nextclade

Align sequences to user provided reference, outputting the alignment to a file

$ nextclade run [path/to/sequences.fa] [[-r|--input-ref]] [path/to/reference.fa] [[-o|--output-fasta]] [path/to/alignment.fa]

$ nextclade run [path/to/fasta] [[-d|--dataset-name]] [dataset_name] [[-t|--output-tsv]] [path/to/report.tsv]

$ nextclade dataset list

$ nextclade dataset get [[-n|--name]] sars-cov-2 [[-o|--output-dir]] [path/to/directory]

$ nextclade run [[-D|--input-dataset]] [path/to/dataset_directory] [[-O|--output-all]] [path/to/output_directory] [path/to/sequences.fasta]

$ nextclade run [[-d|--dataset-name]] [dataset_name] [[-t|--output-tsv]] [path/to/output_tsv] -- [path/to/input_fasta_1 path/to/input_fasta_2 ...]

$ nextclade run --retry-reverse-complement [[-d|--dataset-name]] [dataset_name] [[-t|--output-tsv]] [path/to/output_tsv] [path/to/input_fasta]

nextclade <SUBCOMMAND> [OPTIONS]

run
    Main analysis subcommand for sequences

dataset
    Manage analysis datasets (get, list, update)

--help, -h
    Print help information

--version, -V
    Print version information

--dataset-name <NAME>
    Dataset name (e.g. 'sars-cov-2')

--dataset-url <URL>
    URL to zip dataset

--input-sequences <PATH>
    Input FASTA/FASTQ sequences

--input-reference <PATH>
    Custom reference FASTA

--output-tsv <PATH>
    Tab-separated results

--output-csv <PATH>
    Comma-separated results

--output-json <PATH>
    JSON results

--output-fasta <PATH>
    Aligned FASTA output

--output-tree <PATH>
    Phylogenetic tree (Nexus)

--threads <N>
    Number of CPU threads

--include-endpoint-mutations
    Include mutations outside gene regions

--output-basename <STR>
    Base name for all outputs

Nextclade is a fast, scalable command-line tool for analyzing viral genomes, especially SARS-CoV-2. It processes FASTA/FASTQ input sequences to perform:

• Clade assignment (Nextstrain, WHO)
• Mutation calling (nucleotide/aa substitutions)
• Quality control (scoring missing data, divergences)
• Alignment to reference genomes
• Pango lineage inference

Using predefined datasets (genes, references, trees), it outputs results in TSV, CSV, JSON, aligned FASTA, and phylogenetic trees (Nexus/Newick). Supports multi-threading for high-throughput surveillance.

Part of the Nextstrain ecosystem, it's used globally for COVID-19 tracking. Datasets auto-update for latest variants. Install via Conda, Cargo, or binaries. Handles thousands of sequences efficiently on standard hardware.

Requires internet for default datasets; download locally for offline use.
Large inputs may need significant RAM (>8GB recommended).
Primarily optimized for SARS-CoV-2; check dataset compatibility for other viruses.

Prebuilt collections (reference, genes, tree) for viruses. Use nextclade dataset get --name sars-cov-2 to download.

Core TSV/JSON fields: clade, pango_lineage, qc_overall_score, substitutions, aa_substitutions.

Via Conda: conda install -c bioconda nextclade; Cargo: cargo install nextclade; or prebuilt binaries.

Developed by Nextstrain team in 2020 for COVID-19 surveillance. First release coincided with pandemic; evolved with variants (Alpha to Omicron+). Weekly dataset updates since 2021. Version 2.x introduced multi-virus support.

nextalign(1), pangolin(1), iqtree(1)