] + fn test_process_secondary_toggle() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let mut params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: true, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); - // Test complete novel junction with 2 junctions + // Test that secondary gets processed let mut record = Record::default(); - let r6_name: ReadName = "complete_twice1".parse().unwrap(); - *record.read_name_mut() = Some(r6_name); + let r1_name: ReadName = "secondary1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(200); - *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x100.into(), true); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 5); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); - // Test complete novel junction with 2 junctions (again for more read support) + // Test that secondary gets ignored + params.count_secondary = false; + let mut record = Record::default(); - let r6_name: ReadName = "complete_twice2".parse().unwrap(); - *record.read_name_mut() = Some(r6_name); + let r2_name = "secondary2".parse().unwrap(); + *record.read_name_mut() = Some(r2_name); *record.reference_sequence_id_mut() = Some(0); - *record.alignment_start_mut() = Position::new(200); - *record.cigar_mut() = "10M10N10M10N10M".parse().unwrap(); + *record.alignment_start_mut() = Position::new(1); + *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); + record.flags_mut().set(0x100.into(), true); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 1); assert_eq!(results.records.not_spliced, 0); assert_eq!(results.records.bad_mapq, 0); + } + + #[test] + fn test_process_mapq_toggle() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let mut params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); - // Test fails MAPQ filter + // Test that mapq gets processed let mut record = Record::default(); - let r7_name: ReadName = "low_mapq".parse().unwrap(); - *record.read_name_mut() = Some(r7_name); + let r1_name: ReadName = "mapq1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = MappingQuality::new(20); + *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.bad_mapq, 1); + assert_eq!(results.records.bad_mapq, 0); + + // Test that mapq gets ignored + params.min_mapq = Some(MappingQuality::new(61).unwrap()); - // Test missing MAPQ let mut record = Record::default(); - let r8_name: ReadName = "bad_mapq".parse().unwrap(); - *record.read_name_mut() = Some(r8_name); + let r2_name = "mapq2".parse().unwrap(); + *record.read_name_mut() = Some(r2_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); - *record.mapping_quality_mut() = None; + *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 6); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.bad_mapq, 2); + assert_eq!(results.records.bad_mapq, 1); + } - // Test that intron is too short + #[test] + fn test_process_intron_too_short() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); + + // Test that intron too short gets processed let mut record = Record::default(); - let r9_name: ReadName = "short".parse().unwrap(); - *record.read_name_mut() = Some(r9_name); + let r1_name: ReadName = "short1".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "5M5N5M".parse().unwrap(); + *record.cigar_mut() = "10M5N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 7); // Still gets processed, will be filtered later - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 1); // processed at first, gets filtered later + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 0); - assert_eq!(results.records.bad_mapq, 2); + assert_eq!(results.records.bad_mapq, 0); + } - // Test that that reads not spliced are ignored + #[test] + fn test_process_multiple_junctions() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); + + // Test that multiple junctions are processed let mut record = Record::default(); - let r10_name: ReadName = "not_spliced".parse().unwrap(); - *record.read_name_mut() = Some(r10_name); + let r1_name: ReadName = "long_read".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M".parse().unwrap(); + *record.cigar_mut() = "10M10N10M10N10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 7); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.bad_mapq, 2); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 0); + assert_eq!(results.junction_annotations.known.len(), 1); + assert_eq!( + results.junction_annotations.known.get("sq1").unwrap().len(), + 3 + ); + assert_eq!( + results + .junction_annotations + .known + .get("sq1") + .unwrap() + .get(&(Position::new(11).unwrap(), Position::new(21).unwrap())) + .unwrap(), + &1 + ); + assert_eq!( + results + .junction_annotations + .known + .get("sq1") + .unwrap() + .get(&(Position::new(31).unwrap(), Position::new(41).unwrap())) + .unwrap(), + &1 + ); + assert_eq!( + results + .junction_annotations + .known + .get("sq1") + .unwrap() + .get(&(Position::new(51).unwrap(), Position::new(61).unwrap())) + .unwrap(), + &1 + ); + } + + #[test] + fn test_process_unspliced_read() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let exons = create_test_exons(); + let header = create_test_header(); - // Test unannoted reference + // Test that unspliced gets ignored let mut record = Record::default(); - let r11_name: ReadName = "unannotated1".parse().unwrap(); - *record.read_name_mut() = Some(r11_name); - *record.reference_sequence_id_mut() = Some(1); + let r1_name: ReadName = "unspliced".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); + *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); - *record.cigar_mut() = "10M10N10M".parse().unwrap(); + *record.cigar_mut() = "10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 8); - assert_eq!(results.records.filtered_by_flags, 2); + assert_eq!(results.records.processed, 0); + assert_eq!(results.records.filtered_by_flags, 0); assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.bad_mapq, 2); + assert_eq!(results.records.bad_mapq, 0); + } + + #[test] + fn test_process_unannotated_reference() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + let rand_header = Header::builder() + .set_header(Map::::new(Version::new(1, 6))) + .add_reference_sequence( + "sq1_random".parse().unwrap(), + Map::::new(NonZeroUsize::try_from(800).unwrap()), + ) + .build(); + let exons = create_test_exons(); - // Test unannoted reference (again for more read support) + // Test that unannotated reference gets processed let mut record = Record::default(); - let r11_name: ReadName = "unannotated2".parse().unwrap(); - *record.read_name_mut() = Some(r11_name); - *record.reference_sequence_id_mut() = Some(1); + let r1_name: ReadName = "unannotated".parse().unwrap(); + *record.read_name_mut() = Some(r1_name); + *record.reference_sequence_id_mut() = Some(0); *record.alignment_start_mut() = Position::new(1); *record.cigar_mut() = "10M10N10M".parse().unwrap(); *record.mapping_quality_mut() = MappingQuality::new(60); record.flags_mut().set(0x4.into(), false); - process(&record, &exons, &header, ¶ms, &mut results).unwrap(); - assert_eq!(results.records.processed, 9); - assert_eq!(results.records.filtered_by_flags, 2); - assert_eq!(results.records.not_spliced, 1); - assert_eq!(results.records.bad_mapq, 2); + process(&record, &exons, &rand_header, ¶ms, &mut results).unwrap(); + assert_eq!(results.records.processed, 1); + assert_eq!(results.records.filtered_by_flags, 0); + assert_eq!(results.records.not_spliced, 0); + assert_eq!(results.records.bad_mapq, 0); + assert_eq!(results.junction_annotations.unannotated_reference.len(), 1); + assert_eq!( + results + .junction_annotations + .unannotated_reference + .get("sq1_random") + .unwrap() + .len(), + 1 + ); + assert_eq!