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README_for_DiscoVir_outputs.md

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README for DiscoVir Outputs

Thanks for using Nephele's DiscoVir virome pipeline! See our pipeline details page for more information, and for additional questions and help, please email us at [email protected] (include your jobID and log messages for help with errors).

Log file

  • logfile.txt: main log file for the pipeline. DiscoVir uses Snakemake for the workflow, and logfile.txt has the Snakemake output for each step in the pipeline. It will list any errors (look for "Error"), and direct you to the step's individual log file for more information.

Folders

There are two main folder types in the output of the DiscoVir pipeline:

  • Sample folders: per-sample output
  • Combined folders: outputs of analysis on sequences combined from across the entire dataset & tables merged from the individual sample results.

Here we will highlight the final output of each pipeline step as well as other important files. For a complete list of the outputs of each tool (including any intermediate files), please see the individual tool's documentation linked below.

Sample folders

Each sample has an individual folder which contains subfolders for each step in the pipeline that analyzes the individual sample's sequences.

The subfolders inside the sample folder are:

  • genomad: outputs of geNomad.

    • {sample}.genomad.log: log file to check for all (STDOUT and STDERR) messages from this pipeline step.
    • {sample}_summary: final output of genomad
      • {sample}_summary/{sample}_virus_summary.tsv: summary table of viral sequences identified
      • {sample}_summary/{sample}_virus.fna: FASTA of viral sequences (with host trimmed, if necessary)
      • {sample}_summary/{sample}_virus_genes.tsv: summary table of genes predicted from viral sequences
      • {sample}_summary/{sample}_virus_proteins.faa: amino acid/protein FASTA of viral genes
    • abund_genomad/{sample}_virus.count.CDS.cpm.txt: abundance estimates of the viral sequences.
      • read counts are estimated using the ht-seq algorithm from the tool VERSE , along with estimated copies per million (cpm), and reads per kilobase (rpk)
      • cpm is calculated the same way as tpm, but here we use as a generic term in case of DNA-seq.
    • abund_genomad/{sample}_virus_genes.count.CDS.cpm.txt: abundance estimates of viral genes

  • checkv: outputs of CheckV

    • {sample}.checkv.log: log file
    • quality_summary.tsv: summary of quality of all viral sequences
    • combined.fna: viral sequences identified by CheckV (with headers given by CheckV)
    • checkv_filtered_genomad_viruses.fna: viral sequences filtered based on the checkv quality user option. this file uses the original genomad headers which allows the pipeline to more easily track the sequences through analysis.

  • dramv: outputs of running DRAM-v annotate step only using the viral sequences predicted by geNomad and optionally filtered by CheckV ( checkv/checkv_filtered_genomad_viruses.fna). DRAM-v finds genes and annotates them.

    • {sample}.dramv.log: log file

    • dramv-annotate/annotations.tsv: table of DRAM-v gene annotations

    • dramv-annotate/genes.{faa,fna,gff}: AA and nucleotide FASTA files as well as associated gff file for the genes. the coordinates in the gff are vis-a-vis the original geNomad sequences found in checkv/checkv_filtered_genomad_viruses.fna.

    • abund_dramv/{sample}_dramv.count.gene.cpm.txt: abundance estimates of genes found by DRAM-V

  • amgs (optional): if the run_amgs user option is chosen, this folder has the DRAM-v predicted AMGs (auxiliary metabolic genes). To predict the AMGs, the pipeline first runs VirSorter2 on checkv_filtered_genomad_viruses.fna to generate the table needed by DRAM-v to predict AMGs. VirSorter2 filters out many sequences identified as viral by other tools, so this is an optional step.

