Usage

Input Types

StrainCascade accepts two mutually exclusive input types:

Flag Input Type Accepted Formats
-i Sequencing reads .fasta, .fa, .fna, .fastq, .fastq.gz, .bam
-a Pre-assembled genomes .fasta, .fa, .fna

Both flags accept a single file or a directory containing multiple files. When a directory is provided, all matching files are processed sequentially.

Sequencing Types

Parameter Platform Description
pacbio-hifi PacBio HiFi/CCS reads (default)
pacbio-corr PacBio Corrected subreads
pacbio-raw PacBio Raw subreads
nano-hq ONT High-quality (Q20+, R10.4+)
nano-corr ONT Corrected reads
nano-raw ONT Raw reads (older chemistry) 
straincascade -i reads.fastq.gz -s nano-hq -t 32

Hybrid Assembly

Providing paired Illumina short reads activates two features:

  1. Unicycler Hybrid Assembly (SC6) — combines long and short reads for improved assembly
  2. Short-Read Polishing (SC12) — BWA-MEM alignment + Polypolish consensus correction
straincascade -i longreads.fastq.gz \
    -sr1 R1.fastq.gz -sr2 R2.fastq.gz \
    -o output/ -s pacbio-hifi -t 32

When only long reads are provided, Unicycler falls back to long-read-only mode (miniasm+Racon).

External Assemblies

Provide pre-existing assemblies from external tools for inclusion in the de novo phylogenetic tree (SC16):

straincascade -i reads.fastq.gz -ea /path/to/external_assemblies/ -o output/

Assembly Selection Algorithm

Two algorithms are available for selecting the best assembly from the multi-assembler suite:

Algorithm Method
contig (default) MAD-based outlier filtering on genome size, then minimum contig count
continuity Bivariate MAD filtering on both genome size and contig count 
straincascade -i reads.fastq.gz -sa continuity

Locus Tag

By default, StrainCascade generates an automatic INSDC-compliant locus tag from the GTDB-Tk taxonomic classification. You can override this:

straincascade -i reads.fastq.gz -l MYORG
Note

INSDC locus tag prefixes must contain 3–12 alphanumeric uppercase characters and start with a letter.

Reproducibility Modes

Mode Threads Entropy Use Case
--heuristic (default) User-specified System Fast, standard analysis
--deterministic Forced to 1 Fixed file Bit-identical reproducibility 
# Deterministic mode
straincascade -i reads.fastq.gz --deterministic

# Heuristic mode (default)
straincascade -i reads.fastq.gz --heuristic -t 32

Result Types

Type Description
main (default) Standard output files
all All intermediate and final files
R R-friendly output format 
straincascade -i reads.fastq.gz -r all

Force Overwrite

By default, existing output files are overwritten. To skip existing outputs:

straincascade -i reads.fastq.gz -f no

Updating StrainCascade

# Update software scripts
straincascade -us

# Update Apptainer images
straincascade -uai

# Update databases
straincascade -udb

For instructions on full reinstallation, see Installation.

Caveats

  • Conda environment and PATH in SBATCH jobs: When submitting sbatch jobs, your conda environment will not remain active automatically. If you have multiple installations, activate the correct environment in your SBATCH script to ensure the appropriate paths are used.

  • Resource requirements: StrainCascade is resource-intensive and will overwhelm standard desktop or laptop computers. Use an HPC environment with at least 32 cores and 96 GB RAM for optimal performance.

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