( + results + .junction_annotations + .unannotated_reference + .get("sq1_random") + .unwrap() + .get(&(Position::new(11).unwrap(), Position::new(21).unwrap())) + .unwrap(), + &1 + ); + } - // Test summarize - summarize(&mut results, ¶ms); + #[test] + fn test_summarize() { + // Setup + let mut results = results::JunctionAnnotationResults::default(); + let params = JunctionAnnotationParameters { + min_intron_length: 10, + min_read_support: 2, + min_mapq: Some(MappingQuality::new(30).unwrap()), + no_supplementary: false, + count_secondary: false, + count_duplicates: false, + }; + results.junction_annotations.known.insert( + "sq1".to_string(), + HashMap::from([ + ((Position::new(11).unwrap(), Position::new(21).unwrap()), 4), + ((Position::new(31).unwrap(), Position::new(41).unwrap()), 1), + ((Position::new(21).unwrap(), Position::new(41).unwrap()), 3), + ]), + ); + results.junction_annotations.partial_novel.insert( + "sq1".to_string(), + HashMap::from([ + ((Position::new(11).unwrap(), Position::new(37).unwrap()), 3), + ((Position::new(11).unwrap(), Position::new(15).unwrap()), 2), + ]), + ); + results.junction_annotations.complete_novel.insert( + "sq1".to_string(), + HashMap::from([( + (Position::new(103).unwrap(), Position::new(117).unwrap()), + 2, + )]), + ); + results.junction_annotations.unannotated_reference.insert( + "sq1_random".to_string(), + HashMap::from([((Position::new(1).unwrap(), Position::new(11).unwrap()), 5)]), + ); - assert_eq!(results.summary.total_rejected_junctions, 1); - assert_eq!(results.summary.intron_too_short, 1); - assert_eq!(results.summary.junctions_with_not_enough_read_support, 1); - assert_eq!(results.summary.known_junctions, 1); - assert_eq!(results.summary.known_junctions_read_support, 2); + // Test that results are summarized correctly + summarize(&mut results, ¶ms); + assert_eq!(results.summary.known_junctions, 2); + assert_eq!(results.summary.known_junctions_read_support, 7); assert_eq!(results.summary.partial_novel_junctions, 1); - assert_eq!(results.summary.partial_novel_junctions_read_support, 2); - assert_eq!(results.summary.complete_novel_junctions, 2); - assert_eq!(results.summary.complete_novel_junctions_read_support, 4); + assert_eq!(results.summary.partial_novel_junctions_read_support, 3); + assert_eq!(results.summary.complete_novel_junctions, 1); + assert_eq!(results.summary.complete_novel_junctions_read_support, 2); assert_eq!(results.summary.unannotated_reference_junctions, 1); assert_eq!( results.summary.unannotated_reference_junctions_read_support, - 2 + 5 ); assert_eq!(results.summary.total_junctions, 5); - assert_eq!(results.summary.total_junctions_read_support, 10); - assert_eq!(results.summary.known_junctions_percent, 25.0); + assert_eq!(results.summary.total_junctions_read_support, 17); + assert_eq!(results.summary.known_junctions_percent, 50.0); assert_eq!(results.summary.partial_novel_junctions_percent, 25.0); - assert_eq!(results.summary.complete_novel_junctions_percent, 50.0); - assert_eq!(results.summary.average_junction_read_support, 2.0); - assert_eq!(results.summary.average_known_junction_read_support, 2.0); + assert_eq!(results.summary.complete_novel_junctions_percent, 25.0); + assert_eq!(results.summary.average_junction_read_support, 3.4); + assert_eq!(results.summary.average_known_junction_read_support, 3.5); assert_eq!( results.summary.average_partial_novel_junction_read_support, - 2.0 + 3.0 ); assert_eq!( results.summary.average_complete_novel_junction_read_support, diff --git a/src/derive/junction_annotation/results.rs b/src/derive/junction_annotation/results.rs index 9e363cc..0ce8f86 100644 --- a/src/derive/junction_annotation/results.rs +++ b/src/derive/junction_annotation/results.rs @@ -16,7 +16,7 @@ pub type JunctionCounter = HashMap; pub type JunctionsMap = HashMap; /// Lists of annotated junctions. -#[derive(Clone, Default)] +#[derive(Clone, Debug, Default)] pub struct JunctionAnnotations { /// Known junctions. The outer key is the referece name, and the value is another /// HashMap. The inner key is the (start, end) coordinates of a junction, @@ -90,7 +90,7 @@ impl Serialize for JunctionAnnotations { /// General record metrics that are tallied as a part of the /// junction-annotation subcommand. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Debug, Default, Serialize)] pub struct RecordMetrics { /// The number of records that have been fully processed. /// This is the number of spliced records that have been considered. @@ -205,7 +205,7 @@ pub struct SummaryResults { /// Main Results struct. This struct aggregates all of the minor metrics structs /// outlined in this file so they can be kept track of as a unit. -#[derive(Clone, Default, Serialize)] +#[derive(Clone, Default, Debug, Serialize)] pub struct JunctionAnnotationResults { /// Lists of annotated junctions. pub junction_annotations: JunctionAnnotations, From 7ab84f29bf52a82da451670298f2e416947015b2 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Thu, 22 Feb 2024 12:09:31 -0500 Subject: [PATCH 83/91] stlye: Michael M. feedback --- src/derive/junction_annotation/compute.rs | 47 +++++++++++------------ 1 file changed, 22 insertions(+), 25 deletions(-) diff --git a/src/derive/junction_annotation/compute.rs b/src/derive/junction_annotation/compute.rs index 60bde5b..e50ce99 100644 --- a/src/derive/junction_annotation/compute.rs +++ b/src/derive/junction_annotation/compute.rs @@ -188,29 +188,25 @@ pub fn process( intron_end_known = true; } - // TODO: Better way to do this? - increment_junction_map( - match (intron_start_known, intron_end_known) { - (true, true) => { - // We found both ends of the intron. - // This is a Known Junction. - &mut results.junction_annotations.known - } - (true, false) | (false, true) => { - // We found one end of the intron, - // but not the other. - // This is a Partial Novel Junction. - &mut results.junction_annotations.partial_novel - } - (false, false) => { - // We found neither end of the intron. - // This is a Complete Novel Junction. - &mut results.junction_annotations.complete_novel - } - }, - seq_name, - junction, - ) + let junction_map = match (intron_start_known, intron_end_known) { + (true, true) => { + // We found both ends of the intron. + // This is a Known Junction. + &mut results.junction_annotations.known + } + (true, false) | (false, true) => { + // We found one end of the intron, + // but not the other. + // This is a Partial Novel Junction. + &mut results.junction_annotations.partial_novel + } + (false, false) => { + // We found neither end of the intron. + // This is a Complete Novel Junction. + &mut results.junction_annotations.complete_novel + } + }; + increment_junction_map(junction_map, seq_name, junction) } // Operations that increment the reference position (beside Skip which is handled above). Kind::Match | Kind::Deletion | Kind::SequenceMatch | Kind::SequenceMismatch => { @@ -445,6 +441,7 @@ mod tests { junction_map.insert( "sq1".to_string(), HashMap::from([ + ((Position::new(1).unwrap(), Position::new(10).unwrap()), 3), ((Position::new(1).unwrap(), Position::new(11).unwrap()), 1), ((Position::new(1).unwrap(), Position::new(5).unwrap()), 1), ]), @@ -467,9 +464,9 @@ mod tests { assert_eq!(junction_map.len(), 1); assert_eq!(junction_map.get("sq1"), None); assert_eq!(junction_map.get("sq2").unwrap().len(), 1); - assert_eq!(metrics.intron_too_short, 1); + assert_eq!(metrics.intron_too_short, 2); assert_eq!(metrics.junctions_with_not_enough_read_support, 2); - assert_eq!(metrics.total_rejected_junctions, 2); + assert_eq!