Spae

Phage toolkit to detect phage candidates for phage therapy

Overview

This snakemake workflow was built using Snaketool [https://doi.org/10.1371/journal.pcbi.1010705], to assemble and annotate phage sequences. Currently this tool is being developed for phage genomes. The steps include,

• Quality control that removes adaptor sequences, low quality reads and host contimanination (optional).
• Assembly
• Contig quality checks; read coverage, viral or not, completeness, and assembly graph components.
• Phage genome annotation'
• Annotation of the phage genome

Complete list of programs used for each step is mention in the phagegenomeassembly.CITATION file.

Install

Pre-requisites
- gcc - conda - libgl1-mesa-dev (ubuntu- for Bandage) - libxcb-xinerama0 (ubuntu- for Bandage)

Install Setting up a new conda environment

conda create -n spae python=3.11
conda activate spae
conda install -n base -c conda-forge mamba #if you dont already have mamba installed

Steps for installing spae workflow

git clone https://github.com/linsalrob/spae.git
cd spae
pip install -e .
#confirm the workflow is installed by running the below command 
spae --help

Installing databases

Run command,

spae install

Install the databases to a directory, phagegenomeassembly/workflow/databases

This workflow requires the - Pfam35.0 database to run viral_verify for contig classification. - CheckV database to test for phage completeness - Pharokka databases - Phyteny models

This step takes approximately 1hr 30min to install

Running the workflow

The command spae run will run QC, assembly and annoation

Commands to run Only one command needs to be submitted to run all the above steps: QC, assembly and assembly stats

#For illumina reads, place the reads both forward and reverse reads to one directory
spae run --input test/illumina-subset --output example

#For nanopore reads, place the reads, one file per sample in a directory
spae run --input test/nanopore-subset --sequencing longread --output example 

#To run either of the commands on the cluster, add --profile slurm to the command. For instance here is the command for longreads/nanopore reads 
#Before running this below command, makse sure have slurm config files setup, here is a tutorial, https://fame.flinders.edu.au/blog/2021/08/02/snakemake-profiles-updated 
spae run --input test/nanopore-subset --preprocess longread --output example --profile slurm 

Output - Assmbled phage genome saved to "{outut-directory}/genome/{sample}/{sample}.fasta - Annotations of the phage genome are saved to "{outut-directory}/pharokka/phynteny/phynteny.gbk"

Steps to work on

Phage aspect of the code - subsampling a frgamented genome and assembling again - make output as subsampled - if fragmented - keep but mark as fragmented output - add anti-immunity gene search with pharokka - add final report as output with candidates for phage therapy

bacteria aspect of the code - support bacterial host assembly and annotation - search for immunity proteins

Issues and Questions

This is still a work in progress, so if you come across any issues or errors, report them under Issues.