BactiSeq: Introduction

Bactiseq is a bacterial isolate whole genome sequence analysis pipeline. Bactiseq delivers analysis from Read quality analysis all the way through to gene comparision between samples. BactiSeq accepts reads from PacBio HiFi in the form of Fastq and BAM files, Nanopore reads, and Illumina reads. BactiSeq is capabale of Hybrid assembly or Long read/Short read only assembly.

Tools used depend on the input data

Pipeline overview

The pipeline is built using Nextflow and processes data using the following steps:

Bactiseq Pipeline Workflow: Processes and Modules per Entry Data Type

Input Data Type	Module	Software Tool(s)	Version
Pacbio HiFi	Assembly Module
	if SAM/BAM file Convert to Fastq	GATK4 SamToFastq	4.6.2.0
	1. Quality Assessment (Raw)	FastQC, Nanoplot, Seqkit stats	0.12.1, 1.46.1, 2.9.0
	2. Quality Control & Trimming	HiFiAdaptFilt, Chopper	3.0.0, 0.9.0
	3. Quality Assessment (Trimmed)	FastQC, Nanoplot, Seqkit stats	0.12.1, 1.46.1, 2.9.0
	4. Assembly	Flye	2.9.5
	5. Taxonomy Identification	Gambit, Kraken2	1.1.0, 2.1.5
	6. Polishing (Optional)	Pilon, Racon	1.24, 1.5.0
Nanopore reads	Assembly Module
	1. Quality Assessment (Raw)	FastQC, Nanoplot, Seqkit stats	0.12.1, 1.46.1, 2.9.0
	2. Quality Control & Trimming	HiFiAdaptFilt, Chopper	3.0.0, 0.9.0
	3. Quality Assessment (Trimmed)	FastQC, Nanoplot, Seqkit stats	0.12.1, 1.46.1, 2.9.0
	4. Assembly	Flye	2.9.5
	5. Taxonomy Identification	Gambit, Kraken2	1.1.0, 2.1.5
	6. Polishing (Optional)	Nextpolish, Medaka	1.4.1, 1.8.0
Illumina reads	Assembly Module
	1. Quality Assessment (Raw)	FastQC, Seqkit stats	0.12.1, 2.9.0
	2. Quality Control & Trimming	bbduk	39.18
	3. Quality Assessment (Trimmed)	FastQC, Seqkit stats	0.12.1, 2.9.0
	4. Assembly	Unicycler, Spades	0.5.1, 4.1.0
	5. Taxonomy Identification	Gambit, Kraken2	1.1.0, 2.1.5
	6. Polishing (Optional)	Pilon, Racon	1.24, 1.5.0
Assembled Genome	Conversion Module
	Convert file formats to FASTA	Any2fasta	0.4.2
All Data	Assembly Quality Assessment Module
	Quality assessment	CheckM2, BUSCO, QUAST	1.1.0, 5.8.3, 5.3.0
	Annotation Module
	1. Genome annotation	Bakta, Prokka	1.10.4, 1.14.6
	2. AMR gene detection	RGI, AMRFinderPlus	6.0.3, 3.12.8
	3. Virulence gene detection	Abricate	1.0.1
	4. Plasmid detection	Mobsuite	3.1.9
	5. MLST detection	tseemann/MLST	2.23.0
	Visualization Module
	1. Circular genome view	Cgview	2.0.3
	2. Assembly graph view	Bandage	0.9.0
	If SAM file, convert to BAM	SAMTOOLS	1.21
	3. Read alignment coverage	tinycov	0.4.0
	4. MLST distribution (pie chart)	Custom Python script	Python v3.12
	5. Heatmap of genes in common	Custom Python script	Biopython v1.81
	6. Plasmids and their lengths	Custom Python script	Pandas v2.0.0
	7. Number of reads and Average read quality	Custom Python script	Seaborn v0.12.0
	8. Binary heatmap of AMR/virulence genes	Custom Python scripts	Pandas v2.0.0
	Database Download Module
	1. Bakta database	bakta_db	1.10.4
	2. AMRFinderPlus database	amrfinder_update	3.12.8
	3. CheckM2 database	CheckM2	14897628
	4. RGI database	RGI	6.0.3
	5. Busco database	Busco	5.8.3 (bacteria_odb10)
	6. Kraken2 database	WGET	8gb standard Kraken2 database
	7. Gambit database	WGET	1.0

BactiSeq: Documentation

BactiSeq documentation is split into the following pages:

• Getting started
  • An overview on the pre-requisites, and how to install and use the pipeline
• BactiSeq Databases
  • An overview on the type of databases necessary for the pipeline to run
• Usage
  • An overview of how the pipeline works, how to run it and a description of all of the different command-line flags.
• Output
  • An overview of the different results produced by the pipeline and how to interpret them.

You can find a lot more documentation about installing, configuring and running nf-core pipelines on the website: https://nf-co.re