(metrics.total_rejected_junctions, 3); } #[test] From 68f999c8af0c8c715a448c317b815f954c3c2761 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 22 Mar 2024 09:25:59 -0400 Subject: [PATCH 84/91] Apply suggestions from code review Co-authored-by: Clay McLeod <3411613+claymcleod@users.noreply.github.com> --- src/derive/command.rs | 4 +--- src/derive/command/encoding.rs | 2 +- src/derive/command/endedness.rs | 6 +++--- 3 files changed, 5 insertions(+), 7 deletions(-) diff --git a/src/derive/command.rs b/src/derive/command.rs index 8ded0d3..4b42d0d 100644 --- a/src/derive/command.rs +++ b/src/derive/command.rs @@ -38,12 +38,10 @@ pub enum DeriveSubcommand { Readlen(self::readlen::DeriveReadlenArgs), /// Derives the strandedness of the RNA-Seq file. - /// This subcommand requires a GFF file. Strandedness(self::strandedness::DeriveStrandednessArgs), /// Annotates junctions in the file. - /// This subcommand requires a GFF file with features to annotate. - /// This subcommand does not "derive" anything, but is included here for + /// Note that, technically, this command doesn't derive anything—it will moved in the future to a better home. /// convenience. JunctionAnnotation(self::junction_annotation::JunctionAnnotationArgs), } diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index 5b3ee52..7489365 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -20,7 +20,7 @@ pub struct DeriveEncodingArgs { #[arg(value_name = "BAM")] src: PathBuf, - /// Only examine the first n records in the file. + /// Examine the first `n` records in the file. #[arg(short, long, value_name = "USIZE")] num_records: Option, } diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 8cf78a5..378fb71 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -29,7 +29,7 @@ pub struct DeriveEndednessArgs { num_records: Option, /// Distance from 0.5 split between number of f+l- reads and f-l+ reads - /// allowed to be called 'Paired-End'. Default of `0.0` only appropriate + /// allowed to be called 'Paired-End'. The default value of `0.0` is only appropriate /// if the whole file is being processed. #[arg(long, value_name = "F64", default_value = "0.0")] paired_deviance: f64, @@ -37,12 +37,12 @@ pub struct DeriveEndednessArgs { /// Calculate and output Reads-Per-Template. This will produce a more /// sophisticated estimate for endedness, but uses substantially more memory. #[arg(long, default_value = "false")] - calc_rpt: bool, + calculate_reads_per_template: bool, /// Round RPT to the nearest INT before comparing to expected values. /// Appropriate if using `-n` > 0. Unrounded value is reported in output. #[arg(long, default_value = "false")] - round_rpt: bool, + round_reads_per_template: bool, } /// Main function for the `ngs derive endedness` subcommand. From 451a2a4a30d316522b064a591a45c4db6b268125 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 22 Mar 2024 09:32:42 -0400 Subject: [PATCH 85/91] fix(derive/endedness): complete rename from last commit --- src/derive/command/endedness.rs | 9 +++++++-- 1 file changed, 7 insertions(+), 2 deletions(-) diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 378fb71..78082c7 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -81,7 +81,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { let read_group = get_read_group(&record, Some(&mut found_rgs)); - if args.calc_rpt { + if args.calculate_reads_per_template { let read_name_map = read_names.get_or_insert_with(HashMap::new); match record.read_name() { Some(rn) => { @@ -145,7 +145,12 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } // (2) Derive the endedness based on the ordering flags gathered. - let result = compute::predict(ordering_flags, read_names, paired_deviance, args.round_rpt); + let result = compute::predict( + ordering_flags, + read_names, + paired_deviance, + args.round_reads_per_template, + ); // (3) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). From aa0880d27fd1c403a9eb58230e705762ab39bceb Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 22 Mar 2024 14:49:49 -0400 Subject: [PATCH 86/91] feat: use NonZeroUsize for Number of Records --- src/convert/command.rs | 5 +++-- src/derive/command/encoding.rs | 5 +++-- src/derive/command/endedness.rs | 7 ++++--- src/derive/command/instrument.rs | 13 +++++++++---- src/derive/command/readlen.rs | 13 +++++++++---- src/qc/command.rs | 5 +++-- src/utils/args.rs | 7 ++++--- src/utils/display.rs | 2 +- 8 files changed, 36 insertions(+), 21 deletions(-) diff --git a/src/convert/command.rs b/src/convert/command.rs index ea20904..4f06f7b 100644 --- a/src/convert/command.rs +++ b/src/convert/command.rs @@ -1,5 +1,6 @@ //! Functionality related to the `ngs convert` command itself. +use std::num::NonZeroUsize; use std::path::PathBuf; use anyhow::bail; @@ -31,8 +32,8 @@ pub struct ConvertArgs { to: PathBuf, /// Number of records to process before exiting the conversion. - #[arg(short = 'n', long, value_name = "USIZE")] - num_records: Option, + #[arg(short = 'n', long, value_name = "NonZeroUsize")] + num_records: Option, /// If available, the FASTA reference file used to generate the file. #[arg(short, long)] diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index 7489365..1949a6c 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -6,6 +6,7 @@ use noodles::bam; use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; use std::io::BufReader; +use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -21,8 +22,8 @@ pub struct DeriveEncodingArgs { src: PathBuf, /// Examine the first `n` records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + #[arg(short, long, value_name = "NonZeroUsize")] + num_records: Option, } /// Main function for the `ngs derive encoding` subcommand. diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 78082c7..da14afd 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -4,6 +4,7 @@ use anyhow::Context; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::{HashMap, HashSet}; +use std::num::NonZeroUsize; use std::path::PathBuf; use std::sync::Arc; use tracing::{info, trace}; @@ -24,9 +25,9 @@ pub struct DeriveEndednessArgs { #[arg(value_name = "BAM")] src: PathBuf, - /// Only examine the first n records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + /// Examine the first `n` records in the file. + #[arg(short, long, value_name = "NonZeroUsize")] + num_records: Option, /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. The default value of `0.0` is only appropriate diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index a0776f8..ccbe520 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -4,6 +4,7 @@ use anyhow::bail; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; +use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -21,9 +22,10 @@ pub struct DeriveInstrumentArgs { #[arg(value_name = "BAM")] src: PathBuf, - /// Only examine the first n records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + /// Examine the first `n` records in the file. + /// If `0`, all records are examined. + #[arg(short, long, value_name = "USIZE", default_value = "10000000")] + num_records: usize, } /// Main function for the `ngs derive instrument` subcommand. @@ -41,7 +43,10 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { // (1) Collect instrument names and flowcell names from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = NumberOfRecords::from(args.num_records); + let num_records = match args.num_records { + 0 => NumberOfRecords::All, + _ => NumberOfRecords::Some(NonZeroUsize::new(args.num_records).unwrap()), + }; let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index f5c2246..031abe4 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -4,6 +4,7 @@ use anyhow::Context; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::HashMap; +use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -21,9 +22,10 @@ pub struct DeriveReadlenArgs { #[arg(value_name = "BAM")] src: PathBuf, - /// Only examine the first n records in the file. - #[arg(short, long, value_name = "USIZE")] - num_records: Option, + /// Examine the first `n` records in the file. + /// If `0`, all records are examined. + #[arg(short, long, value_name = "USIZE", default_value = "10000000")] + num_records: usize, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. #[arg(short, long, value_name = "F64", default_value = "0.7")] @@ -46,7 +48,10 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = NumberOfRecords::from(args.num_records); + let num_records = match args.num_records { + 0 => NumberOfRecords::All, + _ => NumberOfRecords::Some(NonZeroUsize::new(args.num_records).unwrap()), + }; let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { diff --git a/src/qc/command.rs b/src/qc/command.rs index d628c00..c6993d7 100644 --- a/src/qc/command.rs +++ b/src/qc/command.rs @@ -1,6 +1,7 @@ //! Functionality related to the `ngs qc` command itself. use std::fs::File; +use std::num::NonZeroUsize; use std::path::PathBuf; use std::rc::Rc; @@ -50,8 +51,8 @@ pub struct QcArgs { /// to process per sequence in the second pass. /// /// This is generally only used for testing purposes. - #[arg(short = 'n', long, value_name = "USIZE")] - num_records: Option, + #[arg(short = 'n', long, value_name = "NonZeroUsize")] + num_records: Option, /// Directory to output files to. Defaults to current working directory. #[arg(short = 'o', long, value_name = "PATH")] diff --git a/src/utils/args.rs b/src/utils/args.rs index 094819a..7bd3ce0 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -1,6 +1,7 @@ //! Utilities related to the parsing of arguments. use std::fmt::Display; +use std::num::NonZeroUsize; use noodles::{bgzf::writer::CompressionLevel, sam::record::MappingQuality}; use tracing::debug; @@ -17,11 +18,11 @@ pub enum NumberOfRecords { /// Designates that we should review _some_ of the records in the file. The /// exact count of records is stored in the `usize`. - Some(usize), + Some(NonZeroUsize), } -impl From> for NumberOfRecords { - fn from(num_records: Option) -> Self { +impl From> for NumberOfRecords { + fn from(num_records: Option) -> Self { match num_records { Some(n) => { debug!