Campylobacter CRISPRscape

Automated bioinformatics workflow for the identification and characterisation of Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) arrays from bacterial genomes, with a focus on Campylobacter coli and C. jejuni.

For more detailed documentation, please look at the project website. Here you also find the user manual, which includes a quick start guide as well as a detailed step-by-step description. Or look at the computer-generated Wiki with integrated chatbot assistant at DeepWiki!

Index

1. Roadmap
2. Workflow description
3. Project (file) organisation
4. Licence
5. Citation

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Release roadmap

• Future additions:
  • Extract orphan CRISPR arrays from CCTyper -> send to CRISPRidentify
  • CRISPR spacer target prediction
    • map to
      • masked ATB genomes (KMA)
      • PLSDB (SpacePHARER)
      • PhageScope (SpacePHARER)
      • VIRE (t.b.d.)
      • MEGAISurv metagenomes (t.b.d)
    • mini-benchmark different mapping algorithms?
      • Sassy
      • KMA
      • SpacePHARER
    • (where feasible) connect spacer hits with functional annotations
      • Bakta annotations from ATB are available!
  • Integrate downstream analyses with Snakemake?
    • run RMarkdown/Quarto notebooks automatically
  • Build a database like this spacerdb?
  • Create a command-line tool with snaketool
  • Separate 'main' workflow steps from 'optional/extra' steps?

Workflow description

The analysis is recorded as a Snakemake workflow. Its dependencies (bioinformatics tools) are handled by Snakemake using the conda package manager, or rather its successor mamba. If you have not yet done so, please install mamba following the instructions found here: https://mamba.readthedocs.io/en/latest/installation/mamba-installation.html.

After installing mamba, snakemake can be installed using their instructions: https://snakemake.readthedocs.io/en/stable/getting_started/installation.html#full-installation (Note: the workflow was tested with Snakemake version 8.20.3 and is expected to work with any version since 5.)

When Snakemake has been set up, you can test if the workflow is ready to be run (dry-run) with:

snakemake --profile config -n

If that returns no errors, run the workflow by removing the -n (dry-run) option:

snakemake --profile config

Note that the workflow is currently configured to run on the local machine (not on a high-performance computing (HPC) cluster or grid) and uses a maximum of 60 CPU threads. The number of threads to use can be configured in: config/config.yaml (overall workflow) and config/parameters.yaml (per step/tool).

In its current state, the workflow:

1. Determines MultiLocus Sequence Types for Campylobacter jejuni/coli using the public typing scheme from pubMLST and pyMLST (version 2.2.1)

2. Identifies CRISPR-Cas loci with CCTyper (version 1.8.0) and resulting loci are processed with CRISPRidentify (forked from version 1.2.1) to reduce false positives.

   • this includes extra scripts to collect CRISRP-Cas information and extract sequences from the genome fasta files

3. Collects all CRISPR spacers and creates clusters of identical spacers using CD-HIT-EST (version 4.8.1)

4. Predicts whether contigs of the species of interest derive from chromosomal DNA, plasmids or viruses using both geNomad (version 1.8.0) and Jaeger (version 1.1.26).

5. Predicts the potential targets of spacers and whether they target chromosomal DNA (of input genomes), plasmid or viruses using Spacepharer (version 5.c2e680a) and kma (version 1.5.0).

Further steps are added to the workflow after testing!

Preparing input genomes

Before running the workflow, the user needs to prepare input genomes. These may be downloaded from AllTheBacteria using a convenient script included with CRISPRscape:

• Prepare genomes

Project organisation

.
├── .gitignore
├── CITATION.cff
├── LICENSE
├── README.md
├── bin                <- Code and programs used in this projec
├── config             <- Configuration of Snakemake workflow
├── doc                <- Project documentation, notes and experiment records
├── log                <- Log files from programs
├── resources          <- Folder for databases and input files
├── results            <- Workflow output, all results
└── workflow           <- Files describing the Snakemake workflow
    ├── Snakefile      <- Main workflow description file (Python-based)
    ├── envs           <- Software environments (conda) for dependencies
    ├── notebooks      <- Analysis notebooks for downstream statistics/visualisation
    ├── rules          <- Workflow modules with processing steps
    └── scripts        <- Scripts used in the workflow

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Licence

This project is licensed under the terms of the New BSD licence.

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Citation

Please cite this project as described in the citation file.