Advantages of long- and short-reads sequencing for the hybrid investigation of the Mycobacterium tuberculosis genome

Script for the analysis showed in the paper : https://doi.org/10.3389/fmicb.2023.1104456

Settings

1. Use the scripts/0_req.txt to build the conda environment for the analysis

2. Create a folder reads with three subfolders: LRS, SRS and Hybrid and put your .fastq files in the respective folders

3. Install MTBseq from https://github.com/ngs-fzb/MTBseq_source

4. Install Ratatosk from https://github.com/DecodeGenetics/Ratatosk

5. Install R with the packages tidyverse and rstatix

Generate Hybrid reads

From the main directory launch the script 1_Hybrid_generate-reads.sh

Running MTBseq

From the main directory launch the three scripts: 2_Hyb_mtbseq.sh 2_LRS_mtbseq.sh 2_SRS_mtbseq.sh

Genome Coverage

From the main directory launch the script: 3a_coverage.sh

Open R and and use the code in 3b_Coverage.R to compare the breadth coverage at 8x with the different approaches

Variant calling and Comparative Analysis

The variants were already called with the MTBseq step. They can be found in the relative folder (e.g reads/SRS/Called). From the main directory launch the script: 4_tree.sh This will produce a folder Trees containing the transmission trees of the samples according the different approaches.

For visualization use the site: https://achtman-lab.github.io/GrapeTree/MSTree_holder.html

Drug resistance

For each of the approaches enter the folder reads/*/Called. From there launch the script 5_WHO-resistance.py

The results will be in the file qcr26_all_full.csv

De Novo Assembly

From the main irectory launch the script 6a_Assembly.sh

The assemblies will be in the folder Assembly

For comparison and visualization use the R script 6b_assembly.R