("Reading a maximum of {} records.", n); diff --git a/src/utils/display.rs b/src/utils/display.rs index 7ee4f56..1ffbbca 100644 --- a/src/utils/display.rs +++ b/src/utils/display.rs @@ -74,7 +74,7 @@ impl RecordCounter { /// (if it exists, otherwise it loops forever). pub fn time_to_break(&self, limit: &NumberOfRecords) -> bool { match limit { - NumberOfRecords::Some(v) => self.count >= *v, + NumberOfRecords::Some(v) => self.count >= ::from(*v), NumberOfRecords::All => false, } } From 406a7b88836dff99f1e20e980b9ebb73f1230da1 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Fri, 22 Mar 2024 21:32:49 -0400 Subject: [PATCH 87/91] feat(utils/args): improved behavior for NumberOfRecords CL utility --- src/convert/command.rs | 12 ++++--- src/derive/command/encoding.rs | 14 ++++---- src/derive/command/endedness.rs | 14 ++++---- src/derive/command/instrument.rs | 18 +++++----- src/derive/command/readlen.rs | 18 +++++----- src/qc/command.rs | 14 ++++---- src/utils/args.rs | 59 ++++++++++++++++++++++++++++++++ 7 files changed, 107 insertions(+), 42 deletions(-) diff --git a/src/convert/command.rs b/src/convert/command.rs index 4f06f7b..76a7a43 100644 --- a/src/convert/command.rs +++ b/src/convert/command.rs @@ -1,6 +1,5 @@ //! Functionality related to the `ngs convert` command itself. -use std::num::NonZeroUsize; use std::path::PathBuf; use anyhow::bail; @@ -32,8 +31,13 @@ pub struct ConvertArgs { to: PathBuf, /// Number of records to process before exiting the conversion. - #[arg(short = 'n', long, value_name = "NonZeroUsize")] - num_records: Option, + #[arg( + short, + long, + default_value_t, + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, /// If available, the FASTA reference file used to generate the file. #[arg(short, long)] @@ -92,7 +96,7 @@ pub fn convert(args: ConvertArgs) -> anyhow::Result<()> { // Number of Records // //===================// - let max_records = NumberOfRecords::from(args.num_records); + let max_records = args.num_records; //==========================// // Bioinformatics File Pair // diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index 1949a6c..8c591cf 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -6,7 +6,6 @@ use noodles::bam; use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; use std::io::BufReader; -use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -22,8 +21,13 @@ pub struct DeriveEncodingArgs { src: PathBuf, /// Examine the first `n` records in the file. - #[arg(short, long, value_name = "NonZeroUsize")] - num_records: Option, + #[arg( + short, + long, + default_value_t, + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, } /// Main function for the `ngs derive encoding` subcommand. @@ -42,9 +46,7 @@ pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { // (1) Collect quality scores from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = NumberOfRecords::from(args.num_records); let mut counter = RecordCounter::default(); - for result in reader.lazy_records() { let record = result?; @@ -54,7 +56,7 @@ pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { } counter.inc(); - if counter.time_to_break(&num_records) { + if counter.time_to_break(&args.num_records) { break; } } diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index da14afd..80b0561 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -4,7 +4,6 @@ use anyhow::Context; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::{HashMap, HashSet}; -use std::num::NonZeroUsize; use std::path::PathBuf; use std::sync::Arc; use tracing::{info, trace}; @@ -26,8 +25,13 @@ pub struct DeriveEndednessArgs { src: PathBuf, /// Examine the first `n` records in the file. - #[arg(short, long, value_name = "NonZeroUsize")] - num_records: Option, + #[arg( + short, + long, + default_value_t, + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, /// Distance from 0.5 split between number of f+l- reads and f-l+ reads /// allowed to be called 'Paired-End'. The default value of `0.0` is only appropriate @@ -67,9 +71,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { // (1) Collect ordering flags (and QNAMEs) from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = NumberOfRecords::from(args.num_records); let mut counter = RecordCounter::default(); - for result in reader.records(&header.parsed) { let record = result?; @@ -129,7 +131,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { } counter.inc(); - if counter.time_to_break(&num_records) { + if counter.time_to_break(&args.num_records) { break; } } diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index ccbe520..1448005 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -4,7 +4,6 @@ use anyhow::bail; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::HashSet; -use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -23,9 +22,13 @@ pub struct DeriveInstrumentArgs { src: PathBuf, /// Examine the first `n` records in the file. - /// If `0`, all records are examined. - #[arg(short, long, value_name = "USIZE", default_value = "10000000")] - num_records: usize, + #[arg( + short, + long, + default_value = "10000000", + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, } /// Main function for the `ngs derive instrument` subcommand. @@ -43,12 +46,7 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { // (1) Collect instrument names and flowcell names from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = match args.num_records { - 0 => NumberOfRecords::All, - _ => NumberOfRecords::Some(NonZeroUsize::new(args.num_records).unwrap()), - }; let mut counter = RecordCounter::default(); - for result in reader.records(&header.parsed) { let record = result?; @@ -76,7 +74,7 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { } counter.inc(); - if counter.time_to_break(&num_records) { + if counter.time_to_break(&args.num_records) { break; } } diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index 031abe4..c13e923 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -4,7 +4,6 @@ use anyhow::Context; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::HashMap; -use std::num::NonZeroUsize; use std::path::PathBuf; use tracing::info; @@ -23,9 +22,13 @@ pub struct DeriveReadlenArgs { src: PathBuf, /// Examine the first `n` records in the file. - /// If `0`, all records are examined. - #[arg(short, long, value_name = "USIZE", default_value = "10000000")] - num_records: usize, + #[arg( + short, + long, + default_value = "10000000", + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, /// Majority vote cutoff value as a fraction between [0.0, 1.0]. #[arg(short, long, value_name = "F64", default_value = "0.7")] @@ -48,12 +51,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { // (1) Collect read lengths from reads within the // file. Support for sampling only a portion of the reads is provided. - let num_records = match args.num_records { - 0 => NumberOfRecords::All, - _ => NumberOfRecords::Some(NonZeroUsize::new(args.num_records).unwrap()), - }; let mut counter = RecordCounter::default(); - for result in reader.records(&header.parsed) { let record = result?; let len = record.sequence().len(); @@ -61,7 +59,7 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { *read_lengths.entry(len).or_default() += 1; counter.inc(); - if counter.time_to_break(&num_records) { + if counter.time_to_break(&args.num_records) { break; } } diff --git a/src/qc/command.rs b/src/qc/command.rs index c6993d7..6fac58d 100644 --- a/src/qc/command.rs +++ b/src/qc/command.rs @@ -1,7 +1,6 @@ //! Functionality related to the `ngs qc` command itself. use