EBI-Metagenomics/envident

main @ 269afbd

Introduction

EBI-Metagenomics/envident EBI-Metagenomic's eDNA analysis pipeline. This pipeline is designed for the analysis of environmental DNA (eDNA) sequencing data, implementing a comprehensive workflow for quality control, primer identification, Amplicon Sequence Variant (ASV) calling and taxonomic profiling using modern bioinformatics tools. Currently the pipeline supports analysis of COI metabarcoding reads.

Default steps in EnvIdent

Quality Control and Preprocessing:

• Raw reads quality assessment using FastQC
• Reads quality control and filtering using fastp
• Minimum read count filtering (configurable threshold)

Primer Analysis:

• Automatic primer identification using PIMENTO
• Primer trimming using Cutadapt
• Primer validation and reporting

Taxonomic Profiling:

• Pfam-based COI (Cytochrome C Oxidase subunit I) profiling using HMMER
• Reads percentage threshold filtering for marker gene identification (configurable threshold)

ASV Analysis:

• Amplicon Sequence Variant (ASV) calling using DADA2
• ASV taxonomic classification using MAPseq
• Krona chart visualization for taxonomic results

Reporting and Quality Control:

• Comprehensive MultiQC reports
• Failed and passed runs tracking
• Software version reporting

Usage

[!NOTE] If you are new to Nextflow and nf-core, please refer to this page on how to set-up Nextflow. Make sure to test your setup with -profile test before running the workflow on actual data.

Requirements

The pipeline requires:

• Nextflow (≥24.04.2)
• Docker, Singularity, or Conda for software management
• Access to reference databases
• Primer database formatted for PIMENTO - a FASTA file with contig ids ending with F for forward primers and R for reverse primers. See here for an example

Input Format

The input data should be eDNA sequencing reads (paired-end or single-end) in FASTQ format, specified using a CSV samplesheet:

sample,fastq_1,fastq_2,single_end
sample1,/path/to/sample1_R1.fastq.gz,/path/to/sample1_R2.fastq.gz,false
sample2,/path/to/sample2.fastq.gz,,true

Basic execution

nextflow run EBI-Metagenomics/envident \
    -r main \
    -profile example_slurm \
    --input samplesheet.csv \
    --outdir results

Pipeline output

Example output structure for a sample (sample1). The qc_passed and qc_failed csvs are only present if you have samples that passed or failed:

results/
├── sample1/
│   ├── asv/
│   │   ├── sample1_DADA2-BOLD_asv_read_counts.tsv
│   │   ├── sample1_DADA2-MIDORI_asv_read_counts.tsv
│   │   └── sample1_dada2_stats.tsv
│   │   └── sample1_asvs.fasta
│   ├── hmmsearch-COI/
│   │   ├── sample1_Pfam-A.domtbl
│   │   └── sample1_Pfam-A.txt
│   ├── primer-identification/
│   │   └── sample1.cutadapt.json
│   ├── qc/
│   │   ├── sample1_seqfu.tsv
│   │   └── sample1.fastp.json
│   │   └── sample1.merged.fastq.gz
│   │   └── sample1_suffix_header_err.json
│   ├── taxonomy-summary/
│   │   ├── DADA2-BOLD/
│   │   |   ├── ERR8441464_DADA2-BOLD_asv_krona_counts.txt
│   │   |   ├── ERR8441464_DADA2-BOLD_asv_taxa.tsv
│   │   |   ├── ERR8441464_DADA2-BOLD.html
│   │   |   └── ERR8441464_DADA2-BOLD.mseq
│   │   ├── DADA2-MIDORI/
│   │   |   ├── ERR8441464_DADA2-MIDORI_asv_krona_counts.txt
│   │   |   ├── ERR8441464_DADA2-MIDORI_asv_taxa.tsv
│   │   |   ├── ERR8441464_DADA2-MIDORI.html
│   │   |   └── ERR8441464_DADA2-MIDORI.mseq
├── pipeline_info/
│   ├── execution_report_YYYY-MM-DD_HH-mm-ss.html
│   ├── execution_timeline_YYYY-MM-DD_HH-mm-ss.html
│   ├── execution_trace_YYYY-MM-DD_HH-mm-ss.txt
│   ├── params_YYYY-MM-DD_HH-mm-ss.json
│   ├── pipeline_dag_YYYY-MM-DD_HH-mm-ss.html
│   └── envident_software_mqc_versions.yml
├── multiqc_report.html
├── qc_passed_runs.csv
└── qc_failed_runs.csv

Credits

EBI-Metagenomics/envident was written by Christina Vasilopoulou and Jennifer Mattock.

Citations

This pipeline uses code developed and maintained by the nf-core community, reused here under the MIT license.

The nf-core framework for community-curated bioinformatics pipelines.

Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia, Paolo Di Tommaso & Sven Nahnsen.

Nat Biotechnol. 2020 Feb 13. doi: 10.1038/s41587-020-0439-x.

Please cite this pipeline using the following DOI: 10.48546/workflowhub.workflow.2177.1