# nf-core/funcscan: Output

## Introduction

The output of nf-core/funcscan provides reports for each of the functional groups:

- antibiotic resistance genes (tools: [ABRicate](https://github.com/tseemann/abricate), [AMRFinderPlus](https://www.ncbi.nlm.nih.gov/pathogens/antimicrobial-resistance/AMRFinder), [DeepARG](https://bitbucket.org/gusphdproj/deeparg-ss/src/master), [fARGene](https://github.com/fannyhb/fargene), [RGI](https://card.mcmaster.ca/analyze/rgi) – summarised by [hAMRonization](https://github.com/pha4ge/hAMRonization))
- antimicrobial peptides (tools: [Macrel](https://github.com/BigDataBiology/macrel), [AMPlify](https://github.com/bcgsc/AMPlify), [ampir](https://ampir.marine-omics.net), [hmmsearch](http://hmmer.org) – summarised by [AMPcombi](https://github.com/Darcy220606/AMPcombi))
- biosynthetic gene clusters (tools: [antiSMASH](https://docs.antismash.secondarymetabolites.org), [DeepBGC](https://github.com/Merck/deepbgc), [GECCO](https://gecco.embl.de), [hmmsearch](http://hmmer.org) – summarised by [comBGC](#combgc))

As a general workflow, we recommend to first look at the summary reports ([ARGs](#hamronization), [AMPs](#ampcombi), [BGCs](#combgc)), to get a general overview of what hits have been found across all the tools of each functional group. After which, you can explore the specific output directories of each tool to get more detailed information about each result. The tool-specific output directories also includes the output from the functional annotation steps of either [prokka](https://github.com/tseemann/prokka), [pyrodigal](https://github.com/althonos/pyrodigal), [prodigal](https://github.com/hyattpd/Prodigal), or [Bakta](https://github.com/oschwengers/bakta) if the `--save_annotations` flag was set.

Similarly, all downloaded databases are saved (i.e. from [antiSMASH](https://docs.antismash.secondarymetabolites.org), [AMRFinderPlus](https://www.ncbi.nlm.nih.gov/pathogens/antimicrobial-resistance/AMRFinder), [Bakta](https://github.com/oschwengers/bakta), [DeepARG](https://bitbucket.org/gusphdproj/deeparg-ss/src/master), and/or [AMPcombi](https://github.com/Darcy220606/AMPcombi)) into the output directory `<outdir>/downloads/` if the `--save_databases` flag was set.

Furthermore, for reproducibility, versions of all software used in the run is presented in a [MultiQC](http://multiqc.info) report.

The directories listed below will be created in the results directory (specified by the `--outdir` flag) after the pipeline has finished. All paths are relative to this top-level output directory. The default directory structure of nf-core/funcscan is:

```console
results/
├── annotation/
|   ├── bakta/
|   ├── prodigal
|   ├── prokka/
|   └── pyrodigal/
├── amp/
|   ├── ampir/
|   ├── amplify/
|   ├── hmmsearch/
|   └── macrel/
├── arg/
|   ├── abricate/
|   ├── amrfinderplus/
|   ├── deeparg/
|   ├── fargene/
|   ├── hamronization/
|   └── rgi/
├── bgc/
|   ├── antismash/
|   ├── deepbgc/
|   ├── gecco/
|   └── hmmsearch/
├── reports/
|   ├── ampcombi/
|   ├── comBGC/
|   └── hamronization_summarize/
├── databases/
├── multiqc/
└── pipeline_info/
work/
```

## Pipeline overview

The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes prokaryotic sequence data through the following steps:

ORF prediction and annotation with any of:

- [Pyrodigal](#pyrodigal) (default) – for open reading frame prediction.
- [Prodigal](#prodigal) – for open reading frame prediction.
- [Prokka](#prokka) – open reading frame prediction and functional protein annotation.
- [Bakta](#bakta) – open reading frame prediction and functional protein annotation.

Antimicrobial Resistance Genes (ARGs):

- [ABRicate](#abricate) – antimicrobial resistance gene detection, based on alignment to one of several databases.
- [AMRFinderPlus](#amrfinderplus) – antimicrobial resistance gene detection, using NCBI’s curated Reference Gene Database and curated collection of Hidden Markov Models.
- [DeepARG](#deeparg) – antimicrobial resistance gene detection, using a deep learning model.
- [fARGene](#fargene) – antimicrobial resistance gene detection, using Hidden Markov Models.
- [RGI](#rgi) – antimicrobial resistance gene detection, based on alignment to the CARD database.

