Annotating contigs

This workflow can be used to transfer taxonomic annotations to virus contigs. The workflow is written in Snakemake. So, if you interupt it in the middle you can start again from where you left off. following command runs skadi with default parameters.

skadi contigs -i <input>.fasta -o outdir

Advance options

However, you can customize skadi with following cmd line options. For example, consider setting --nuc-search mmseqs_tblastx for more sensitive tANI calculations.

Options:
  -i, --input PATH                input file/s to run skadi on  [required]
  -o, --output PATH               dir to store skadi results  [required]
  -d, --database PATH             dir to skadi database
  --nuc_search [blastn|tbalstx|mmseqs_blastn|mmseqs_tblastx]
                                  nucelotide search algorithm (blastn,
                                  tblastx, mmseqs_blastn, mmseqs_tblastx).
                                  [default: mmseqs_blastn]
  --prot_search [blast|diamond|mmseqs]
                                  protein search algorithm (blast, diamond,
                                  mmseqs).  [default: mmseqs]
  --prof_search [mmseqs|hmmer]    nucelotide seach algorithm (mmseqs, hmmer).
                                  [default: mmseqs]
  -j, --jobs INTEGER              use at most this many jobs in parallel (see
                                  cluster submission for more details).
                                  [default: 8]
  --tapif FLOAT                   assign sequences above this tapi threshold
                                  to families  [default: 0.3]
  --tapio FLOAT                   assign sequences above this tapi threshold
                                  to orders  [default: 0.15]
  --tapic FLOAT                   assign sequences above this tapi threshold
                                  to classes  [default: 0.15]
  --tapip FLOAT                   assign sequences above this tapi threshold
                                  to phyla  [default: 0.15]
  --tapik FLOAT                   assign sequences above this tapi threshold
                                  to kingdoms  [default: 0.15]
  --taaig FLOAT                   assign sequences above this taai threshold
                                  to genera  [default: 0.49]
  --taaif FLOAT                   assign sequences above this taai threshold
                                  to families  [default: 0.3]
  --taaio FLOAT                   assign sequences above this taai threshold
                                  to orders  [default: 0.3]
  --taaic FLOAT                   assign sequences above this taai threshold
                                  to classes  [default: 0.3]
  --taaip FLOAT                   assign sequences above this taai threshold
                                  to phyla  [default: 0.3]
  --taaik FLOAT                   assign sequences above this taai threshold
                                  to kingdoms  [default: 0.3]
  --tanis FLOAT                   assign sequences above this taai threshold
                                  to species  [default: 0.81]
  --tanig FLOAT                   assign sequences above this taai threshold
                                  to genera  [default: 0.49]
  --batch INTEGER                 number of records to process at a time
                                  [default: 5000]
  -n, --dryrun                    Test execution.
  -h, --help                      Show this message and exit.

Results

Results can be found in the results directory within the output directory. A typical layout is:

outdir/
├── logs
├── nuc
│   ├── input_genome_ani.tsv
│   └── input_genome.m8
├── prof
│   ├── input_fasta_prof_api.tsv
│   └── input_fasta_prof.m8
├── prot
│   ├── input_fasta.faa
│   ├── input_fasta.gff
│   ├── input_fasta_prot_aai.tsv
│   └── input_fasta_prot.m8
├── results
│   ├── *input_fasta_ictv.csv
│   └── input_fasta.tsv
└── tmp

• results/inpuy_fasta.tsv contains the putative taxonomic annotations for the input contigs.

• nuc/input_genome_ani.tsv contains all the tANI calculations before applying taxon-level tANI cutoffs.

• prot/input_fasta_prot_aai.tsv contains all txAAI calculations before applying taxon-level txAAI cutoffs.

• prof/input_fasta_prof_api.tsv contains all txAPI calculations before applying taxon-level txAPI cutoffs.

• To know more about tANI, txAAI and txAPI, refer Virus contig annotation pipeline

Note

As a rough guideline, it takes around 4 hours to run skadi on the ICTV Taxonomy challenge dataset on a typical laptop.

SKADI output

skadi reports the predicted taxonomic lineage for each query sequence in a structured results table. Each row corresponds to a single sequence and lists the assigned taxa across all ICTV hierarchical ranks, from realm down to species, whenever a match is detected.

The Score column contains the computed similarity metric—tANI, txAAI, or txAPI, depending on which database level produced the annotation. These values range from 0 to 1, with 1.0 indicating a perfect match to a reference at that rank. The Method column specifies which comparison level generated the score:

ani → nucleotide-level comparison (tANI)

aai → protein-level comparison (txAAI)

api → profile HMM comparison (txAPI)

Higher levels of the table represent broader taxonomic classifications (e.g., Realm, Kingdom, Phylum), while lower levels provide increasingly specific annotations (e.g., Family, Genus, Species). Empty fields indicate that no confident match was found at that particular rank.