std::fs::File; -use std::num::NonZeroUsize; use std::path::PathBuf; use std::rc::Rc; @@ -51,8 +50,13 @@ pub struct QcArgs { /// to process per sequence in the second pass. /// /// This is generally only used for testing purposes. - #[arg(short = 'n', long, value_name = "NonZeroUsize")] - num_records: Option, + #[arg( + short, + long, + default_value_t, + value_name = "'all' or a positive, non-zero integer" + )] + num_records: NumberOfRecords, /// Directory to output files to. Defaults to current working directory. #[arg(short = 'o', long, value_name = "PATH")] @@ -202,8 +206,6 @@ pub fn qc(args: QcArgs) -> anyhow::Result<()> { // Number of Records // //===================// - let num_records = NumberOfRecords::from(args.num_records); - app( src, reference_fasta, @@ -211,7 +213,7 @@ pub fn qc(args: QcArgs) -> anyhow::Result<()> { reference_genome, output_prefix, output_directory, - num_records, + args.num_records, feature_names, only_facet, vaf_file_path, diff --git a/src/utils/args.rs b/src/utils/args.rs index 7bd3ce0..8d5bedf 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -12,6 +12,7 @@ use tracing::debug; /// Utility enum to designate whether we are reviewing all records in the file /// or just some of them. +#[derive(Clone, Debug)] pub enum NumberOfRecords { /// Designates that we should review _all_ of the records in the file. All, @@ -21,6 +22,64 @@ pub enum NumberOfRecords { Some(NonZeroUsize), } +impl std::default::Default for NumberOfRecords { + fn default() -> Self { + Self::All + } +} + +impl std::fmt::Display for NumberOfRecords { + fn fmt(&self, f: &mut std::fmt::Formatter<'_>) -> std::fmt::Result { + match self { + NumberOfRecords::All => write!(f, "all"), + NumberOfRecords::Some(value) => write!(f, "{value}"), + } + } +} + +/// An error type for parsing the number of records. +#[derive(Debug)] +pub enum NumberOfRecordsError { + /// The number of records is invalid. + Invalid(String), +} + +impl std::fmt::Display for NumberOfRecordsError { + fn fmt(&self, f: &mut std::fmt::Formatter<'_>) -> std::fmt::Result { + match self { + NumberOfRecordsError::Invalid(value) => write!(f, "invalid number of reads: {value}"), + } + } +} + +impl std::error::Error for NumberOfRecordsError {} + +impl std::str::FromStr for NumberOfRecords { + type Err = NumberOfRecordsError; + + fn from_str(s: &str) -> Result { + match s.to_lowercase().as_str() { + "all" => Ok(NumberOfRecords::All), + _ => s + .parse::() + .map_err(|_| { + NumberOfRecordsError::Invalid(String::from( + "must be a positive, non-zero integer or 'all'", + )) + }) + .and_then(|num| { + NonZeroUsize::new(num) + .ok_or_else(|| { + NumberOfRecordsError::Invalid(String::from( + "integers must be positive and non-zero", + )) + }) + .map(NumberOfRecords::Some) + }), + } + } +} + impl From> for NumberOfRecords { fn from(num_records: Option) -> Self { match num_records { From f83015f0f496cc47269865786dcb793dc425886f Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Sat, 23 Mar 2024 18:08:09 -0400 Subject: [PATCH 88/91] feat(derive): report by read group where appropriate and feasible --- src/derive/command/encoding.rs | 2 +- src/derive/command/endedness.rs | 6 +- src/derive/command/instrument.rs | 35 +++-- src/derive/command/readlen.rs | 25 +++- src/derive/instrument/compute.rs | 211 +++++++++++++++++++++++++------ src/derive/readlen/compute.rs | 169 ++++++++++++++++++++----- 6 files changed, 359 insertions(+), 89 deletions(-) diff --git a/src/derive/command/encoding.rs b/src/derive/command/encoding.rs index 8c591cf..ceaef6b 100644 --- a/src/derive/command/encoding.rs +++ b/src/derive/command/encoding.rs @@ -32,7 +32,7 @@ pub struct DeriveEncodingArgs { /// Main function for the `ngs derive encoding` subcommand. pub fn derive(args: DeriveEncodingArgs) -> anyhow::Result<()> { - info!("Starting derive readlen subcommand."); + info!("Starting derive encoding subcommand."); let file = std::fs::File::open(args.src); let reader = file diff --git a/src/derive/command/endedness.rs b/src/derive/command/endedness.rs index 80b0561..b7d38c6 100644 --- a/src/derive/command/endedness.rs +++ b/src/derive/command/endedness.rs @@ -141,13 +141,13 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { counter.get().to_formatted_string(&Locale::en) ); - // (1.5) Validate the read group information. + // (2) Validate the read group information. let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); for rg_id in rgs_in_header_not_records { ordering_flags.insert(Arc::new(rg_id), OrderingFlagsCounts::new()); } - // (2) Derive the endedness based on the ordering flags gathered. + // (3) Derive the endedness based on the ordering flags gathered. let result = compute::predict( ordering_flags, read_names, @@ -155,7 +155,7 @@ pub fn derive(args: DeriveEndednessArgs) -> anyhow::Result<()> { args.round_reads_per_template, ); - // (3) Print the output to stdout as JSON (more support for different output + // (4) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); println!("{}", output); diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 1448005..5913694 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -1,10 +1,10 @@ //! Functionality relating to the `ngs derive instrument` subcommand itself. -use anyhow::bail; use clap::Args; use num_format::{Locale, ToFormattedString}; -use std::collections::HashSet; +use std::collections::{HashMap, HashSet}; use std::path::PathBuf; +use std::sync::Arc; use tracing::info; use crate::derive::instrument::compute; @@ -13,6 +13,7 @@ use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; +use crate::utils::read_groups::{get_read_group, validate_read_group_info, ReadGroupPtr}; /// Clap arguments for the `ngs derive instrument` subcommand. #[derive(Args)] @@ -34,9 +35,10 @@ pub struct DeriveInstrumentArgs { /// Main function for the `ngs derive instrument` subcommand. pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { let src = args.src; - let mut instrument_names = HashSet::new(); - let mut flowcell_names = HashSet::new(); + let mut instrument_names: HashMap> = HashMap::new(); + let mut flowcell_names: HashMap> = HashMap::new(); let mut metrics = compute::RecordMetrics::default(); + let mut found_rgs = HashSet::new(); info!("Starting derive instrument subcommand."); @@ -49,24 +51,25 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; + let read_group = get_read_group(&record, Some(&mut found_rgs)); if let Some(read_name) = record.read_name() { let name: &str = read_name.as_ref(); match name.parse::() { Ok(read) => { - instrument_names.insert(read.instrument_name); + instrument_names + .entry(read_group.clone()) + .or_default() + .insert(read.instrument_name); metrics.found_instrument_name += 1; if let Some(fc) = read.flowcell { - flowcell_names.insert(fc); + flowcell_names.entry(read_group).or_default().insert(fc); metrics.found_flowcell_name += 1; } } Err(_) => { - bail!( - "Could not parse Illumina-formatted query names for read: {}", - name - ); + metrics.bad_read_name += 1; } } } else { @@ -85,12 +88,20 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { ); metrics.total_records = counter.get(); - // (2) Derive the predict instrument results based on these detected + // (2) Validate the read group information. + let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); + for rg_id in rgs_in_header_not_records { + let rg_ptr = Arc::new(rg_id); + instrument_names.insert(rg_ptr.clone(), HashSet::new()); + flowcell_names.insert(rg_ptr, HashSet::new()); + } + + // (3) Derive the instrument results based on the detected // instrument names and flowcell names. let mut result = compute::predict(instrument_names, flowcell_names); result.records = metrics; - // (3) Print the output to stdout as JSON (more support for different output + // (4) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); println!