Antimicrobial Peptides (AMPs):

- [ampir](#ampir) – antimicrobial peptide detection, based on a supervised statistical machine learning approach.
- [amplify](#amplify) – antimicrobial peptide detection, using a deep learning model.
- [hmmsearch](#hmmsearch) – antimicrobial peptide detection, based on hidden Markov models.
- [Macrel](#macrel) – antimicrobial peptide detection, using a machine learning approach.

Biosynthetic Gene Clusters (BGCs):

- [antiSMASH](#antismash) – biosynthetic gene cluster detection.
- [deepBGC](#deepbgc) - biosynthetic gene cluster detection, using a deep learning model.
- [GECCO](#gecco) – biosynthetic gene cluster detection, using Conditional Random Fields (CRFs).
- [hmmsearch](#hmmsearch) – biosynthetic gene cluster detection, based on hidden Markov models.

Output Summaries:

- [AMPcombi](#ampcombi) – summary of antimicrobial peptide gene output from various detection tools.
- [hAMRonization](#hamronization) – summary of antimicrobial resistance gene output from various detection tools.
- [comBGC](#combgc) – summary of biosynthetic gene cluster output from various detection tools.
- [MultiQC](#multiqc) – report of all software and versions used in the pipeline.
- [Pipeline information](#pipeline-information) – report metrics generated during the workflow execution.

## Tool details

### Annotation tools

[Pyrodigal](#pyrodigal), [Prodigal](#prodigal), [Prokka](#prokka), [Bakta](#bakta)

#### Prodigal

<details markdown="1">
<summary>Output files</summary>

- `prodigal/`
  - `<samplename>/`:
    - `*.gff`: annotation in GFF3 format, containing both sequences and annotations
    - `*.fna`: nucleotide FASTA file of the input contig sequences
    - `*.faa`: protein FASTA file of the translated CDS sequences
    - `*.gbk`: annotation in GBK format, containing both sequences and annotations

> Descriptions taken from the [Prodigal documentation](https://github.com/hyattpd/prodigal/wiki)

</details>

[Prodigal](https://github.com/hyattpd/Prodigal) annotates whole (meta-)genomes by identifying ORFs in a set of genomic DNA sequences. The output is used by some of the functional screening tools.

#### Pyrodigal

<details markdown="1">
<summary>Output files</summary>

- `pyrodigal/`
  - `<samplename>/`:
    - `*.gff`: annotation in GFF3 format, containing both sequences and annotations
    - `*.fna`: nucleotide FASTA file of the input contig sequences
    - `*.faa`: protein FASTA file of the translated CDS sequences

> Descriptions taken from the [Pyrodigal documentation](https://pyrodigal.readthedocs.io/)

</details>

[Pyrodigal](https://github.com/althonos/pyrodigal) annotates whole (meta-)genomes by identifying ORFs in a set of genomic DNA sequences. It produces the same results as [Prodigal](#prodigal) while being more resource-optimized, thus faster. Unlike Prodigal, Pyrodigal cannot produce output in GenBank format. The output is used by some of the functional screening tools.

#### Prokka

<details markdown="1">
<summary>Output files</summary>

- `prokka/`
  - `<samplename>/`
    - `*.gff`: annotation in GFF3 format, containing both sequences and annotations
    - `*.gbk`: standard Genbank file derived from the master .gff
    - `*.fna`: nucleotide FASTA file of the input contig sequences
    - `*.faa`: protein FASTA file of the translated CDS sequences
    - `*.ffn`: nucleotide FASTA file of all the prediction transcripts (CDS, rRNA, tRNA, tmRNA, misc_RNA)
    - `*.sqn`: an ASN1 format "Sequin" file for submission to Genbank
    - `*.fsa`: nucleotide FASTA file of the input contig sequences, used by "tbl2asn" to create the .sqn file
    - `*.tbl`: feature Table file, used by "tbl2asn" to create the .sqn file
    - `*.err`: unacceptable annotations - the NCBI discrepancy report
    - `*.log`: logging output that Prokka produced during its run
    - `*.txt`: statistics relating to the annotated features found
    - `*.tsv`: tab-separated file of all features

> Descriptions directly from the [Prokka documentation](https://github.com/tseemann/prokka#output-files)

</details>

[Prokka](https://github.com/tseemann/prokka) performs whole genome annotation to identify features of interest in a set of (meta-)genomic DNA sequences. The output is used by some of the functional screening tools.

