COGclassifier

Table of Contents

• Overview
• Installation
• Workflow
• Usage
• Output Contents
• Customize Charts

Overview

COG(Cluster of Orthologous Genes) is a database that plays an important role in the annotation, classification, and analysis of microbial gene function. Functional annotation, classification, and analysis of each gene in newly sequenced bacterial genomes using the COG database is a common task. However, there was no COG functional classification command line software that is easy-to-use and capable of producing publication-ready figures. Therefore, I developed COGclassifier to fill this need. COGclassifier can automatically perform the processes from searching query sequences into the COG database, to annotation and classification of gene functions, to generation of publication-ready figures (See figure below).

Fig.1: Barchart of COG funcitional category classification result for E.coli

Fig.2: Piechart of COG funcitional category classification result for E.coli

Installation

COGclassifier is implemented in Python3. RPS-BLAST(v2.13.0) binary is bundled in COGclassifier (src/cogclassifier/bin).

Install bioconda package:

conda install -c conda-forge -c bioconda cogclassifier

Install PyPI stable package:

pip install cogclassifier

Workflow

Description of COGclassifier's automated workflow. This workflow was created based in part on cdd2cog.

1. Download COG & CDD resources

Download 4 required COG & CDD files from FTP site.

• fun-20.tab (https://ftp.ncbi.nih.gov/pub/COG/COG2020/data/fun-20.tab)
  Descriptions of COG functional categories.

  Show more information

  Tab-delimited plain text file with descriptions of COG functional categories
  Columns:

  1. Functional category ID (one letter)
  2. Hexadecimal RGB color associated with the functional category
  3. Functional category description
  Each line corresponds to one functional category. The order of the categories is meaningful (reflects a hierarchy of functions; determines the order of display)

  (From https://ftp.ncbi.nih.gov/pub/COG/COG2020/data/Readme.2020-11-25.txt)

• cog-20.def.tab (https://ftp.ncbi.nih.gov/pub/COG/COG2020/data/cog-20.def.tab)
  COG descriptions such as 'COG ID', 'COG functional category', 'COG name', etc...

  Show more information

  Tab-delimited plain text file with COG descriptions
  Columns:

  1. COG ID
  2. COG functional category (could include multiple letters in the order of importance)
  3. COG name
  4. Gene associated with the COG (optional)
  5. Functional pathway associated with the COG (optional)
  6. PubMed ID, associated with the COG (multiple entries are semicolon-separated; optional)
  7. PDB ID of the structure associated with the COG (multiple entries are semicolon-separated; optional)
  Each line corresponds to one COG. The order of the COGs is arbitrary (displayed in the lexicographic order)

  (From https://ftp.ncbi.nih.gov/pub/COG/COG2020/data/Readme.2020-11-25.txt)

• cddid.tbl.gz (https://ftp.ncbi.nih.gov/pub/mmdb/cdd/)
  Summary information about the CD(Conserved Domain) model.

  Show more information

  "cddid.tbl.gz" contains summary information about the CD models in this distribution, which are part of the default "cdd" search database and are indexed in NCBI's Entrez database. This is a tab-delimited text file, with a single row per CD model and the following columns:

  PSSM-Id (unique numerical identifier)
  CD accession (starting with 'cd', 'pfam', 'smart', 'COG', 'PRK' or "CHL')
  CD "short name"
  CD description
  PSSM-Length (number of columns, the size of the search model)

  (From https://ftp.ncbi.nih.gov/pub/mmdb/cdd/README)

• Cog_LE.tar.gz (https://ftp.ncbi.nih.gov/pub/mmdb/cdd/little_endian/)
  COG database, a part of CDD(Conserved Domain Database), for RPS-BLAST search.

2. RPS-BLAST search against COG database

Run query sequences RPS-BLAST against COG database [Default: E-value = 1e-2]. Best-hit (=lowest e-value) blast results are extracted and used in next functional classification step.

3. Classify query sequences into COG functional category

From best-hit results, extract relationship between query sequences and COG functional category as described below.