("{}", output); diff --git a/src/derive/command/readlen.rs b/src/derive/command/readlen.rs index c13e923..1cf46d8 100644 --- a/src/derive/command/readlen.rs +++ b/src/derive/command/readlen.rs @@ -4,7 +4,9 @@ use anyhow::Context; use clap::Args; use num_format::{Locale, ToFormattedString}; use std::collections::HashMap; +use std::collections::HashSet; use std::path::PathBuf; +use std::sync::Arc; use tracing::info; use crate::derive::readlen::compute; @@ -13,6 +15,7 @@ use crate::utils::args::NumberOfRecords; use crate::utils::display::RecordCounter; use crate::utils::formats::bam::ParsedBAMFile; use crate::utils::formats::utils::IndexCheck; +use crate::utils::read_groups::{get_read_group, validate_read_group_info, ReadGroupPtr}; /// Clap arguments for the `ngs derive readlen` subcommand. #[derive(Args)] @@ -41,7 +44,8 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let majority_vote_cutoff = cutoff_in_range(args.majority_vote_cutoff, 0.0..=1.0) .with_context(|| "Majority vote cutoff is not within acceptable range")?; - let mut read_lengths = HashMap::new(); + let mut read_lengths: HashMap> = HashMap::new(); + let mut found_rgs = HashSet::new(); info!("Starting derive readlen subcommand."); @@ -54,9 +58,14 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { let mut counter = RecordCounter::default(); for result in reader.records(&header.parsed) { let record = result?; + let read_group = get_read_group(&record, Some(&mut found_rgs)); let len = record.sequence().len(); - *read_lengths.entry(len).or_default() += 1; + *read_lengths + .entry(read_group) + .or_default() + .entry(len) + .or_default() += 1; counter.inc(); if counter.time_to_break(&args.num_records) { @@ -69,10 +78,16 @@ pub fn derive(args: DeriveReadlenArgs) -> anyhow::Result<()> { counter.get().to_formatted_string(&Locale::en) ); - // (2) Derive the consensus read length based on the read lengths gathered. - let result = compute::predict(read_lengths, counter.get(), majority_vote_cutoff).unwrap(); + // (2) Validate the read group information. + let rgs_in_header_not_records = validate_read_group_info(&found_rgs, &header.parsed); + for rg_id in rgs_in_header_not_records { + read_lengths.insert(Arc::new(rg_id), HashMap::new()); + } + + // (3) Derive the consensus read length based on the read lengths gathered. + let result = compute::predict(read_lengths, majority_vote_cutoff); - // (3) Print the output to stdout as JSON (more support for different output + // (4) Print the output to stdout as JSON (more support for different output // types may be added in the future, but for now, only JSON). let output = serde_json::to_string_pretty(&result).unwrap(); println!("{}", output); diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index c51070d..4256f96 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -5,6 +5,7 @@ use serde::Serialize; use std::collections::{HashMap, HashSet}; use crate::derive::instrument::{flowcells, instruments}; +use crate::utils::read_groups::ReadGroupPtr; /// Generalized struct for holding instrument detection results. #[derive(Debug, Default, Serialize)] @@ -58,15 +59,61 @@ pub struct RecordMetrics { /// due to a missing or invalid read name. pub bad_read_name: usize, - /// The total number of records that contained a valid + /// The total number of records that contained a parseable /// instrument name in their read name. pub found_instrument_name: usize, - /// The total number of records that contained a valid + /// The total number of records that contained a parseable /// flowcell name in their read name. pub found_flowcell_name: usize, } +/// Struct holding the per read group results for an `ngs derive instrument` +/// subcommand call. +#[derive(Debug, Serialize)] +pub struct ReadGroupDerivedInstrumentResult { + /// The read group that these results are associated with. + pub read_group: String, + + /// Whether or not the `ngs derive instrument` subcommand succeeded + /// for this read group. + pub succeeded: bool, + + /// The possible instruments detected for this read group, if derivable. + pub instruments: Option>, + + /// The level of confidence that the tool has concerning these results. + pub confidence: String, + + /// Status of the evidence that supports (or lack thereof) these predicted + /// instruments, if available. + pub evidence: Option, + + /// A general comment field, if available. + pub comment: Option, + + /// The results of the instrument name look-ups for this read group. + pub instrument_name_queries: Vec, + + /// The results of the flowcell name look-ups for this read group. + pub flowcell_name_queries: Vec, +} + +impl Default for ReadGroupDerivedInstrumentResult { + fn default() -> Self { + ReadGroupDerivedInstrumentResult { + read_group: String::new(), + succeeded: false, + instruments: None, + confidence: "unknown".to_string(), + evidence: None, + comment: None, + instrument_name_queries: Vec::new(), + flowcell_name_queries: Vec::new(), + } + } +} + /// Struct holding the final results for an `ngs derive instrument` subcommand /// call. #[derive(Debug, Serialize)] @@ -88,11 +135,10 @@ pub struct DerivedInstrumentResult { /// A general comment field, if available. pub comment: Option, - /// The results of the instrument name look-ups. - pub instrument_name_queries: Vec, - - /// The results of the flowcell name look-ups. - pub flowcell_name_queries: Vec, + /// Vector of [`ReadGroupDerivedInstrumentResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, /// Metrics related to how read records were processed. pub records: RecordMetrics, @@ -106,8 +152,7 @@ impl Default for DerivedInstrumentResult { confidence: "unknown".to_string(), evidence: None, comment: None, - instrument_name_queries: Vec::new(), - flowcell_name_queries: Vec::new(), + read_groups: Vec::new(), records: RecordMetrics::default(), } } @@ -186,15 +231,15 @@ pub fn predict_instrument( } /// Combines evidence from the instrument id detection and flowcell id detection -/// to produce a [`DerivedInstrumentResult`]. +/// to produce a [`ReadGroupDerivedInstrumentResult`]. pub fn resolve_instrument_prediction( iid_results: InstrumentDetectionResults, fcid_results: InstrumentDetectionResults, -) -> DerivedInstrumentResult { +) -> ReadGroupDerivedInstrumentResult { let possible_instruments_by_iid = iid_results.possible_instruments.unwrap_or_default(); let possible_instruments_by_fcid = fcid_results.possible_instruments.unwrap_or_default(); - let mut result = DerivedInstrumentResult::default(); + let mut result = ReadGroupDerivedInstrumentResult::default(); // (1) If the set of possible instruments as determined by the instrument id // is empty _and_ we have seen at least one machine, then the only possible @@ -284,19 +329,45 @@ pub fn resolve_instrument_prediction( /// return a result for the derived instruments. This may fail, and the /// resulting [`DerivedInstrumentResult`] should be evaluated accordingly. pub fn predict( - instrument_names: HashSet, - flowcell_names: HashSet, + instrument_names: HashMap>, + flowcell_names: HashMap>, ) -> DerivedInstrumentResult { let instruments = instruments::build_instrument_lookup_table(); let flowcells = flowcells::build_flowcell_lookup_table(); - let (iid_results, instrument_name_queries) = predict_instrument(instrument_names, &instruments); - let (fcid_results, flowcell_name_queries) = predict_instrument(flowcell_names, &flowcells); + let mut rg_results = Vec::new(); + let mut all_instrument_names = HashSet::new(); + let mut all_flowcell_names = HashSet::new(); - let mut final_results = resolve_instrument_prediction(iid_results, fcid_results); - final_results.instrument_name_queries = instrument_name_queries; - final_results.flowcell_name_queries = flowcell_name_queries; - final_results + for rg in instrument_names.keys() { + all_instrument_names.extend(instrument_names[rg].iter().cloned()); + all_flowcell_names.extend(flowcell_names[rg].iter().cloned()); + + let (rg_iid_results, rg_instrument_name_queries) = + predict_instrument(instrument_names[rg].clone(), &instruments); + let (rg_fcid_results, rg_flowcell_name_queries) = + predict_instrument(flowcell_names[rg].clone(), &flowcells); + + let mut rg_result = resolve_instrument_prediction(rg_iid_results, rg_fcid_results); + rg_result.read_group = rg.to_string(); + rg_result.instrument_name_queries = rg_instrument_name_queries; + rg_result.flowcell_name_queries = rg_flowcell_name_queries; + rg_results.push(rg_result); + } + + let (iid_results, _) = predict_instrument(all_instrument_names, &instruments); + let (fcid_results, _) = predict_instrument(all_flowcell_names, &flowcells); + + let overall_prediction = resolve_instrument_prediction(iid_results, fcid_results); + DerivedInstrumentResult { + succeeded: overall_prediction.succeeded, + instruments: overall_prediction.instruments, + confidence: overall_prediction.confidence, + evidence: overall_prediction.evidence, + comment: overall_prediction.comment, + read_groups: rg_results, + ..DerivedInstrumentResult::default() + } } #[cfg(test)] @@ -335,8 +406,14 @@ mod tests { #[test] fn test_derive_instrument_novaseq_succesfully() { - let detected_iids = HashSet::from(["A00000".to_string()]); - let detected_fcids = HashSet::from(["H00000RXX".to_string()]); + let detected_iids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["A00000".to_string()]), + )]); + let detected_fcids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H00000RXX".to_string()]), + )]); let result = predict(detected_iids, detected_fcids); assert!