#### Bakta

<details markdown="1">
<summary>Output files</summary>

- `bakta/`
  - `<samplename>.gff3`: annotations & sequences in GFF3 format
  - `<samplename>.gbff`: annotations & sequences in (multi) GenBank format
  - `<samplename>.ffn`: feature nucleotide sequences as FASTA
  - `<samplename>.fna`: replicon/contig DNA sequences as FASTA
  - `<samplename>.embl`: annotations & sequences in (multi) EMBL format
  - `<samplename>.faa`: CDS/sORF amino acid sequences as FASTA
  - `<samplename>_hypothetical.faa`: further information on hypothetical protein CDS as simple human readble tab separated values
  - `<samplename>_hypothetical.tsv`: hypothetical protein CDS amino acid sequences as FASTA
  - `<samplename>.tsv`: annotations as simple human readble TSV
  - `<samplename>.txt`: summary in TXT format

> Descriptions directly from the [Bakta documentation](https://github.com/oschwengers/bakta#output).

</details>

[Bakta](https://github.com/oschwengers/bakta) is a tool for the rapid & standardised annotation of bacterial genomes and plasmids from both isolates and MAGs. It provides dbxref-rich, sORF-including and taxon-independent annotations in machine-readable JSON & bioinformatics standard file formats for automated downstream analysis. The output is used by some of the functional screening tools.

### AMP detection tools

[ampir](#ampir), [AMPlify](#amplify), [hmmsearch](#hmmsearch), [Macrel](#macrel)

#### ampir

<details markdown="1">
<summary>Output files</summary>

- `ampir/`
  - `<samplename>.ampir.faa`: predicted AMP sequences in FAA format
  - `<samplename>.ampir.tsv`: predicted AMP metadata in TSV format, contains contig name, sequence and probability score

</details>

[ampir](https://github.com/Legana/ampir) (**a**nti**m**icrobial **p**eptide **p**rediction **i**n **r**) was designed to predict antimicrobial peptides (AMPs) from any given size protein dataset. ampir uses a supervised statistical machine learning approach to predict AMPs. It incorporates two support vector machine classification models, “precursor” and “mature” that have been trained on publicly available antimicrobial peptide data.

#### AMPlify

<details markdown="1">
<summary>Output files</summary>

- `amplify/`
  - `*_results.tsv`: table of contig amino-acid sequences with prediction result (AMP or non-AMP) and information on sequence length, charge, probability score, AMPlify log-scaled score)

</details>

[AMPlify](https://github.com/bcgsc/AMPlify) is an attentive deep learning model for antimicrobial peptide prediction. It takes in contig annotations (as protein sequences) and classifies them as either AMP or non-AMP.

#### hmmsearch

<details markdown="1">
<summary>Output files</summary>

- `hmmersearch/`
  - `*.txt.gz`: human readable output summarizing hmmsearch results
  - `*.sto.gz`: optional multiple sequence alignment (MSA) in Stockholm format
  - `*.tbl.gz`: optional tabular (space-delimited) summary of per-target output
  - `*.domtbl.gz`: optional tabular (space-delimited) summary of per-domain output

</details>

[HMMER/hmmsearch](http://hmmer.org) is used for searching sequence databases for sequence homologs, and for making sequence alignments. It implements methods using probabilistic models called profile hidden Markov models (profile HMMs). `hmmsearch` is used to search one or more profiles against a sequence database.

#### Macrel

<details markdown="1">
<summary>Output files</summary>

- `macrel_contigs/`
  - `*.smorfs.faa.gz`: zipped fasta file containing amino acid sequences of small peptides (<100 aa, small open reading frames) showing the general gene prediction information in the contigs
  - `*.all_orfs.faa.gz`: zipped fasta file containing amino acid sequences showing the general gene prediction information in the contigs
  - `prediction.gz`: zipped file, with all predicted and non-predicted AMPs in a table format
  - `*.md`: readme file containing tool specific information (e.g. citations, details about the output, etc.)
  - `*_log.txt`: log file containing the information pertaining to the run

</details>

[Macrel](https://github.com/BigDataBiology/macrel) is a tool that mines antimicrobial peptides (AMPs) from (meta)genomes by predicting peptides from genomes (provided as contigs) and outputs predicted antimicrobial peptides that meet specific criteria/thresholds.

### ARG detection tools

[ABRicate](#abricate), [AMRFinderPlus](#amrfinderplus), [DeepARG](#deeparg), [fARGene](#fargene), [RGI](#rgi)

#### ABRicate

<details markdown="1">
<summary>Output files</summary>

- `abricate/`
  - `*.{csv,tsv}`: search results in tabular format

</details>

[ABRicate](https://github.com/tseemann/abricate) screens contigs for antimicrobial resistance or virulence genes. It comes bundled with multiple databases: NCBI, CARD, ARG-ANNOT, Resfinder, MEGARES, EcOH, PlasmidFinder, Ecoli_VF and VFDB.

