Json producing Assembly driven microbial Sequence analysis pipeline to support Epitypification and Normalize classification decisions

JASEN produces results for epidemiological and surveillance purposes. JASEN has been tested using MRSA, but should work well with any bacteria with a stable cgMLST scheme.

Requirements

• Singularity
• Nextflow (curl -s https://get.nextflow.io | bash)

Recommended

• Conda
• Singularity Remote Login
• Mamba. Is available from conda. Install in base environment.

Development deployment (self-contained)

• git clone --recurse-submodules --single-branch --branch master https://github.com/genomic-medicine-sweden/JASEN.git && cd JASEN # Copies code locally
• bash -i deploy/deploy_conda.sh # Creates development environment
• bash -i deploy/deploy_references.sh # Downloads self-test databases
• (Optional) singularity remote login # Access to OCI regestries
• cd container && sudo bash -i build_container.sh && cd .. # Creates singularity images
• Edit the root parameter in configs/nextflow.base.config
• Edit the path to the test files in assets/test_data/samplelist.csv

Usage

Simple self-test

./nextflow run main.nf -entry bacterial_default -profile staphylococcus_aureus -config configs/nextflow.base.config --csv=assets/test_data/samplelist.csv

Start a new analysis with samples defined in assets/test_data/samplelist.csv using the staphylococcus_aureus profile. If nextflow has been added to the PATH, one should start the command with nextflow instead of ./nextflow.

Input files are defined in a csv file with the following format. All samples need to be of the same "type", meaning that they can be analyzed with the same analysis profile, defined in the nextflow config.

id,read1,read2
p1,assets/test_data/sequencing_data/saureus_10k/saureus_large_R1_001.fastq.gz,assets/test_data/sequencing_data/saureus_10k/saureus_large_R2_001.fastq.gz

Component Breakdown

QC

Species detection is performed using Kraken2 together with Bracken. The database used is a standard Kraken database built with

kraken2-build --standard --db $DBNAME

Low levels of Intra-species contamination or erronous mapping is removed using bwa and filtering away the heterozygous mapped bases.

Genome coverage is estimated by mapping with bwa mem and using a bed file containing the cgMLST loci.

A value on the evenness of coverage is calculated as an interquartile range.

Epidemiological typing

For de novo assembly SPAdes is used. QUAST is used for extracting QC data from the assembly.

The cgMLST reference scheme used, is branched off cgmlst.net At the moment this fork is not synced back with new allele numbers. For extracting alleles chewBBACA is used. Number of missing loci is calculated and used as a QC parameter.

Traditional 7-locus MLST is calculated using mlst.

Virulence and resistance markers

ARIBA is used as the tool to detect genetic markes. The database for virulence markes is VFDB.

Report and visualisation

The QC data is aggregated in a web service CDM (repo coming) and the cgMLST is visualized using a web service cgviz that is combined with graptetree for manipulating trees (repo coming).

Tips

It is recommended that you use latest versions of software tools, however if you are running an older version of Singularity and you get an error FATAL: could not open image JASEN/container/*.sif: image format not recognized! check the permissions set on image *.sif. Make sure you have the permission to execute it.