(result.succeeded); @@ -354,8 +431,23 @@ mod tests { #[test] fn test_derive_instrument_conflicting_instrument_ids() { - let detected_iids = HashSet::from(["A00000".to_string(), "D00000".to_string()]); - let detected_fcids = HashSet::from(["H00000RXX".to_string()]); + let detected_iids = HashMap::from([ + ( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["A00000".to_string()]), + ), + ( + ReadGroupPtr::from("RG2".to_string()), + HashSet::from(["D00000".to_string()]), + ), + ]); + let detected_fcids = HashMap::from([ + ( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H00000RXX".to_string()]), + ), + (ReadGroupPtr::from("RG2".to_string()), HashSet::new()), + ]); let result = predict(detected_iids, detected_fcids); assert!(!result.succeeded); @@ -372,8 +464,23 @@ mod tests { #[test] fn test_derive_instrument_conflicting_flowcell_ids() { - let detected_iids = HashSet::from(["A00000".to_string()]); - let detected_fcids = HashSet::from(["H00000RXX".to_string(), "B0000".to_string()]); + let detected_iids = HashMap::from([ + ( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["A00000".to_string()]), + ), + (ReadGroupPtr::from("RG2".to_string()), HashSet::new()), + ]); + let detected_fcids = HashMap::from([ + ( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H00000RXX".to_string()]), + ), + ( + ReadGroupPtr::from("RG2".to_string()), + HashSet::from(["B0000".to_string()]), + ), + ]); let result = predict(detected_iids, detected_fcids); assert!(!result.succeeded); @@ -388,8 +495,12 @@ mod tests { #[test] fn test_derive_instrument_medium_instrument_evidence() { - let detected_iids = HashSet::from(["A00000".to_string()]); - let detected_fcids = HashSet::new(); + let detected_iids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["A00000".to_string()]), + )]); + let detected_fcids = + HashMap::from([(ReadGroupPtr::from("RG1".to_string()), HashSet::new())]); let result = predict(detected_iids, detected_fcids); assert!(result.succeeded); @@ -404,8 +515,12 @@ mod tests { #[test] fn test_derive_instrument_low_instrument_evidence() { - let detected_iids = HashSet::from(["K00000".to_string()]); - let detected_fcids = HashSet::new(); + let detected_iids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["K00000".to_string()]), + )]); + let detected_fcids = + HashMap::from([(ReadGroupPtr::from("RG1".to_string()), HashSet::new())]); let result = predict(detected_iids, detected_fcids); assert!(result.succeeded); @@ -423,8 +538,12 @@ mod tests { #[test] fn test_derive_instrument_medium_flowcell_evidence() { - let detected_iids = HashSet::new(); - let detected_fcids = HashSet::from(["H00000RXX".to_string()]); + let detected_iids = + HashMap::from([(ReadGroupPtr::from("RG1".to_string()), HashSet::new())]); + let detected_fcids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H00000RXX".to_string()]), + )]); let result = predict(detected_iids, detected_fcids); assert!(result.succeeded); @@ -439,8 +558,12 @@ mod tests { #[test] fn test_derive_instrument_low_flowcell_evidence() { - let detected_iids = HashSet::new(); - let detected_fcids = HashSet::from(["H0000ADXX".to_string()]); + let detected_iids = + HashMap::from([(ReadGroupPtr::from("RG1".to_string()), HashSet::new())]); + let detected_fcids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H0000ADXX".to_string()]), + )]); let result = predict(detected_iids, detected_fcids); assert!(result.succeeded); @@ -459,8 +582,14 @@ mod tests { #[test] fn test_derive_instrument_conflicting_flowcell_and_instrument_evidence() { - let detected_iids = HashSet::from(["K00000".to_string()]); - let detected_fcids = HashSet::from(["H00000RXX".to_string()]); + let detected_iids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["K00000".to_string()]), + )]); + let detected_fcids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["H00000RXX".to_string()]), + )]); let result = predict(detected_iids, detected_fcids); assert!(!result.succeeded); @@ -475,8 +604,14 @@ mod tests { #[test] fn test_derive_instrument_no_matches() { - let detected_iids = HashSet::from(["QQQQQ".to_string()]); - let detected_fcids = HashSet::from(["ZZZZZZ".to_string()]); + let detected_iids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["QQQQQ".to_string()]), + )]); + let detected_fcids = HashMap::from([( + ReadGroupPtr::from("RG1".to_string()), + HashSet::from(["ZZZZZZ".to_string()]), + )]); let result = predict(detected_iids, detected_fcids); assert!(!result.succeeded); diff --git a/src/derive/readlen/compute.rs b/src/derive/readlen/compute.rs index 97a3227..591092d 100644 --- a/src/derive/readlen/compute.rs +++ b/src/derive/readlen/compute.rs @@ -1,8 +1,51 @@ //! Module holding the logic for computing the consensus read length. -use anyhow::bail; use serde::Serialize; use std::collections::HashMap; +use tracing::warn; + +use crate::utils::read_groups::ReadGroupPtr; + +/// Struct holding the per read group results for an `ngs derive readlen` +/// subcommand call. +#[derive(Debug, Serialize)] +pub struct ReadGroupDerivedReadlenResult { + /// The read group that these results are associated with. + pub read_group: String, + + /// Whether or not the `ngs derive readlen` subcommand succeeded + /// for this read group. + pub succeeded: bool, + + /// The consensus read length, if derivable. + pub consensus_read_length: Option, + + /// The majority vote percentage of the consensus read length. + pub majority_pct_detected: f64, + + /// Status of the evidence that supports (or does not support) the + /// consensus read length. + pub evidence: Vec<(usize, usize)>, +} + +impl ReadGroupDerivedReadlenResult { + /// Creates a new [`ReadGroupDerivedReadlenResult`]. + pub fn new( + read_group: String, + succeeded: bool, + consensus_read_length: Option, + majority_pct_detected: f64, + evidence: Vec<(usize, usize)>, + ) -> Self { + ReadGroupDerivedReadlenResult { + read_group, + succeeded, + consensus_read_length, + majority_pct_detected, + evidence, + } + } +} /// Struct holding the final results for an `ngs derive readlen` subcommand /// call. @@ -17,6 +60,11 @@ pub struct DerivedReadlenResult { /// The majority vote percentage of the consensus read length. pub majority_pct_detected: f64, + /// Vector of [`ReadGroupDerivedReadlenResult`]s. + /// One for each read group in the BAM, + /// and potentially one for any reads with an unknown read group. + pub read_groups: Vec, + /// Status of the evidence that supports (or does not support) the /// consensus read length. pub evidence: Vec<(usize, usize)>, @@ -28,70 +76,122 @@ impl DerivedReadlenResult { succeeded: bool, consensus_read_length: Option, majority_pct_detected: f64, + read_groups: Vec, evidence: Vec<(usize, usize)>, ) -> Self { DerivedReadlenResult { succeeded, consensus_read_length, majority_pct_detected, + read_groups, evidence, } } } -/// Main method to evaluate the collected read lengths and -/// return a result for the consensus read length. This may fail, and the -/// resulting [`DerivedReadlenResult`] should be evaluated accordingly. -pub fn predict( - read_lengths: HashMap, - num_samples: usize, +/// Predicts the consensus read length for a given read group based on the +/// read lengths and a majority vote cutoff. +pub fn predict_readlen( + read_group: String, + read_lengths: &HashMap, majority_vote_cutoff: f64, -) -> Result { - if num_samples == 0 { - bail!