#### AMRFinderPlus

<details markdown="1">
<summary>Output files</summary>

- `amrfinderplus/`
  - `*.tsv`: search results in tabular format

</details>

[AMRFinderPlus](https://www.ncbi.nlm.nih.gov/pathogens/antimicrobial-resistance/AMRFinder) relies on NCBI’s curated Reference Gene Database and curated collection of Hidden Markov Models. It identifies antimicrobial resistance genes, resistance-associated point mutations, and select other classes of genes using protein annotations and/or assembled nucleotide sequences.

#### DeepARG

<details markdown="1">
<summary>Output files</summary>

- `deeparg/`
  - `*.align.daa*`: Intermediate DIAMOND alignment output
  - `*.align.daa.tsv`: DIAMOND alignment output as .tsv
  - `*.mapping.ARG`: ARG predictions with a probability >= --prob (0.8 default).
  - `*.mapping.potential.ARG`: ARG predictions with a probability < --prob (0.8 default)

</details>

[deepARG](https://bitbucket.org/gusphdproj/deeparg-ss/src/master) uses deep learning to characterize and annotate antibiotic resistance genes in metagenomes. It is composed of two models for two types of input: short sequence reads and gene-like sequences. In this pipeline we use the `ls` model, which is suitable for annotating full sequence genes and to discover novel antibiotic resistance genes from assembled samples. The tool `DIAMOND` is used as an aligner.

#### fARGene

<details markdown="1">
<summary>Output files</summary>

- `fargene/`
  - `fargene_analysis.log`: logging output that fARGene produced during its run
  - `<sample_name>/`:
    - `hmmsearchresults/`: output from intermediate hmmsearch step (only if `--arg_fargene_savetmpfiles` supplied)
    - `predictedGenes/`:
      - `*-filtered.fasta`: nucleotide sequences of predicted ARGs
      - `*-filtered-peptides.fasta`: amino acid sequences of predicted ARGs
    - `results_summary.txt`: text summary of results, listing predicted genes and ORFs for each input file
    - `tmpdir/`: temporary output files and fasta files (only if `--arg_fargene_savetmpfiles` supplied)

</details>

[fARGene](https://github.com/fannyhb/fargene) (**F**ragmented **A**ntibiotic **R**esistance **Gene** Identifier) is a tool that takes either fragmented metagenomic data or longer sequences as input and predicts and delivers full-length antibiotic resistance genes as output. The tool includes developed and optimised models for a number of resistance gene types. By default the pipeline runs all models, thus you will receive output for all models. If only a sub-list or single model is required, this can be specified with the `--hmm-model` flag. Available models are:

- `class_a`: class A beta-lactamases
- `class_b_1_2`: subclass B1 and B2 beta-lactamases
- `class_b3`: subclass B3 beta-lactamases
- `class_c`: class C beta-lactamases
- `class_d_1, class_d_2`: class D beta-lactamases
- `qnr`: quinolone resistance genes
- `tet_efflux`, `tet_rpg`, `tet_enzyme`: tetracycline resistance genes

#### RGI

<details markdown="1">
<summary>Output files</summary>

- `rgi/`
  - `<samplename>.txt`: hit results table separated by '#'
  - `<samplename>.json`: hit results in json format (only if `--arg_rgi_savejson` supplied)
  - `temp/`:
    - `<samplename>.fasta.temp.*.json`: temporary json files, '\*' stands for 'homolog', 'overexpression', 'predictedGenes' and 'predictedGenes.protein' (only if `--arg_rgi_savetmpfiles` supplied).

</details>

[RGI](https://github.com/arpcard/rgi) (**R**esistance **G**ene **I**dentifier) predicts resistome(s) from protein or nucleotide data based on homology and SNP models. It uses reference data from the Comprehensive Antibiotic Resistance Database (CARD).

### BGC detection tools

[antiSMASH](#antismash), [deepBGC](#deepbgc), [GECCO](#gecco), [hmmsearch](#hmmsearch)

#### antiSMASH

<details markdown="1">
<summary>Output files</summary>

- `antismash/`
  - `css`: accessory files for the HTML output
  - `clusterblastoutput.txt` (optional): raw BLAST output of known clusters previously predicted by antiSMASH using the built-in ClusterBlast algorithm
  - `images`: accessory files for the HTML output
  - `index.html`: interactive web view of results in HTML format
  - `js`: accessory files for the HTML output
  - `knownclusterblast/`: directory with MIBiG hits (optional)
    - `*_c*.txt`: tables with MIBiG hits
  - `knownclusterblastoutput.txt` (optional): raw BLAST output of known clusters of the MIBiG database.
  - `regions.js`: sideloaded annotations of protoclusters and/or subregions
  - `*region*.gbk`: nucleotide sequence + annotations in GenBank file format; one file per antiSMASH hit.
  - `<sample name>.gbk`: nucleotide sequence and annotations in GenBank format; converted from input file
  - `<sample name>.json`: nucleotide sequence and annotations in JSON format; converted from GenBank file
  - `<sample name>.log`: logging output that antiSMASH produced during its run
  - `<sample name>.zip`: compressed version of the output folder in zip format

