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Viral metagenomics analyses

Snakevir is a workflow developed in snakemake that enables viral metagenomic analysis of short-read data. You can give your own parameters/database in the config.yaml file to customize the workflow for your own data.

Warning

For the moment, snakevir only searches and analyzes viral data.

Read processing

First, snakevir will process reads. Library adapters and low-quality sequences in reads were removed with Cutadapt. Reads from rRNA sequences from diptera and bacteria were then removed through mapping with BWA against SILVA rRNA bases (bacterial databases: SSURefNr99 and LSURef (release 138), dipteran database, (release 138))

Included tools :

  • Cutadapt version == 3.4

  • Flash version == 1.2.11

  • BWA version == 0.7.17

  • samtools version == 1.12

  • picard version == 2.25.4

Assembly

The remaining reads were de-novo assembled with Megahit version. The resulting contigs and non-assembled reads were used in a second de-novo assembly with CAP3.

Included tools :

  • megahit version == 1.2.9

  • cap3 version == 10.2011

Virus identification

A contig was considered a virus-like sequence if a viral sequence was found as the first best hit by Diamond after comparison with the NCBI nr database (e-value cutoff = 10-3). Then, virus-like contigs were screened for sequences potentially derived from endogenous viral elements (EVEs) using BLASTn against NCBI nt (e-value cutoff = 10-3). Contigs with a non-viral best hit with a coverage above 25% were considered as potential EVEs and removed.

Included tools :

  • blast version == 2.11.0

  • diamond version == 2.0.9

Warning

  • For the moment, this workflow doesn’t work with diamond version >= 2.1.0 because the taxonmap option change (need to use this option on makeblastdb command).

Counting table

SnakeVir quantified the number of reads per virus-like contig and library through mapping with BWA. Duplicate reads had been previously removed with the markdup tool in Samtools. Contigs sharing the same best hit were grouped into a viral operational taxonomic unit (vOTU) as previously described.

Warning

ADD a description for coverage 10x

Included tools :

  • BWA version == 0.7.17

  • samtools version == 1.12

  • picard version == 2.25.4

Note

The grouping of viral contigs into a viral operational taxonomic unit (vOTU), as described above, is currently being added.