("No read lengths were detected in the file."); - } +) -> ReadGroupDerivedReadlenResult { + let mut read_lengths: Vec<(usize, usize)> = + read_lengths.iter().map(|(k, v)| (*k, *v)).collect(); - // Sort the read lengths by their key for output. - let mut read_lengths: Vec<(usize, usize)> = read_lengths.into_iter().collect(); read_lengths.sort_by(|a, b| b.0.cmp(&a.0)); - let max_read_length = read_lengths[0].0; - let max_count = read_lengths[0].1; + // Tally the number of reads + let num_reads: usize = read_lengths.iter().map(|(_, count)| count).sum(); - let consensus_read_length = max_read_length; - let majority_detected = max_count as f64 / num_samples as f64; + let (majority_detected, consensus_read_length) = match num_reads == 0 { + true => { + warn!("No reads were detected for read group: {}", read_group); + (0.0, None) + } + false => ( + read_lengths[0].1 as f64 / num_reads as f64, + Some(read_lengths[0].0), + ), + }; match majority_detected >= majority_vote_cutoff { - true => anyhow::Ok(DerivedReadlenResult::new( + true => ReadGroupDerivedReadlenResult::new( + read_group, true, - Some(consensus_read_length), + consensus_read_length, majority_detected * 100.0, read_lengths, - )), - false => anyhow::Ok(DerivedReadlenResult::new( + ), + false => ReadGroupDerivedReadlenResult::new( + read_group, false, None, majority_detected * 100.0, read_lengths, - )), + ), } } +/// Main method to evaluate the collected read lengths and +/// return a result for the consensus read length. This may fail, and the +/// resulting [`DerivedReadlenResult`] should be evaluated accordingly. +pub fn predict( + read_lengths: HashMap>, + majority_vote_cutoff: f64, +) -> DerivedReadlenResult { + // Iterate over the read lengths and predict the consensus read length. + let mut rg_results = Vec::new(); + let mut overall_lengths = HashMap::new(); + + for (read_group, lengths) in read_lengths { + let result = predict_readlen(read_group.to_string(), &lengths, majority_vote_cutoff); + rg_results.push(result); + + for (length, count) in lengths { + *overall_lengths.entry(length).or_default() += count; + } + } + + let overall_result = predict_readlen( + "overall".to_string(), + &overall_lengths, + majority_vote_cutoff, + ); + + // Sort the read lengths by their key for output. + let mut overall_lengths: Vec<(usize, usize)> = overall_lengths.into_iter().collect(); + overall_lengths.sort_by(|a, b| b.0.cmp(&a.0)); + + DerivedReadlenResult::new( + overall_result.succeeded, + overall_result.consensus_read_length, + overall_result.majority_pct_detected, + rg_results, + overall_lengths, + ) +} + #[cfg(test)] mod tests { use super::*; + use std::sync::Arc; #[test] fn test_derive_readlen_from_empty_hashmap() { let read_lengths = HashMap::new(); - let result = predict(read_lengths, 0, 0.7); - assert!(result.is_err()); + let result = predict(read_lengths, 0.7); + assert!(!result.succeeded); + assert_eq!(result.consensus_read_length, None); + assert_eq!(result.majority_pct_detected, 0.0); + assert_eq!(result.evidence, Vec::new()); } #[test] fn test_derive_readlen_when_all_readlengths_equal() { - let read_lengths = HashMap::from([(100, 10)]); - let result = predict(read_lengths, 10, 1.0).unwrap(); + let read_lengths = + HashMap::from([(Arc::new("RG1".to_string()), HashMap::from([(100, 10)]))]); + let result = predict(read_lengths, 1.0); assert!(result.succeeded); assert_eq!(result.consensus_read_length, Some(100)); assert_eq!(result.majority_pct_detected, 100.0); @@ -100,8 +200,11 @@ mod tests { #[test] fn test_derive_readlen_success_when_not_all_readlengths_equal() { - let read_lengths = HashMap::from([(101, 1000), (100, 5), (99, 5)]); - let result = predict(read_lengths, 1010, 0.7).unwrap(); + let read_lengths = HashMap::from([( + Arc::new("RG1".to_string()), + HashMap::from([(101, 1000), (100, 5), (99, 5)]), + )]); + let result = predict(read_lengths, 0.7); assert!(result.succeeded); assert_eq!(result.consensus_read_length, Some(101)); assert!(result.majority_pct_detected > 99.0); @@ -110,8 +213,14 @@ mod tests { #[test] fn test_derive_readlen_fail_when_not_all_readlengths_equal() { - let read_lengths = HashMap::from([(101, 5), (100, 1000), (99, 5)]); - let result = predict(read_lengths, 1010, 0.7).unwrap(); + let read_lengths = HashMap::from([ + ( + Arc::new("RG1".to_string()), + HashMap::from([(101, 5), (99, 5)]), + ), + (Arc::new("RG2".to_string()), HashMap::from([(100, 1000)])), + ]); + let result = predict(read_lengths, 0.7); assert!(!result.succeeded); assert_eq!(result.consensus_read_length, None); assert!(result.majority_pct_detected < 0.7); From cf3a8692fdf0f824c52556e794334b22a79136a4 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 25 Mar 2024 06:18:43 -0400 Subject: [PATCH 89/91] chore: removing dead code --- src/derive/strandedness/compute.rs | 4 ---- src/utils/args.rs | 13 +------------ 2 files changed, 1 insertion(+), 16 deletions(-) diff --git a/src/derive/strandedness/compute.rs b/src/derive/strandedness/compute.rs index 9be8f3c..19a64b4 100644 --- a/src/derive/strandedness/compute.rs +++ b/src/derive/strandedness/compute.rs @@ -469,10 +469,6 @@ mod tests { assert!(disqualify_gene(&record, &exons)); // disqualified } - #[test] - fn test_query_and_filter() { // TODO - } - #[test] fn test_classify_read() { // Set up diff --git a/src/utils/args.rs b/src/utils/args.rs index 8d5bedf..ac6f309 100644 --- a/src/utils/args.rs +++ b/src/utils/args.rs @@ -3,7 +3,7 @@ use std::fmt::Display; use std::num::NonZeroUsize; -use noodles::{bgzf::writer::CompressionLevel, sam::record::MappingQuality}; +use noodles::bgzf::writer::CompressionLevel; use tracing::debug; //===================// @@ -150,14 +150,3 @@ pub fn arg_in_range(arg: f64, range: std::ops::RangeInclusive) -> anyhow::R ), } } - -// TODO dead code, not used. Doesn't work as written. -/// Utility method to parse command line integers and ensure they are -/// within the range [0, 255) and return them as MappingQualities. -pub fn parse_min_mapq(s: &str) -> Result, std::num::ParseIntError> { - let value = s.parse()?; - match value { - 0..=254 => Ok(Some(MappingQuality::new(value).unwrap())), - 255 => Ok(None), - } -} From 7bbb7e5cbb1470a152a7d3d6e7463ce881b1d67f Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 25 Mar 2024 10:52:51 -0400 Subject: [PATCH 90/91] tests(derive/instrument): assert that read groups succeed --- src/derive/instrument/compute.rs | 6 ++++++ 1 file changed, 6 insertions(+) diff --git a/src/derive/instrument/compute.rs b/src/derive/instrument/compute.rs index 4256f96..173e11d 100644 --- a/src/derive/instrument/compute.rs +++ b/src/derive/instrument/compute.rs @@ -460,6 +460,9 @@ mod tests { "multiple instruments were detected in this file via the instrument id".to_string() ) ); + // We can't know which read group will be first in the vector. + // But both should succeed. + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); } #[test] @@ -491,6 +494,9 @@ mod tests { result.comment, Some("multiple instruments were detected in this file via the flowcell id".to_string()) ); + // We can't know which read group will be first in the vector. + // But both should succeed. + assert!(result.read_groups[0].succeeded && result.read_groups[1].succeeded); } #[test] From 6091af41ec52a988e8a4d3e3a83b0200b115b755 Mon Sep 17 00:00:00 2001 From: Andrew Frantz Date: Mon, 26 Aug 2024 17:07:28 -0400 Subject: [PATCH 91/91] fix(instrument): properly init flowcell entries --- src/derive/command/instrument.rs | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/src/derive/command/instrument.rs b/src/derive/command/instrument.rs index 5913694..4ef85ab 100644 --- a/src/derive/command/instrument.rs +++ b/src/derive/command/instrument.rs @@ -63,8 +63,10 @@ pub fn derive(args: DeriveInstrumentArgs) -> anyhow::Result<()> { .or_default() .insert(read.instrument_name); metrics.found_instrument_name += 1; + // Get or init the flowcell set for this read group. + let fc_entry = flowcell_names.entry(read_group).or_default(); if let Some(fc) = read.flowcell { - flowcell_names.entry(read_group).or_default().insert(fc); + fc_entry.insert(fc); metrics.found_flowcell_name += 1; } }