</details>

[antiSMASH](https://docs.antismash.secondarymetabolites.org) (**anti**biotics & **S**econdary **M**etabolite **A**nalysis **SH**ell) is a tool for rapid genome-wide identification, annotation and analysis of secondary metabolite biosynthesis gene clusters in bacterial genomes. It identifies biosynthetic loci covering all currently known secondary metabolite compound classes in a rule-based fashion using profile HMMs and aligns the identified regions at the gene cluster level to their nearest relatives from a database containing experimentally verified gene clusters (MIBiG).

#### deepBGC

<details markdown="1">
<summary>Output files</summary>

- `deepbgc/`
  - `README.txt`: Summary of output files generated
  - `LOG.txt`: Log output of DeepBGC
  - `*.antismash.json`: AntiSMASH JSON file for sideloading
  - `*.bgc.gbk`: Sequences and features of all detected BGCs in GenBank format
  - `*.bgc.tsv`: Table of detected BGCs and their properties
  - `*.full.gbk`: Fully annotated input sequence with proteins, Pfam domains (PFAM_domain features) and BGCs (cluster features)
  - `*.pfam.tsv`: Table of Pfam domains (pfam_id) from given sequence (sequence_id) in genomic order, with BGC detection scores
  - `evaluation/`
    - `*.bgc.png`: Detected BGCs plotted by their nucleotide coordinates
    - `*.pr.png`: Precision-Recall curve based on predicted per-Pfam BGC scores
    - `*.roc.png`: ROC curve based on predicted per-Pfam BGC scores
    - `*.score.png`: BGC detection scores of each Pfam domain in genomic order

</details>

[deepBGC](https://github.com/Merck/deepbgc) detects BGCs in bacterial and fungal genomes using deep learning. DeepBGC employs a Bidirectional Long Short-Term Memory Recurrent Neural Network and a word2vec-like vector embedding of Pfam protein domains. Product class and activity of detected BGCs is predicted using a Random Forest classifier.

#### GECCO

<details markdown="1">
<summary>Output files</summary>

- `gecco/`
  - `*.genes.tsv/`: TSV file containing detected/predicted genes with BGC probability scores
  - `*.features.tsv`: TSV file containing identified domains
  - `*.clusters.tsv`: TSV file containing coordinates of predicted clusters and BGC types
  - `*_cluster_*.gbk`: GenBank file (if clusters were found) containing sequence with annotations; one file per GECCO hit

</details>

[GECCO](https://gecco.embl.de) is a fast and scalable method for identifying putative novel Biosynthetic Gene Clusters (BGCs) in genomic and metagenomic data using Conditional Random Fields (CRFs).

### Summary tools

[AMPcombi](#ampcombi), [hAMRonization](#hamronization), [comBGC](#combgc), [MultiQC](#multiqc), [pipeline information](#pipeline-information)

#### AMPcombi

<details markdown="1">
<summary>Output files</summary>

- `ampcombi/`
  - `ampcombi_complete_summary.csv.gz`: summarised output from all AMP workflow tools (except hmmer_hmmsearch) in compressed csv format
  - `ampcombi.log`: a log file generated by ampcombi
  - `*_ampcombi.csv`: summarised output in csv for each sample
  - `*_amp.faa*`: fasta file containing the amino acid sequences for all AMP hits for each sample
  - `*_diamond_matches.txt*`: alignment file generated by DIAMOND for each sample

