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Marker Gene Search

marker-search identifies RNA virus marker genes (primarily RNA-dependent RNA polymerase) in contigs using profile HMMs.

flowchart TD
  subgraph IN["<b>Input</b>"]
  IN1["FASTA (Contigs or proteins - nuceic or proteins) <br>  (.fa/.fasta/.faa/.fna)/.fas"]
  end

  subgraph P["<b>Marker Search Pipeline</b>"]
  T["Translate / ORF prediction<br>  (six-frame / pyrodigal / bbmap)"]
  H["HMM Search<br> (pyhmmer / hmmsearch)"]
  R["Resolve / Filter Hits<br>  (merge/split/drop/none)"]
  end

  subgraph OUT["<b>Outputs</b>"]
    O1["marker_hits.tsv (tabular)"]
    O2["per-db outputs (if resolve=none)"]
    O3["resolved_hits.json/tsv"]
  end

  IN1 --> T --> H --> R --> O1
  R --> O2
  R --> O3

  classDef inputStyle fill:#f0f9ff,stroke:#0366d6,color:#03396c;
  classDef pipelineStyle fill:#fff7f0,stroke:#b85c00,color:#7a3b00;
  classDef outputStyle fill:#f0fff4,stroke:#0b8a3e,color:#0b6624;

  class IN1 inputStyle
  class T,H,R pipelineStyle
  class O1,O2,O3 outputStyle

Options

Common

  • -i, --input: Input fasta file (required)
  • -o, --output: Output directory (default: current_directory/marker_search_out)
  • -t, --threads: Number of threads (default: 1)
  • -M, --memory: Memory limit (default: "6g")
  • -g, --log-file: Path to log file (default: current_directory/marker_search_logfile.txt)
  • --keep-tmp: Keep temporary files (flag)
  • -db, --database: Database(s) to search (default: "NeoRdRp_v2.1,genomad")
  • Options include: NeoRdRp_v2.1, RdRp-scan, RVMT, Pfam_RTs_RdRp, genomad, all
  • You may also pass a custom path to an HMM file, HMM directory, or MSA directory
  • -ie, --inc-evalue: Maximum e-value (default: 0.05)
  • -s, --score: Minimum score (default: 20)
  • -am, --aa-method: Translation strategy (six_frame, pyrodigal, bbmap)
  • -td, -tempdir, --temp-dir: Temporary directory path

Hit Resolution (Not Fully Implemented)

  • -rm, --resolve-mode: Multiple profile match handling (default: "simple")
  • merge: Merge overlapping hits
  • one_per_range: One hit per range
  • one_per_query: One hit per query
  • split: Split overlapping domains
  • drop_contained: Drop contained hits
  • none: No overlap resolution
  • simple: Chain drop_contained with split
  • -mo, --min-overlap-positions: Minimum overlap positions (default: 10)

Citations

Tools

  • pyhmmer: Python HMMER bindings

  • Citation: https://doi.org/10.1093/bioinformatics/btad214

  • pyrodigal: Python Prodigal-GV bindings

  • Citation: https://doi.org/10.21105/joss.04296

  • BBMap: ORF prediction

  • Citation: https://sourceforge.net/projects/bbmap/files/BBMap_39.08.tar.gz

Databases

  • NeoRdRp_v2.1: RdRp profiles
  • Citation: https://doi.org/10.1264/jsme2.ME22001

  • GitHub: https://github.com/shoichisakaguchi/NeoRdRp

  • RdRp-scan: RdRp profiles

  • Citation: https://doi.org/10.1093/ve/veac082
  • GitHub: https://github.com/JustineCharon/RdRp-scan

  • Note: Incorporates PALMdb (https://doi.org/10.7717/peerj.14055)

  • RVMT: RNA Virus MetaTranscriptomes

  • Citation: https://doi.org/10.1016/j.cell.2022.08.023
  • GitHub: https://github.com/UriNeri/RVMT

  • Zenodo: https://zenodo.org/record/7368133

  • TSA_2018: Transcriptome Shotgun Assembly

  • Citation: https://doi.org/10.1093/molbev/msad060

  • Data: https://drive.google.com/drive/folders/1liPyP9Qt_qh0Y2MBvBPZQS6Jrh9X0gyZ

  • Pfam_A_37: Protein families

  • Citation: https://doi.org/10.1093/nar/gkaa913
  • Data: https://ftp.ebi.ac.uk/pub/databases/Pfam/releases/Pfam37.0/Pfam-A.hmm.gz

  • Note: RdRps and RT profiles: PF04197.17, PF04196.17, PF22212.1, PF22152.1, PF22260.1, PF05183.17, PF00680.25, PF00978.26, PF00998.28, PF02123.21, PF07925.16, PF00078.32, PF07727.19, PF13456.11

  • geNomad: Plasmid, virus, or host classification

  • Citation: https://doi.org/10.1038/s41587-023-01953-y
  • Data: https://doi.org/10.5281/zenodo.6994741
  • GitHub: https://github.com/apcamargo/genomad
  • Note: RNA virus marker proteins extracted from geNomad v1.9