cazomevolve 0.0.1

A bioinforamtic package for investigating the evolution of CAZomes

cazomevolve

Cazome Evolve (cazomevolve) invetigates the evolution of CAZomes by:

• searching for CAZy families that associated more than expected from their lineage
• projecting the CAZome composition onto a plot
• building a dendogram using distances calculated from the CAZome composition

This repo houses all scripts required for calculating, exploring and visually representing the covariance of CAZy family annotations within genomic assemblies.

Contents

1. Overview
2. Installation
   • Requirements
3. Current Developments
4. Directories

Overview

Carbohydrate Active enZymes are a subset of proteins that generate, modify and/or degrade carbohydrates. CAZy (www.cazy.org) is the most comprehensive CAZyme database, grouping proteins by sequence similarity into CAZy families. Cazy Family coVariance investigator (cfv_investigator) investigates the covariance of CAZy family annotations within proteomes across all species annotated by CAZy, and evaluates taxonomic specific covaraince of CAZy families.

cazomevolve is a bioinformatic package (still in development) for:

1. Retrieving of genomic accessions from which proteins catalogued within CAZy are derived from, and the associated taxonomic data of the source organism
2. Tracking frequency of CAZy family annotations for all genomic assemblies identified in step 1
3. Calculation of covariance of CAZy family annotations across all genomic assemblies identified in step 1, and for taxonomic specific groups (at the kingdom, genus and species taxonomic level)
4. Generating dataframes of the number of CAZymes per CAZy family for each genomic assembly
5. Generating a presence/absence matrix for each CAZy family in each genomic assembly

 

Installation

1. Create a virtual environment with dependencies, then activate the environment - where venv_name is an chosen name for the virtual environment conda create -n <venv_name> python=3.9
conda activate <venv_name>

2. Clone the repository git clone https://github.com/HobnobMancer/cazomevolve.git

3. Install pyrewton pip3 install -e <path to directory containing setup.py file>
   Do not forget to use the -e option when install using pip3, otherwise each time pyrewton is invoked a ModuleNotFound error will be raised. Pass the path to the directory containign the setup.py file not the path to the setup.py file; if you are currently in the root directory of the repoistory where the file is located, simply use '.' to indicate the current working directory.

 

Requirements

POISx or Mac OS, or linux emulator
Python version 3.8+
Miniconda3 or Anaconda managed microenvironment, incorporated code checkers are included in list form in 'requirements.txt'.
Miniconda3 environment file is also available in the GitHub repository: 'environment.yml'.
For all required Python libraries please read 'requirements.txt'.

• ncbi-genome-download
• saintBioutils

 

Method

Download genomes

Already have a list of genomic version accessions

If you already have a list of genomic version accessions in a plain text file, using the Python package ncbi-genome-download to download the genomic assemblies in .gbff (used to annotate the CAZome) and .fna (used to reconstruct the phylogenetic tree) formats - do not use the --flat-format option, leave the genomic assemblies compressed.

Retrieve all genomic assemblies associated with a specific term

To download load all genomic assemblies associated with a term of interest, such as Pectobacteriaceae (so as to download all Pectobacteriaceae assemblies), use the Python script cazomevolve/genomes/download_genomes.py. The script takes 4 required arguments:

1. User email address (required by NCBI)
2. The term of interest
3. The file formats to download the genomic assemblies in. Each file format is defined by its file extension. Separate file formats with a single comma, for example gbff,fna
4. Path to an output directory (this will be build by cazomevolve).

By default if the output directory exists, cazomevolve will crash. To write to an existing output directory use the -f/--force flag. By default, cazomevolve will delete all existing data in the existing output directory. To retain the data available in the existing output directory use the -n/--nodelete flag.

Extract protein seqs

Use the Python script cazomevolve/genomes/extract_prot_seqs.py to extract the protein sequences from annotations in the genomic assemblies.

One multisequence FASTA file is produced per genomic assembly, listing all protein sequences extracted from the respective genomic assembly.