</details>

<details markdown="1">
<summary>AMP summary table header descriptions</summary>

| Table column              | Description                                                                                                                                                                                                                                                                                                                                                                                                                        |
| ------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| `name`                    | Name of the sample                                                                                                                                                                                                                                                                                                                                                                                                                 |
| `contig_id`               | Contig header                                                                                                                                                                                                                                                                                                                                                                                                                      |
| `prob_macrel`             | Probability associated with the AMP prediction using `MACREL`                                                                                                                                                                                                                                                                                                                                                                      |
| `prob_neubi`              | Probability associated with the AMP prediction using `NEUBI`                                                                                                                                                                                                                                                                                                                                                                       |
| `prob_ampir`              | Probability associated with the AMP prediction using `AMPIR`                                                                                                                                                                                                                                                                                                                                                                       |
| `prob_amplify`            | Probability associated with the AMP prediction using `AMPLIFY`                                                                                                                                                                                                                                                                                                                                                                     |
| `evalue_hmmer`            | Expected number of false positives (nonhomologous sequences) with a similar of higher score. This stands for how significant the hit is, the lower the evalue, the more significant the hit                                                                                                                                                                                                                                        |
| `aa_sequence`             | Amino acid sequence that forms part of the contig and is AMP encoding                                                                                                                                                                                                                                                                                                                                                              |
| `target_id`               | [DRAMP](http://dramp.cpu-bioinfor.org/) ID within the database found to be similar to the predicted AMP by `DIAMOND` alignment                                                                                                                                                                                                                                                                                                     |
| `pident`                  | Percentage identity of amino acid residues that fully aligned between the `DRAMP` sequence and the predicted AMP sequence                                                                                                                                                                                                                                                                                                          |
| `evalue`                  | Number of alignments of similar or better qualities that can be expected when searching a database of similar size with a random sequence distribution. This is generated by `DIAMOND` alignments using the [DRAMP](http://dramp.cpu-bioinfor.org/) AMP database. The lower the value the more significant that the hit is positive. An e-value of < 0.001 means that the this hit will be found by chance once per 1,0000 queries |
| `Sequence`                | Sequence corresponding to the `DRAMP` ID found to be similar to the predicted AMP sequence                                                                                                                                                                                                                                                                                                                                         |
| `Sequence_length`         | Number of amino acid residues in the `DRAMP` sequence                                                                                                                                                                                                                                                                                                                                                                              |
| `Name`                    | Full name of the peptide copied from the database it was uploaded to                                                                                                                                                                                                                                                                                                                                                               |
| `Swiss_Prot_Entry`        | Entry name of the peptide within the [UniProtKB/Swiss-Prot](https://www.uniprot.org/help/entry_name) database                                                                                                                                                                                                                                                                                                                      |
| `Family`                  | Name of the family, group or class of AMPs this peptide belongs to, e.g. bacteriocins                                                                                                                                                                                                                                                                                                                                              |
| `Gene`                    | Name of the gene (if available in the database) that encodes the peptide                                                                                                                                                                                                                                                                                                                                                           |
| `Source`                  | Name of the source organism (if available in the database) from which the peptide was extracted                                                                                                                                                                                                                                                                                                                                    |
| `Activity`                | Peptide activity, e.g. Antimicrobial, Antibacterial, Anti-Gram+, Anti-Gram-, Insecticidal or Antifungal                                                                                                                                                                                                                                                                                                                            |
| `Protein_existence`       | Peptide status, e.g. only a homology, protein level, predicted or transcript level                                                                                                                                                                                                                                                                                                                                                 |
| `Structure`               | Type of peptide structure, e.g. alpha helix, bridge, etc.                                                                                                                                                                                                                                                                                                                                                                          |
| `Structure_Description`   | Further description of the structure if available                                                                                                                                                                                                                                                                                                                                                                                  |
| `PDB_ID`                  | The ID of an equivalent peptide found in the protein data bank [PDB](https://www.rcsb.org/docs/general-help/organization-of-3d-structures-in-the-protein-data-bank)                                                                                                                                                                                                                                                                |
| `Comments`                | Further details found in the database regarding the peptide                                                                                                                                                                                                                                                                                                                                                                        |
| `Target_Organism`         | Name of the target organism to which the peptide is effective against                                                                                                                                                                                                                                                                                                                                                              |
| `Hemolytic_activity`      | Type of hemolytic activity if any                                                                                                                                                                                                                                                                                                                                                                                                  |
| `Linear/Cyclic/Branched`  | Whether the hit is a linear, cyclic or branched peptide                                                                                                                                                                                                                                                                                                                                                                            |
| `N-terminal_Modification` | Whether it contains N-terminal_Modification                                                                                                                                                                                                                                                                                                                                                                                        |
| `C-termina_Modification`  | Whether it contains C-terminal_Modification                                                                                                                                                                                                                                                                                                                                                                                        |
| `Other_Modifications`     | Whether there are any other modifications found in the peptide structure                                                                                                                                                                                                                                                                                                                                                           |
| `Stereochemistry`         | Type of peptide stereochemistry if available                                                                                                                                                                                                                                                                                                                                                                                       |
| `Cytotoxicity`            | Cytotoxicity mechanism of the peptide if available                                                                                                                                                                                                                                                                                                                                                                                 |
| `Binding_Target`          | Peptide binding target, e.g. lipid, cell membrane or chitin binding                                                                                                                                                                                                                                                                                                                                                                |
| `Pubmed_ID`               | Pubmed ID if a publication is associated with the peptide                                                                                                                                                                                                                                                                                                                                                                          |
| `Reference`               | Citation of the associated publication if available                                                                                                                                                                                                                                                                                                                                                                                |
| `Author`                  | Authors' names associated with the publication or who have uploaded the peptide                                                                                                                                                                                                                                                                                                                                                    |
| `Title`                   | Publication title if available                                                                                                                                                                                                                                                                                                                                                                                                     |

</details>

[AMPcombi](https://github.com/Darcy220606/AMPcombi) summarizes the results of **antimicrobial peptide (AMP)** prediction tools (AMPIR, AMPLIFY, MACREL, and other non nf-core tools) into a single table and aligns the hits against a reference AMP database for functional and taxonomic classification.

