This is the experimental commandline version of the TBProfiler described here: https://genomemedicine.biomedcentral.com/articles/10.1186/s13073-015-0164-0 and is available here: http://tbdr.lshtm.ac.uk
This repository contains a complete rewrite of the web version of TBProfiler. It allows the use of profiling through a command line interface and contains some additional functionality such as the ability to process minION data.
The pipeline aligns reads to the H37Rv reference BWA or minimap2 and then looks at the coverage across a number of different candidate regions. We also predict the number of reads supporting drug resistance variants as an insight into hetero-resistance (not applicable for minION data)
Version 0.3.0 features many changes to the code including the modularisation through creation of specific classes and functions for various functions. This will make it easier to maintain and add additional functionality. Support for minION has also been added. In the process some old functionality has not been added yet and some required arguments have changed. Please download v0.2.1 from the releases if you require the old code.
git clone --recursive https://github.com/jodyphelan/TBProfiler.git
cd TBProfiler
bash install_prerequisites.sh
echo "export PATH=$PWD:$PATH" >> ~/.bashrc
For OSX use:
bash osX_install_prerequisites.sh
The first argument indicates the analysis type to perform. At the moment we currently only support the calling of small variants or the detection of large deletions.
Run whole pipeline:
tb-profiler profile -1 /path/to/reads_1.fastq.gz -2 /path/to/reads_2.fastq.gz -p prefix
The prefix is usefull when you need to run more that one sample. This will store BAM files, summary pileup and result files in respective directories. Results are output in text format and json format.
Example run:
mkdir test_run; cd test_run
wget ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR166/009/ERR1664619/ERR1664619_1.fastq.gz
wget ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR166/009/ERR1664619/ERR1664619_2.fastq.gz
../tb-profiler profile -1 ERR1664619_1.fastq.gz -2 ERR1664619_2.fastq.gz -t 4 -p ERR1664619
cat results/ERR1664619.results.txt
By using the -a option you can specify to use an existing BAM file instead of fastq files.
Warning!!!: The BAM files must have been created using the ensembl version of the genome which can be downloaded here:
ftp://ftp.ensemblgenomes.org/pub/release-32/bacteria//fasta/bacteria_0_collection/mycobacterium_tuberculosis_h37rv/dna/Mycobacterium_tuberculosis_h37rv.ASM19595v2.dna.toplevel.fa.gz
The results from numerous runs can be collated into one table using the following command:
tb-profiler collate samples_file out_file
Where samples_file is a list of prefixes of previously run samples and out_file is the name of the output file.
The pipeline searches for small variants and big deletions associated with drug resistance. It will also report the lineage.
To add new mutations navigate to the db directory and edit the drdb.txt file.
Add a new line corresponding to the desired variant with the following columns:
- Drug - Drug name with no spaces
- Genomic position - If more than one position affected, seperate with "/".
- Reference nucleotides - String of nucleotides with length equal to the number of bases affected.
- Alternate nucleotides - String of nucleotides with length equal to the number of bases affected.
- Gene name
- Mutation - String with the mutation
After editing the file run the parse_drdb.py <prefix> script using a prefix to generate a new database.
To use the database use the --db <prefix> option in tb-profiler.
ETHIONAMIDE 1674484/1674485 AT CC inhA Ile95Pro
ISONIAZID 2156118 C T katG_promoter C-7T
PYRAZINAMIDE 2288953 CC C pncA CC289C
Several files are produced by the tb-profile collate function. Among these are several config files that can be used with iTOL (http://itol.embl.de/) to annotate phylogenetic trees. A small tree and config files have been placed in the example_data directory. To use navigate to the iTOL website and upload the tbprofiler.tree file using the upload button on the navigation bar. Once this has been uploaded you will be taken to a visualisation of the tree. To add the annotation, click on the '+' button on the lower right hand corner and select the iTOL config files. You should now see a figure similar to the one below. The following annotations are included:
- Lineage
- Drug resistance classes (Sensitive, drug-resistant, MDR, XDR)
- Drug resistance calls for individual drugs, were filled circles represent resistance.
Modularisation of code into classes and functions Support for minION
- Collate data fix
- Generation of ITOL data files for visualisation
- Allow for the choice of BWA or SNAP for Linux users
- Calling of deletions before small variant calling to avoid low quality variants around deletion breakpoints
- Allow users to provide BAM file as input
- Ability to print out version
If you would like to cite this work please use:
Coll, F. et al. Rapid determination of anti-tuberculosis drug resistance from whole-genome sequences. Genome Med. 5:51, (2015).