ResMAG - name pending

ResMAG is a state-of-the-art and user-friendly Snakemake workflow designed for the analysis of metagenomic data. It integrates multiple bioinformatics tools and algorithms to facilitate key steps in metagenome analysis, including bin refinement, metagenome-assembled genome (MAG) reconstruction, taxonomic classification of MAGs, and identification of antibiotic resistance genes.

Key Features

Binning Techniques: Employ a collection of five state-of-the-art binning tools to partition metagenomic contigs into individual bins, allowing for comprehensive and accurate analysis.

MAG Reconstruction: Utilize cutting-edge algorithms to reconstruct high-quality metagenome-assembled genomes (MAGs) from sequencing data.

Taxonomic Classification: Apply advanced taxonomic classification methods to assign taxonomic labels to MAGs and identify the microbial community composition within the metagenomic samples.

Antibiotic Resistance Gene Identification: Perform in-depth analysis to detect and characterize antibiotic resistance genes within the metagenomic data, providing valuable insights into antimicrobial resistance profiles.

Performance Refinement: Continuously optimize the pipeline by incorporating the latest advancements in metagenomics research, ensuring the highest accuracy and efficiency in metagenomic data analysis.

Overview

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   'theme':'base',
   'themeVariables': {
      'secondaryColor': '#fff',
      'tertiaryColor': '#fff',
      'tertiaryBorderColor' : '#fff'}
   }}%%

flowchart TB;

   subgraph " "
      direction TB

      %% Nodes
      A[/short reads/]
      B["<b>QC</b> <br> <i>fastp and fastQC<i>"]
      C["<b>Filtering human reads</b> <br> <i>minimap2<i>"]
      L["optional <br> <b>Filtering host reads</b> <br> <i>minimap2<i>"]
      N["<b>taxonomic classification</b> <br> <i>Kaiju<i>"]
      D["<b>Assembly</b> <br> <i>MegaHIT</i>"]
      E["<b>Binning</b>  <br> <i>MetaBAT and MetaCoAG<i>"]
      F["<b>Bin refinement</b> <br> <i>DAS Tool<i>"]
      G["<b>Bin quality control</b> <br> <i>CheckM2<i>"]
      M[/MAGs/]
      H["<b>Resistance analysis</b> <br> <i>CARD<i>"]
      I["<b>taxonomic classification</b> <br> <i>GTDBTk<i>"]
      J[/MultiQC report/]
      K[/Assembly summary/]

      %% input & output node design
      classDef in_output fill:#fff,stroke:#cde498,stroke-width:4px
      class A,J,K,M in_output
      %% rule node design
      classDef rule fill:#cde498,stroke:#000
      class B,C,D,E,F,G,H,I,L,N rule

      %% Node links
      A --> B
      B --> C
      B --- J
      C --> L
      L --> D
      C --> N
      L --> N
      C --> D
      D --> E
      D --- K
      E --> F
      F --> G
      G --"filtering"--> M
      M --- H
      M --- I

   end

Loading

Usage

Preparations

Clone workflow

To prepare the workflow clone it to your desired working folder via git or your preferred IDE

git clone https://github.com/IKIM-Essen/metagenomics_res.git

Download GTDB

The GTDB needs to be downloaded and decompressed, it requires about 110 Gb.

1. Change to the cloned workflow directory
2. Create a new folder resources/gtdb/ and change to this directory
3. Download the latest version of GTDB or if you have already downloaded a version of GTDB move the gtdbtk_data.tar.gz file to resources/gtdb/
4. Unarchive the downloaded file
5. After successful step 4: the archive can be removed

wget https://data.ace.uq.edu.au/public/gtdb/data/releases/latest/auxillary_files/gtdbtk_package/full_package/gtdbtk_data.tar.gz
tar xvzf gtdbtk_data.tar.gz

Install Snakemake

Create a snakemake environment using mamba via:

mamba create -c conda-forge -c bioconda -n snakemake snakemake=7.32.3

For installation details, see the instructions in the Snakemake documentation.

Configuring workflow

1. Edit the config/config.yaml file:
   • Specify a project name (project-name)
   • Specify filtering options for human reads (human-filtering)
   • Specify host filtering options, if you have a non-human host (host-filtering)
   • Specify options for GTDB database (see Download GTDB)
2. Provide sample information in the config/pep/samples.csv file while keeping the header and the format:

sample_name,fq1,fq2
sample1,path/to/your/fastq/sample1_R1.fastq.gz,path/to/your/fastq/sample1_R2.fastq.gz

Run the workflow

Activate the conda environment: conda activate snakemake

Test your configuration by performing a dry-run via snakemake --use-conda -n

Executing the workflow: snakemake --use-conda --cores $N --rerun-incomplete

using $N cores. It is recommended to use all available cores.

The usage of this workflow is described in the Snakemake Workflow Catalog.

Output

Tools

A list of tools used in the pipeline:

Tool	Link
CheckM2	https://doi.org/10.1038/s41592-023-01940-w
CoverM	https://github.com/wwood/CoverM
DAS Tool	https://doi.org/10.1038/s41564-018-0171-1
fastp	https://doi.org/10.1093/bioinformatics/bty560
FastQC	www.bioinformatics.babraham.ac.uk/projects/fastqc
Kaiju	https://doi.org/10.1038/ncomms11257
MEGAHIT	https://doi.org/10.1093/bioinformatics/btv033
MetaBAT	http://dx.doi.org/10.7717/peerj.1165
MetaCoAG	https://doi.org/10.1101/2021.09.10.459728
minimap2	https://doi.org/10.1093/bioinformatics/bty191
MultiQC	www.doi.org/10.1093/bioinformatics/btw354
pandas	pandas.pydata.org
samtools	https://doi.org/10.1093/gigascience/giab008
Snakemake	www.doi.org/10.12688/f1000research.29032.1

Contact Information

For any questions, or feedback, please contact the project maintainer at [email protected]. We appreciate your input and support in using and improving ResMAG.

Acknowledgements

We would like to express our gratitude towards Katharina Block, Adrian Doerr, Miriam Balzer, Alexander Thomas, Johannes Köster, Ann-Kathrin Doerr and the IKIM who have contributed to the development and testing of ResMAG. Their valuable insights and feedback have been helpful throughout the creation of the workflow.

Citation

A paper is on its way. If you use ResMAG in your work before the paper, then please consider citing this GitHub.

License

ResMAG is released under the BSD-2 Clause. Please review the license file for more details.