#### hAMRonization

<details markdown="1">
<summary>Output files</summary>

- `hamronization/` one of the following:
  - `hamronization_combined_report.json`: summarised output in .json format
  - `hamronization_combined_report.tsv`: summarised output in .tsv format
  - `hamronization_combined_report.html`: interactive output in .html format

</details>
<details markdown="1">
<summary>ARG summary table headers</summary>

| Table column                                      | Description                                                                                                                                                                         |
| ------------------------------------------------- | ----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| `input_file_name`                                 | Name of the file containing the sequence data to be analysed                                                                                                                        |
| `gene_symbol`                                     | Short name of a gene; a single word that does not contain white space characters. It is typically derived from the gene name                                                        |
| `gene_name`                                       | Name of a gene                                                                                                                                                                      |
| `reference_database_name`                         | Identifier of a biological or bioinformatics database                                                                                                                               |
| `reference_database_version`                      | Version of the database containing the reference sequences used for analysis                                                                                                        |
| `reference_accession`                             | Identifier that specifies an individual sequence record in a public sequence repository                                                                                             |
| `analysis_software_name`                          | Name of a computer package, application, method or function used for the analysis of data                                                                                           |
| `analysis_software_version`                       | Version of software used to analyze data                                                                                                                                            |
| `genetic_variation_type`                          | Class of genetic variation detected                                                                                                                                                 |
| `antimicrobial_agent` (optional)                  | A substance that kills or slows the growth of microorganisms, including bacteria, viruses, fungi and protozoans                                                                     |
| `coverage_percentage` (optional)                  | Percentage of the reference sequence covered by the sequence of interest                                                                                                            |
| `coverage_depth` (optional)                       | Average number of reads representing a given nucleotide in the reconstructed sequence                                                                                               |
| `coverage_ratio` (optional)                       | Ratio of the reference sequence covered by the sequence of interest.                                                                                                                |
| `drug_class` (optional)                           | Set of antibiotic molecules, with similar chemical structures, molecular targets, and/or modes and mechanisms of action                                                             |
| `input_gene_length` (optional)                    | Length (number of positions) of a target gene sequence submitted by a user                                                                                                          |
| `input_gene_start` (optional)                     | Position of the first nucleotide in a gene sequence being analysed (input gene sequence)                                                                                            |
| `input_gene_stop` (optional)                      | Position of the last nucleotide in a gene sequence being analysed (input gene sequence)                                                                                             |
| `input_protein_length` (optional)                 | Length (number of positions) of a protein target sequence submitted by a user                                                                                                       |
| `input_protein_start` (optional)                  | Position of the first amino acid in a protein sequence being analysed (input protein sequence)                                                                                      |
| `input_protein_stop` (optional)                   | Position of the last amino acid in a protein sequence being analysed (input protein sequence)                                                                                       |
| `input_sequence_id` (optional)                    | Identifier of molecular sequence(s) or entries from a molecular sequence database                                                                                                   |
| `nucleotide_mutation` (optional)                  | Nucleotide sequence change(s) detected in the sequence being analysed compared to a reference                                                                                       |
| `nucleotide_mutation_interpretation` (optional)   | Description of identified nucleotide mutation(s) that facilitate clinical interpretation                                                                                            |
| `predicted_phenotype` (optional)                  | Characteristic of an organism that is predicted rather than directly measured or observed                                                                                           |
| `predicted_phenotype_confidence_level` (optional) | Confidence level in a predicted phenotype                                                                                                                                           |
| `amino_acid_mutation` (optional)                  | Amino acid sequence change(s) detected in the sequence being analysed compared to a reference                                                                                       |
| `amino_acid_mutation_interpretation` (optional)   | Description of identified amino acid mutation(s) that facilitate clinical interpretation.                                                                                           |
| `reference_gene_length` (optional)                | Length (number of positions) of a gene reference sequence retrieved from a database                                                                                                 |
| `reference_gene_start` (optional)                 | Position of the first nucleotide in a reference gene sequence                                                                                                                       |
| `reference_gene_stop` (optional)                  | Position of the last nucleotide in a reference sequence                                                                                                                             |
| `reference_protein_length` (optional)             | Length (number of positions) of a protein reference sequence retrieved from a database                                                                                              |
| `reference_protein_start` (optional)              | Position of the first amino acid in a reference protein sequence                                                                                                                    |
| `reference_protein_stop` (optional)               | Position of the last amino acid in a reference protein sequence                                                                                                                     |
| `resistance_mechanism` (optional)                 | Antibiotic resistance mechanisms evolve naturally via natural selection through random mutation, but it could also be engineered by applying an evolutionary stress on a population |
| `strand_orientation` (optional)                   | Orientation of a genomic element on the double-stranded molecule                                                                                                                    |
| `sequence_identity` (optional)                    | Sequence identity is the number (%) of matches (identical characters) in positions from an alignment of two molecular sequences                                                     |