1. Best-hit results -> CDD ID
2. CDD ID -> COG ID (From cddid.tbl)
3. COG ID -> COG Functional Category Letter (From cog-20.def.tab)
4. COG Functional Category Letter -> COG Functional Category Definition (From fun-20.tab)

⚠️ If functional category with multiple letters exists, first letter is treated as functional category (e.g. COG4862 has multiple letters KTN. A letter K is treated as functional category).

Using the above information, the number of query sequences classified into each COG functional category is calculated and functional annotation and classification results are output.

Usage

Basic Command

COGclassifier -i [query protein fasta file] -o [output directory]

Options

-i I, --infile I      Input query protein fasta file
-o O, --outdir O      Output directory
-d , --download_dir   Download COG & CDD resources directory (Default: '~/.cache/cogclassifier')
-t , --thread_num     RPS-BLAST num_thread parameter (Default: MaxThread - 1)
-e , --evalue         RPS-BLAST e-value parameter (Default: 1e-02)
-v, --version         Print version information
-h, --help            Show this help message and exit

Example Command

Classify E.coli protein sequences into COG functional category (ecoli.faa):

COGclassifier -i ./example/input/ecoli.faa -o ./ecoli_cog_classifier

Example API

from cogclassifier import cogclassifier

query_fasta_file = "./example/input/ecoli.faa"
outdir = "./ecoli_cog_classifier"
cogclassifier.run(query_fasta_file, outdir)

Output Contents

COGclassifier outputs 4 result text files, 3 html format chart files.

• rpsblast_result.tsv (example)
  RPS-BLAST against COG database result (format = outfmt 6).

• classifier_result.tsv (example)
  Query sequences classified into COG functional category result.
  This file contains all classified query sequences and associated COG information.

  Table of detailed tsv format information (9 columns)

  Columns	Contents	Example Value
  QUERY_ID	Query ID	NP_414544.1
  COG_ID	COG ID of RPS-BLAST top hit result	COG0083
  CDD_ID	CDD ID of RPS-BLAST top hit result	223161
  EVALUE	RPS-BLAST top hit evalue	2.5e-150
  IDENTITY	RPS-BLAST top hit identity	45.806
  GENE_NAME	Abbreviated gene name	ThrB
  COG_NAME	COG gene name	Homoserine kinase
  COG_LETTER	Letter of COG functional category	E
  COG_DESCRIPTION	Description of COG functional category	Amino acid transport and metabolism

• classifier_count.tsv (example)
  Count classified sequences per COG functional category result.

  Table of detailed tsv format information (4 columns)

  Columns	Contents	Example Value
  LETTER	Letter of COG functional category	J
  COUNT	Count of COG classified sequence	259
  COLOR	Symbol color of COG functional category	#FCCCFC
  DESCRIPTION	Description of COG functional category	Translation, ribosomal structure and biogenesis

• classifier_stats.txt (example)
  The percentages of the classified sequences are described as example below.

  86.35% (3575 / 4140) sequences classified into COG functional category.

• classifier_count_barchart.html
  Barchart of COG funcitional category classification result.
  COGclassifier uses Altair visualization library for plotting html format charts.
  In web browser, Altair charts interactively display tooltips and can export image as PNG or SVG format.

  

• classifier_count_piechart.html
  Piechart of COG funcitional category classification result.
  Functional category with percentages less than 1% don't display letter on piechart.

  

• classifier_count_piechart_sort.html
  Piechart with descending sort by count.
  Functional category with percentages less than 1% don't display letter on piechart.

  

Customize Charts

COGclassifier also provides barchart & piechart plotting scripts to customize charts appearence. Each script can plot the following feature charts from classifier_count.tsv. See wiki for details.

• Features of plot_cog_classifier_barchart script (wiki)

  • Adjust figure width, height, barwidth
  • Plot charts with percentage style instead of count number style
  • Fix maximum value of Y-axis
  • Descending sort by count number or not
  • Plot charts from user-customized 'classifier_count.tsv'

• Features of plot_cog_classifier_piechart script (wiki)

  • Adjust figure width, height
  • Descending sort by count number or not
  • Show letter on piechart or not
  • Plot charts from user-customized 'classifier_count.tsv'