Spriggan

Spriggan is a NextFlow pipeline used for assembly of bacterial whole genome sequence data and identification of antibiotic resistance genes.

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 spriggan/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/spriggan -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 or -profile test_full:

nextflow spriggan/main.nf --outdir [path-to-outdir] -profile test[_full],[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

Antimicrobial resistance gene detection

Antimicrobial resistance genes, as well as point mutations, are identified using AMRFinderPlus v3.10.30. Spriggan can generate a table of AMRFinderPlus results for AR genes of interest with the selected_genes parameter. Spriggan will search for matches to the AR genes of interest in the AMRFinderPlus results and make a table called 'selected_ar_genes.tsv.' The list of genes must be separated by | and enclosed in single quotes in the config file. By default the selected_genes parameter is set to: 'NDM|OXA|KPC|IMP|VIM'

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.

Contamination detection

Contamination is detected by classifying reads using Kraken2 v2.0.8 with the Minikraken2_v1_8GB database.

Summary

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

Output files

Example of pipeline output:

spriggan_results
├── amrfinder
│   ├── *.amr.tsv
│   ├── *.fa
│   ├── amrfinder_predictions.tsv
│   ├── amrfinder_summary.tsv
│   └── selected_ar_genes.tsv
├── bbduk
│   ├── *.fastq.gz
│   ├── *.adapter.stats.txt
│   ├── *.bbduk.log
│   ├── *.trim.txt
│   └── bbduk_results.tsv
├── coverage
│   └── coverage_stats.tsv
├── fastqc
│   ├── *.html
│   ├── *.zip
│   └── fastqc_summary.tsv
├── kraken
│   ├── *.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
│   └── spriggan_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
│   └── spriggan_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:
spriggan_report.csv - Summary table of each step in Spriggan
spriggan_multiqc_report.html - HTML report generated by MultiQC
*.contigs.fa - Shovill assembly for each sample
*.amr.tsv - AMR genes identified in each sample by AMRFinderPlus
*.mlst.tsv - MLST scheme identified for each sample

Authors

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

cjjossart/spriggan