S

    galah process

    Runs both analyse and cluster in sequence.

    Determines the MIMAG quality score based on completeness, contamination, rRNA, and tRNA presence. Completeness and contamination are estimated using CheckM2 by default, unless CheckM1/2 quality reports are provided.

    Cluster genomes into ANI-based groups for downstream analysis.

    # Example: process genomes to produce cluster definition and MIMAG summary
CHECKM2DB=CheckM2_database/uniref100.KO.1.dmnd galah process \
    --genome-fasta-files genome1.fna genome2.fna \
    --output-cluster-definition clusters.tsv \
    --output-mimag-summary mimag.tsv

    GENOME INPUT

    -f, --genome-fasta-files PATH ..

    Path(s) to FASTA files of each genome e.g. pathA/genome1.fna pathB/genome2.fa.

    -d, --genome-fasta-directory PATH

    Directory containing FASTA files of each genome.

    -x, --genome-fasta-extension EXT

    File extension of genomes in the directory specified with -d/--genome-fasta-directory. [default: fna]

    --genome-fasta-list PATH

    File containing FASTA file paths, one per line.

    QUALITY PARAMETERS

    --quality-method NAME

    method for finding genome quality. 'checkm2' for CheckM2. [default: checkm2]

    --checkm2-db-path PATH

    Path to CheckM2 database (required for CheckM2 quality method). If not given, will use CHECKM2DB environment variable if set.

    --checkm2-quality-report PATH

    Path to pre-generated CheckM2 quality_report.tsv file. If given, will use this file instead of running quality method.

    --checkm-tab-table PATH

    Path to pre-generated CheckM tab table file. If given, will use this file instead of running quality method.

    RNA PARAMETERS

    --rrna-method NAME

    method for finding rRNA genes. 'barrnap' for Barrnap. [default: barrnap]

    --trna-method NAME

    method for finding tRNA genes. 'trnascan' for tRNAscan-SE. [default: trnascan]

    --barrnap-gff-list PATH

    Path to two-column TSV file mapping genome paths (as given in input) to Barrnap GFF paths (no headers). If given, will use these files instead of running rRNA method.

    --trnascan-out-list PATH

    Path to two-column TSV file mapping genome paths (as given in input) to tRNAscan-SE output paths (no headers). If given, will use these files instead of running tRNA method.

    FILTERING PARAMETERS

    --checkm2-quality-report PATH

    CheckM version 2 quality_report.tsv (i.e. the quality_report.tsv in the output directory output of checkm2 predict ..) for defining genome quality, which is used both for filtering and to rank genomes during clustering.

    --checkm-tab-table PATH

    CheckM tab table (i.e. the output of checkm .. --tab_table -f PATH ..). The information contained is used like --checkm2-quality-report.

    --genome-info PATH

    dRep style genome info table for defining quality. The information contained is used like --checkm2-quality-report.

    --min-completeness FLOAT

    Ignore genomes with less completeness than this percentage. [default: not set]

    --max-contamination FLOAT

    Ignore genomes with more contamination than this percentage. [default: not set]

    --run-checkm2

    Run CheckM2 to generate quality scoring used for clustering. Requires --checkm2-db-path or CHECKM2DB env variable to be set.

    --checkm2-db-path DB_PATH

    Path to CheckM2 database (required for running CheckM2) [default: from CHECKM2DB environment variable]

    CLUSTERING PARAMETERS

    --ani FLOAT

    Overall ANI level to dereplicate at with the primary clusterer. [default: 95]

    --min-aligned-fraction FLOAT

    Min aligned fraction of two genomes for clustering. [default: 15]

    --small-genomes

    Use small-genomes settings in skani calculation. Recommended for sequences < 20kb.

    --fragment-length FLOAT

    Length of fragment used in FastANI calculation (i.e. --fragLen). [default: 3000]

    --quality-formula FORMULA

    Scoring function for genome quality [default: Parks2020_reduced]. One of:

    formuladescription
    Parks2020_reduced(default) A quality formula described in Parks et. al. 2020 https://doi.org/10.1038/s41587-020-0501-8 (Supplementary Table 19) but only including those scoring criteria that can be calculated from the sequence without homology searching: completeness-5*contamination-5*num_contigs/100-5*num_ambiguous_bases/100000
    completeness-4contaminationcompleteness-4*contamination
    completeness-5contaminationcompleteness-5*contamination
    dRepcompleteness-5*contamination+contamination*(strain_heterogeneity/100)+0.5*log10(N50)

    --precluster-ani FLOAT

    Require at least this precluster-derived ANI for preclustering and to avoid primary clustering on distant lineages within preclusters. [default: 90]

    --precluster-method NAME

    method of calculating rough ANI for dereplication. 'finch' for finch MinHash, 'skani' for Skani. [default: skani]

    --cluster-method NAME

    method of calculating ANI. 'fastani' for FastANI, 'skani' for Skani. [default: skani]

    --cluster-contigs

    Cluster contigs within a fasta file instead of genomes. When used, either --small-contigs or --large-contigs must be specified.

    --small-contigs

    Use small-genomes settings in skani when clustering contigs. Recommended for contigs < 20kb. Mutually exclusive with --large-contigs.

    --large-contigs

    Do not use small-genomes settings in skani when clustering contigs. Recommended for contigs >= 20kb. Mutually exclusive with --small-contigs.

    --low-memory

    Reduce memory use by sketching to file and searching it instead.

    --reference-genomes PATH ...

    Reference genomes to cluster against. These should be pre-clustered at the chosen %ANI. If quality is provided for representative selection, values for these genomes must also be provided. Genomes within the precluster ANI cutoff of each reference will be placed in the same precluster. Mutually exclusive with --reference-genomes-list.

    --reference-genomes-list PATH

    File containing paths to reference genomes (one per line). These should be pre-clustered at the chosen %ANI. If quality is provided for representative selection, values for these genomes must also be provided. Genomes within the precluster ANI cutoff of each reference will be placed in the same precluster. Mutually exclusive with --reference-genomes.

    OUTPUT

    --output-mimag-summary PATH

    Output a tsv file summarising the MIMAG status for each genome.

    --output-quality-report PATH

    Output a CheckM2-format quality report TSV file.

    --output-cluster-definition PATH

    Output a file of representative<TAB>member lines.

    --output-representative-fasta-directory PATH

    Symlink representative genomes into this directory.

    --output-representative-fasta-directory-copy PATH

    Copy representative genomes into this directory.

    --output-representative-list PATH

    Print newline separated list of paths to representatives into this file.

    GENERAL PARAMETERS

    -t, --threads INT

    Number of threads. [default: 1]

    -v, --verbose

    Print extra debugging information

    -q, --quiet

    Unless there is an error, do not print log messages

    -h, --help

    Output a short usage message.

    --full-help

    Output a full help message and display in 'man'.

    --full-help-roff

    Output a full help message in raw ROFF format for conversion to other formats.

    EXIT STATUS

    0

    Successful program execution.

    1

    Unsuccessful program execution.

    101

    The program panicked.

    AUTHOR

    Ben J. Woodcroft, Centre for Microbiome Research, Queensland University of Technology

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