Together, these columns provide a detailed view of how confidently and at what resolution skadi was able to assign taxonomy to each query sequence.

Taxonomic assignment summary

SequenceID	Seqlen	Score	Method	Realm (-viria)	Realm_score	Subrealm (-vira)	Subrealm_score	Kingdom (-virae)	Kingdom_score	Subkingdom (-virites)	Subkingdom_score	Phylum (-viricota)	Phylum_score	Subphylum (-viricotina)	Subphylum_score	Class (-viricetes)	Class_score	Subclass (-viricetidae)	Subclass_score	Order (-virales)	Order_score	Suborder (-virineae)	Suborder_score	Family (-viridae)	Family_score	Subfamily (-virinae)	Subfamily_score	Genus (-virus)	Genus_score	Subgenus (-virus)	Subgenus_score	Species (binomial)	Species_score
EU188799	7498	1.0	ani	Monodnaviria				Shotokuvirae				Cossaviricota				Papovaviricetes				Zurhausenvirales				Papillomaviridae		Firstpapillomavirinae		Dyoepsilonpapillomavirus				Dyoepsilonpapillomavirus 1
JQ245696	8601	1.0	ani	Riboviria				Orthornavirae				Kitrinoviricota				Alsuviricetes				Tymovirales				Betaflexiviridae		Quinvirinae		Carlavirus				Carlavirus americanense
MK170446	43094	1.0	ani	Duplodnaviria				Heunggongvirae				Uroviricota				Caudoviricetes								Drexlerviridae				Webervirus				Webervirus wv13
U01060	4615	1.0	ani	Riboviria				Orthornavirae				Duplornaviricota				Chrymotiviricetes				Ghabrivirales		Alphatotivirineae		Orthototiviridae				Totivirus				Totivirus ni
KU687349	39444	1.0	ani	Duplodnaviria				Heunggongvirae				Uroviricota				Caudoviricetes				Autographivirales				Autotranscriptaviridae		Studiervirinae		Kayfunavirus				Kayfunavirus SH3
MF352410	7407	1.0	ani	Riboviria				Orthornavirae				Pisuviricota				Pisoniviricetes				Picornavirales				Picornaviridae		Caphthovirinae		Mischivirus				Mischivirus ehoushre
OL436139	42600	1.0	ani	Duplodnaviria				Heunggongvirae				Uroviricota				Caudoviricetes								Sarkviridae		Guernseyvirinae		Jerseyvirus				Jerseyvirus LP31
MK896629	6949	1.0	ani	Riboviria				Orthornavirae				Negarnaviricota		Polyploviricotina		Bunyaviricetes				Elliovirales				Peribunyaviridae				Orthobunyavirus				Orthobunyavirus bellavistaense
MT074450	110183	0.999	ani	Duplodnaviria				Heunggongvirae				Uroviricota				Caudoviricetes								Demerecviridae		Markadamsvirinae		Epseptimavirus				Epseptimavirus faergetype
CASDWV010003421	3859	1.0	ani	Riboviria				Orthornavirae				Lenarviricota				Leviviricetes				Norzivirales				Atkinsviridae				Lobdovirus				Lobdovirus lutivicinum
AY780919	3095	1.0	ani	Riboviria				Orthornavirae																Birnaviridae				Aquabirnavirus				Aquabirnavirus salmonidae
DQ163914	43365	1.0	ani	Duplodnaviria				Heunggongvirae				Uroviricota				Caudoviricetes								Fredfastierviridae				Jamesmcgillvirus				Jamesmcgillvirus jv119X
CASDWV010000512	3611	1.0	ani	Riboviria				Orthornavirae				Lenarviricota				Leviviricetes				Norzivirales				Solspiviridae				Wulvevirus				Wulvevirus geoadaptatum

Utility workflows

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Calculate txAAI of query contigs to ICTV genomes

skadi utils aai [OPTIONS] -i contigs.m8 -g contigs.gff -d [DBDIR]

Calculate tANI of query contigs to ICTV genomes

skadi utils ani [OPTIONS] -i contigs.m8

Visualization of genome comparisons (coming soon)

Create genome comparison plots (query sequence vs highly similar ICTV genomes):

skadi utils visualize --ani --taxa <taxname> -i contigs.m8 -o outdir

Phage contig annotation plots (coming soon)

skadi utils visualize --phrogs -i contigs.fasta -o outdir

Provirus identification (coming soon)

skadi utils provirus -i contigs.fasta -o outdir