Annotate your own data

One assembly

metapathways run \
  -i sample.fasta \
  -o results \
  -d /data/MPDB

The sample output is results/sample/; the run-wide report is results/reports/. Input can be nucleotide FASTA, compressed FASTA, or amino-acid FASTA with --input_format fasta-amino. GFF and GenBank are output formats, not accepted primary inputs of this CLI.

Choose references and inspect the plan:

metapathways run -i sample.fasta -o results -d /data/MPDB \
  --annotation_dbs swissprot cazy --annotation_algorithm FAST --dryrun

A dry run checks dependencies and writes the execution plan and logs without executing annotation tasks. It can create sample directory scaffolding; it does not produce a completed biological report.

Read mapping

Paired reads in separate files:

metapathways run -i sample.fasta -o results -d /data/MPDB \
  -1 sample_R1.fastq.gz -2 sample_R2.fastq.gz

Interleaved paired reads:

metapathways run -i sample.fasta -o results -d /data/MPDB \
  -1 sample.interleaved.fastq.gz --interleaved

Single-end reads use -1 alone. With no reads, abundance calculation is skipped. -1 and -2 must identify different files; MP now rejects identical paired input paths, including aliases to the same file. The paired command passes the actual reverse file to CoverM.

Historical output generated by the incorrect mate selection must not be treated as corrected merely because it appears in a report. Repeating the annotation command with the corrected reads can reuse completed annotation stages and recompute mapping. The portal copies existing abundance values; it does not certify their provenance or rerun mapping.

Interpreting feature and hit counts

Protein search statistics distinguish alignment rows from distinct query ORFs; multiple hits for one ORF count once in the latter. ORF identifiers are counted in full, without assuming a particular naming format. Counts from different reference databases can overlap.

rRNA prediction and rRNA taxonomic annotation are separate outputs. MP uses barrnap’s GFF annotations to identify the subunit when extracted FASTA headers contain only coordinates, and retains support for older headers containing the subunit name. Alignment lengths include both endpoints and are independent of alignment direction. A predicted rRNA feature need not have a qualifying taxonomic hit.

Multiple assemblies

Pass a directory of FASTA files to -i; each file becomes a sample with its own output directory. Use unique, simple filenames and --samples to select a subset. Read mapping arguments apply to one sample only: run separate commands for samples with different FASTQs. CPU/memory budgets are per invocation, so independent MP commands do not share a global resource limit.

Stage controls

Stage flags accept yes, skip or redo. For example:

metapathways run -i sample.fasta -o results -d /data/MPDB \
  --COMPUTE_TPM redo -1 sample_R1.fastq.gz -2 sample_R2.fastq.gz

yes reuses valid outputs, redo executes the stage again, and skip requires any needed downstream inputs to already exist. --force_redo forces all annotation stages. Changes to tracked inputs or outputs invalidate task receipts. See the workflow guide before skipping dependencies.