bactopia_qc

Tags: fastq qc adapter-removal error-correction subsampling fastp bbduk lighter porechop nanoq fastqc nanoplot sample-scope

Automated quality control, error correction, and read subsampling.

A comprehensive QC pipeline that adapts to the input read type:

• Illumina: Adapter/PhiX removal (Fastp or BBDuk), Error Correction (Lighter), and Subsampling (Rasusa)
• Nanopore: Adapter removal (Porechop), Quality filtering (Nanoq), and Subsampling (Rasusa)
• Hybrid: Processes both short and long reads through their respective pipelines
• Assembly: Passes through simulated reads from assemblies

Generates quality metrics using fastq-scan and optional quality reports using FastQC (Illumina) and NanoPlot (ONT).

Inputs

record (
    meta: Record,
    r1: Path?,
    r2: Path?,
    se: Path?,
    lr: Path?,
    fna: Path?
)

Field	Type	Description
meta	Record	Groovy Record containing sample information (must include runtype, genome_size, species)
r1	Path?	Illumina R1 reads (paired-end forward)
r2	Path?	Illumina R2 reads (paired-end reverse)
se	Path?	Single-end Illumina reads
lr	Path?	Long reads (ONT)
fna	Path?	Assembly file (FASTA) for assembly-based simulations

adapters: Path?
phix: Path?

Name	Type	Description
adapters	Path?	Filepath for custom adapter sequences (FASTA)
phix	Path?	Filepath for custom PhiX sequences (FASTA)

Outputs

record (
    meta: Record,
    r1: Path?,
    r2: Path?,
    se: Path?,
    lr: Path?,
    fna: Path?,
    reads_grouped: Set<Path?>,
    error: Set<Path?>,
    skipped: Path?,
    results: Set<Path>,
    logs: Set<Path?>,
    nf_logs: Set<Path>,
    versions: Set<Path>
)

Field	Type	Description
meta	Record	Sample information record
r1	Path?	QC'd Illumina R1 reads (paired-end forward)
r2	Path?	QC'd Illumina R2 reads (paired-end reverse)
se	Path?	QC'd single-end Illumina reads
lr	Path?	QC'd long reads (ONT)
fna	Path?	Assembly file (FASTA)
reads_grouped	Set<Path?>	All output FASTQs for publishing
error	Set<Path?>	Captured error messages if QC failed (e.g., reads empty after trimming)
skipped	Path?	Marker file indicating QC was skipped for this sample
results	Set<Path>	All output files to be published
logs	Set<Path?>	Optional program specific log files
nf_logs	Set<Path>	Nextflow-specific log files (e.g. .command.{begin
versions	Set<Path>	A YAML formatted file with program versions

Parameters

QC Parameters

Parameter	Type	Default	Description
--use_bbmap	boolean		Illumina reads will be QC'd using BBMap
--use_porechop	boolean	false	Use Porechop to remove adapters from ONT reads
--skip_qc	boolean		The QC step will be skipped and it will be assumed the inputs sequences have already been QCed.
--skip_qc_plots	boolean		QC Plot creation by FastQC or Nanoplot will be skipped
--skip_error_correction	boolean		FLASH error correction of reads will be skipped.
--adapters	string		A FASTA file containing adapters to remove
--adapter_k	integer	23	Kmer length used for finding adapters.
--phix	string		phiX174 reference genome to remove
--phix_k	integer	31	Kmer length used for finding phiX174.
--ktrim	string	r	Trim reads to remove bases matching reference kmers (choices: f, r, l)
--mink	integer	11	Look for shorter kmers at read tips down to this length, when k-trimming or masking.
--hdist	integer	1	Maximum Hamming distance for ref kmers (subs only)
--tpe	string	t	When kmer right-trimming, trim both reads to the minimum length of either (choices: f, t)
--tbo	string	t	Trim adapters based on where paired reads overlap (choices: f, t)
--qtrim	string	rl	Trim read ends to remove bases with quality below trimq. (choices: rl, f, r, l, w)
--trimq	integer	6	Regions with average quality BELOW this will be trimmed if qtrim is set to something other than f
--maq	integer	10	Reads with average quality (after trimming) below this will be discarded
--minlength	integer	35	Reads shorter than this after trimming will be discarded
--ftm	integer	5	If positive, right-trim length to be equal to zero, modulo this number
--tossjunk	string	t	Discard reads with invalid characters as bases (choices: f, t)
--ain	string	f	When detecting pair names, allow identical names (choices: f, t)
--qout	string	33	PHRED offset to use for output FASTQs (choices: 33, 64)
--maxcor	integer	1	Max number of corrections within a 20bp window
--sampleseed	integer	42	Set to a positive number to use as the random number generator seed for sampling
--ont_minlength	integer	1000	ONT Reads shorter than this will be discarded
--ont_minqual	integer	0	Minimum average read quality filter of ONT reads
--porechop_opts	string		Extra Porechop options in quotes
--nanoplot_opts	string		Extra NanoPlot options in quotes
--bbduk_opts	string		Extra BBDuk options in quotes
--fastp_opts	string		Extra fastp options in quotes

Used By

Subworkflows

• bactopia_qc - Perform comprehensive quality control on sequencing reads.

Workflows

• bactopia - Comprehensive bacterial analysis pipeline for complete genomic characterization.
• cleanyerreads - Quality control and optional host read removal from raw sequencing reads.
• staphopia - Comprehensive analysis pipeline for Staphylococcus aureus isolates.

Citations

If you use this in your analysis, please cite the following.

• Bactopia
  Petit III RA, Read TD Bactopia - a flexible pipeline for complete analysis of bacterial genomes. mSystems 5 (2020)

• BBTools
  Bushnell B BBMap short read aligner, and other bioinformatic tools. (Link)

• fastp
  Chen S, Zhou Y, Chen Y, and Gu J fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics, 34(17), i884-i890. (2018)

• FastQC
  Andrews S FastQC: a quality control tool for high throughput sequence data. (WebLink)

• fastq-scan
  Petit III RA fastq-scan: generate summary statistics of input FASTQ sequences. (GitHub)

• Lighter
  Song L, Florea L, Langmead B Lighter: Fast and Memory-efficient Sequencing Error Correction without Counting. Genome Biol. 15(11):509 (2014)

• NanoPlot
  De Coster W, D'Hert S, Schultz DT, Cruts M, Van Broeckhoven C NanoPack: visualizing and processing long-read sequencing data Bioinformatics Volume 34, Issue 15 (2018)

• Nanoq
  Steinig E Nanoq: Minimal but speedy quality control for nanopore reads in Rust (GitHub)

• Porechop
  Wick RR, Judd LM, Gorrie CL, Holt KE. Completing bacterial genome assemblies with multiplex MinION sequencing. Microb Genom. 3(10):e000132 (2017)

• Rasusa
  Hall MB Rasusa: Randomly subsample sequencing reads to a specified coverage. (2019).

Source

View source on GitHub

Version

BACTOPIA_QC:
    - bactopia-qc: 1.0.4