    • vs2: output of VirSorter2
      • vs2/{sample}.vs2.log: log file
      • vs2/for-dramv/final-viral-combined-for-dramv.fna & vs2/for-dramv/viral-affi-contigs-for-dramv.tab: files used by DRAM-v to predict AMGs
    • {sample}.dramv_amgs.log: DRAM-v log file
    • dramv-annotate/: directory of DRAM-v gene annotations - using all DRAM-v databases
    • dramv-distill: directory containing the output of DRAM-v's distill step which identifies AMGs
      • dramv-distill/amg_summary.tsv: table of potential AMGs
    • abund_amgs/{sample}_amgs.count.gene.cpm.txt: abundance estimates of genes

  • diamond (optional): if the run_diamond option is chosen, this folder contains the output of annotating the geNomad-predicted genes by aligning sequences with diamond to NCBI's nr database.

    • {sample}.nr.diamond.tsv: table of top alignments for gene sequences with NCBI nr accession number. For full explanation of all columns see the NCBI BLAST format table under outfmt (DIAMOND uses the BLAST output format).

Combined folders

  • vOTUs:

    • vOTU_sequences.fasta: FASTA of final vOTU sequences.
    • vOTU_table_cpm.tsv: Matrix of abundances (CPM) of vOTUs for each sample.
    • vOTU.krona.html: Krona plots of vOTU taxonomy.
    • vOTU_genomad_virus_summary.tsv: geNomad summary information for vOTUs.
    • vOTU_cpm.biom: biom file of vOTU abundances and taxonomy
    • vOTU_clustering:
      • bbtools_dedupe: outputs of deduplication step with BBTools dedupe.sh
        • all_input_contigs.fasta: FASTA file containing all viral genomes combined from all samples.
        • unique_seqs.fasta: FASTA file containing all unique sequences (deduplicated sequences) from all samples.
        • bbtools_dedupe.log: log file to check for all (STDOUT and STDERR) messages from this pipeline step.
      • mmseqs: output of clustering deduplicated sequences with MMSeqs2
        • cluster_seqs.fasta: Viral genome FASTA sequences grouped by cluster.
        • DB_clu.tsv: Tab separated file displaying IDs of sequences within each cluster.
        • flat_DB_clu.tsv: Tab separated file displaying IDs of sequences within each cluster.
        • representative_sequences.fasta: FASTA sequences of viral genome representing each cluster, which becomes the vOTU. FASTA names include names of all viral genomes within each cluster.
        • representative_sequences.renamed.fasta: FASTA sequences of viral genome representing each cluster, which becomes the vOTU. FASTA names are changed so that they are only the representative sequence.
        • mmseqs2.log: log file
        • DB: Directory containing mmseqs2 outputs and indexes for database.

  • vOTU_Host_predictions_iphop (optional): if the run_iphop option is chosen, this will contain outputs of iPHoP

    • Host_prediction_to_genome_m##.csv: Files containing summary information of host predictions. Host predictions are made at the genome level.
    • Host_prediction_to_genus_m##.csv: Files containing summary information of host predictions. Host predictions are made at the genus level.
    • Host_cpm_table.csv: Matrix of abundances (CPM) of phage hosts of vOTUs for each sample predicted from iphop.
    • Host.krona.html: Krona plots of host taxonomy.

  • gene_tables:

    • dramv_kofam_hits_cpm.tsv: A matrix of abundances (CPM) of kofam hits from DRAM-v.
    • dramv_pfam_hits_cpm.tsv: A matrix of abundances (CPM) of Pfam hits from DRAM-v.
    • dramv_vogdb_hits_cpm.tsv: A matrix of abundances (CPM) of VOGID hits from DRAM-v.
    • dramv_vogdb_heatmap_cpm.pdf: Heatmap of abundances (CPM) of top (by prevalence and abundance) VOGID hits from DRAM-v.
    • amg_cpm.tsv: If the AMG option is selected, this file will be produced containing a matrix of CPM abundances of AMGs for each sample.
    • amg_heatmap_cpm.pdf: Heatmap of abundances (CPM) of AMGs.

Pipeline diagram

discovir pipeline diagram{ height=auto }\