Two position arguments are required:

1. Path to the directory containing the compressed genomic assemblies
2. Path to an output directory to write out all multisequence FASTA files

Reconstruct the phylogenetic tree

A baysian based approach

A distance based approach

Annotate the CAZomes

Option 1: Using cazomevolve and cazy_webscraper

Step 1: Using CAZy -- retrieve the canonical classifications

Use the Python script cazomevolve/cazome/cazy/get_cazy_cazymes.py, or the command cevolve_get_cazy_cazymes to retrieve the CAZy family classifications for proteins extracted from the genomic assemblies, and write two files of tab delimited list of the:

The required args are:

1. Path to the directory containing the FASTA protein sequences files
2. Path to the local CAZyme database compiled using cazy_webscraper
3. Path to an output directory to write out the protein sequences of proteins not listed in the local CAZyme database
4. Path to write out the tab delimited list of CAZy family annotations and associated genomic assembly accessions
5. Path to write out the tab delimited list of CAZy family annotations and associated genome and protein accessions

cazomevolve will build all necessary output directories.

Step 2: Using dbCAN --- retrieve predicted classifications

Use the Python script cazomevolve/cazome/invoke_dbcan.py to use dbCAN to predicte the CAZymes in each FASTA file of protein sequences.

2 positional arguments are required:

1. Input dir: path to directory containing all FASTA files of protein sequences
2. Output dir: path to write out all dbCAN output files. One subdir is created in the output dir for each FASTA file parsed by dbCAN

By default dbCAN version >= 3.0.4 is used (which uses HMMER, DIAMOND and eCAMI). To use dbCAN version 2.0.11 (which uses HMMER, DIAMOND and Hotpep) add the -V2 or --version_2 flag.

To extract the CAZy family predictions from dbCAN version 2 and/or 3, use the Python script cazomevolve/cazome/get_dbcan_cazymes.py, which will write out the CAZy family annotations to a tab delimited list.

Two positional arguments are required:

1. dbCAN dir: path to output dir from invoke_dbcan<num>.py
2. Path to write out tab delimited list - this may already exist and contain the CAZy family annotations from the local CAZyme database. The script will add the predicted CAZy family annotaitons from the dbCAN to the existing file. If a file does not already exist, a new file will be created.

Option 2: Using pyrewton and cazy_webscraper

You can use the Python package pyrewton to annotate the CAZome for a set of genomic assemblies, using cazy_webscraper and dbCAN [Zhange et al., 2018]. pyrewton compiles the canconical and predicted CAZyme classifications into a local SQLite3 database.

Han Zhang, Tanner Yohe, Le Huang, Sarah Entwistle, Peizhi Wu, Zhenglu Yang, Peter K Busk, Ying Xu, Yanbin Yin, dbCAN2: a meta server for automated carbohydrate-active enzyme annotation, Nucleic Acids Research, Volume 46, Issue W1, 2 July 2018, Pages W95–W101, https://doi.org/10.1093/nar/gky418

To retrieve the CAZy family annotations associated with each genomic assembly, execute the following sql command against the local CAZome database compiled using pyrewton:

Export the resulting table as a tsv file or tab delimited list.

Build the input for coinfinder

Find networks of co-evolving CAZy families

Build a presence/abensce and CAZy family number matrices

Build dendograms based upon CAZome compositions, and compare against the phylogenetic tree

Map genome and CAZome distances onto a plot

Directories

Below is a directory plan of this repository, followed by a brief overview of each directories role , to facilitate navigation through the repository.

assets

Directory containing all files needed for the GitHub page, created for easy access to accompanying Jupyter notebooks.

docs

Directory containing files to build documentation hosted at ReadTheDocs.

notebooks

Directory containing all Jupyter notebooks, and html copies used for easier in-browser viewing via the GitHub pages. These notebooks include the data outputs from using cfv_investigator, how to use the package and how the package works.

tests

Directory containing all pytest files for testing pyrewton, including subdirectories for test inputs and targets. Each module/submodule has its own specific test input and target subdirectory.

cazomevolve

Directory containing all pyrewton program modules (including all submodules and Python scripts).

 

Modules

Please find more detailed documentation at for operation and troubleshooting at Read the Docs

This is an overview of the functionalities of each module within pyrewton, as well as basics of operation. For more detailed documentation on the operation of each module and indiviudal Python scripts please see the documentation at Read the Docs

utilities

Contains all functions that are called from other Python scripts for building command-line parsers and loggers. Includes the submodule file_io, which contains functions that are called from other Python scripts for handling directories and files in pyrewton, including retrieving program inputs and creating output directories.

ncbi

Modules that are involved in retrieving handling data from NCBI. This includes retrieval of genomic accession numbers and source organism taxonomic data.

covariance

Modules that build the data set required for calculating the covariance, calculate the covariance of CAZy family annotations for all genomic accessions retrieved and taxonomic specific groups.