tbQC

tbQC is a NextFlow pipeline used for assembly of bacterial whole genome sequence data and identification of species and MLST scheme.

Table of Contents:

Usage
Workflow outline
Read trimming and quality assessment
Genome assembly
Assembly quality assessment
Genome coverage
MLST scheme
Species determination
Summary
Output files

Usage

The pipeline is designed to start from raw Illumina reads. All reads must be in the same directory. Then start the pipeline using:

nextflow tbQC/main.nf --input [path-to-samplesheet] --outdir [path-to-outdir] -profile [docker,singularity,aws]

You can specify a version of the pipeline and run it directly from the github repository by using:

nextflow wslh-bio/tbQC -r [version] --input [path-to-samplesheet] --outdir [path-to-outdir] -profile [docker,singularity,aws]

You can also test the pipeline with example data using -profile test:

nextflow tbQC/main.nf --outdir [path-to-outdir] -profile test,[docker/singularity]

Workflow outline

Read trimming and quality assessment

Read trimming and cleaning is performed using BBtools v38.76 to trim reads of low quality bases and remove PhiX contamination. Then FastQC v0.11.8 is used assess the quality of the raw and cleaned reads.

Genome assembly

Assembly of the cleaned and trimmed reads is performed using Shovill v1.1.0.

Assembly quality assessment

Quality assessment of the assemblies is performed using QUAST v5.0.2

Genome coverage

Mean and median genome coverage is determined by mapping the cleaned reads back their the assembly using BWA v0.7.17-r1188 and calculating depth using samtools v1.10

MLST scheme

MLST scheme is classified using MLST v2.17.6. Multiple schemes are available for specific organisms, and STs from all available schemes are reported for those organisms.

Species determination

Species is determined by classifying reads using Kraken2 v2.0.8 and Mash.

Summary

Results are summarized using MultiQC v1.11 and Pandas v1.3.2.

Output files

Example of pipeline output:

tbqc_results
├── bbduk
│   ├── *.fastq.gz
│   ├── *.adapter.stats.txt
│   ├── *.bbduk.log
│   ├── *.trim.txt
│   └── bbduk_results.tsv
├── coverage
│   └── coverage_stats.tsv
├── fastqc
│   ├── *.html
│   ├── *.zip
│   └── fastqc_summary.tsv
├── kraken
│   ├── *.mash.tsv
│   └── mash_results.tsv
├── mash
│   ├── *.kraken2.txt
│   ├── kraken_results.tsv
│   └── kraken2.log
├── mlst
│   ├── *.alleles.tsv
│   ├── *.mlst.tsv
│   └── mlst_results.tsv
├── multiqc
│   ├── multiqc_data
│   │   ├── *.json
│   │   ├── *.txt
│   │   └── multiqc.log
│   ├── multiqc_plots
│   │   ├── pdf
│   │   │   └── *.pdf
│   │   ├── png
│   │   │   └── *.png
│   │   └── svg
│   │       └── *.svg
│   └── tbqc_multiqc_report.html
├── pipeline_info
│   ├── *.html
│   ├── *.txt
│   ├── samplesheet.valid.csv
│   └── software_versions.yml
├── quast
│   ├── *.quast.report.tsv
│   ├── *.transposed.quast.report.tsv
│   └── quast_results.tsv
├── results
│   └── tbqc_report.csv
├── samtools
│   ├── *.bam
│   ├── *.depth.tsv
│   └── *.stats.txt
└── shovill
    ├── *.contigs.fa
    ├── *.sam
    └── shovill_output
          ├── contigs.gfa
          ├── shovill.corrections
          ├── shovill.log
          └── spades.fasta

Notable result files:
tbqc_report.csv - Summary table of each step in tbqc
tbQC_multiqc_report.html - HTML report generated by MultiQC

Warning: Please provide pipeline parameters via the CLI or Nextflow -params-file option. Custom config files including those provided by the -c Nextflow option can be used to provide any configuration except for parameters; see docs.

Authors

Kelsey Florek, WSLH Senior Genomics and Data Scientist
Abigail Shockey, WSLH Bioinformatician and Data Scientist

Contributions and Support

If you would like to contribute to this pipeline, please see the contributing guidelines.

This pipeline uses code and infrastructure developed and maintained by the nf-core community.

The nf-core framework for community-curated bioinformatics pipelines.

Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia, Paolo Di Tommaso & Sven Nahnsen.

Nat Biotechnol. 2020 Feb 13. doi: 10.1038/s41587-020-0439-x.