</details>

[hAMRonization](https://github.com/pha4ge/hAMRonization) summarizes the outputs of the **antimicrobial resistance gene** detection tools (ABRicate, AMRFinderPlus, DeepARG, fARGene, RGI) into a single unified tabular format. It supports a variety of summary options including an interactive summary.

#### comBGC

<details markdown="1">
<summary>Output files</summary>

- `comBGC/`
  - `combgc_complete_summary.tsv`: summarised output from all BGC detection tools used in tsv format (all samples concatenated).
  - `*/combgc_summary.tsv`: summarised output from all applied BGC detection tools in tsv format for each sample.

</details>

<details markdown="1">
<summary>BGC summary table headers</summary>

| Table column      | Description                                                                                                 |
| ----------------- | ----------------------------------------------------------------------------------------------------------- |
| `Sample_ID`       | ID of the sample                                                                                            |
| `Prediction_tool` | BGC prediction tool (antiSMASH, DeepBGC, and/or GECCO)                                                      |
| `Contig_ID`       | ID of the contig containing the candidate BGC                                                               |
| `Product_class`   | Predicted BGC type/class                                                                                    |
| `BGC_probability` | Confidence of BGC candidate as inferred by the respective tool                                              |
| `BGC_complete`    | Whether BGC sequence is assumed to be complete or truncated by the edge of the contig                       |
| `BGC_start`       | Predicted BGC start position on the ontig                                                                   |
| `BGC_end`         | Predicted BGC end position on the contig                                                                    |
| `BGC_length`      | Length of the predicted BGC                                                                                 |
| `CDS_ID`          | ID of the coding sequence(s) (CDS) from the annotation step (prodigal/prokka/bakta) if provided by the tool |
| `CDS_count`       | Number of CDSs the BGC contains                                                                             |
| `PFAM_domains`    | Inferred PFAM IDs or annotations if provided by the tool                                                    |
| `MIBiG_ID`        | Inferred MIBiG IDs if provided by the tool                                                                  |
| `InterPro_ID`     | Inferred InterPro IDs if provided by the tool                                                               |

</details>

**comBGC** is a tool built for nf-core/funcscan which summarizes the results of the **Biosynthetic Gene Cluster (BGC)** prediction tools (antiSMASH, deepBGC, GECCO) used in the pipeline into one comprehensive tabular summary with standardised headers.

> ℹ️ comBGC does not feature `hmmer_hmmsearch` support. Please check the hmmsearch results directory.

#### MultiQC

<details markdown="1">
<summary>Output files</summary>

- `multiqc/`
  - `multiqc_report.html`: a standalone HTML file that can be viewed in your web browser
  - `multiqc_data/`: directory containing raw parsed data used for MultiQC report rendering
  - `multiqc_plots/`: directory containing any static images from the report in various formats

</details>

[MultiQC](http://multiqc.info) is used in nf-core/funcscan to report the versions of all software used in the given pipeline run, and provides a suggested methods text. This allows for reproducible analysis and transparency in method reporting in publications.

#### Pipeline information

<details markdown="1">
<summary>Output files</summary>

- `pipeline_info/`
  - Reports generated by Nextflow: `execution_report.html`, `execution_timeline.html`, `execution_trace.txt` and `pipeline_dag.dot`/`pipeline_dag.svg`.
  - Reports generated by the pipeline: `pipeline_report.html`, `pipeline_report.txt` and `software_versions.yml`. The `pipeline_report*` files will only be present if the `--email` / `--email_on_fail` parameter's are used when running the pipeline.
  - Reformatted samplesheet files used as input to the pipeline: `samplesheet.valid.csv`.

</details>

[Nextflow](https://www.nextflow.io/docs/latest/tracing.html) provides excellent functionality for generating various reports relevant to the running and execution of the pipeline. This will allow you to troubleshoot errors with the running of the pipeline, and also provide you with other information such as launch commands, run times and resource usage.
