BIB Staphylococcus aureus tutorial

Example of using BIB to identify Staphylococcus aureus strains. To run all the commands at once, install the required software and run the run_tutorial.sh script.

External software required to run the tutorial

1. progressiveMauve and stripSubsetLCBs.
2. hierBAPS
3. bowtie2
4. BitSeq
5. BIB

Tutorial

Steps 1.-6. can take up to an hour to run. The end result ref_seqs_gapless.fasta file is provided if you wish to skip the steps.

The example read file SRR016122.fastq.gz can be downloaded from the European Nucleotide Archive.

1. Obtain the full alignment of the assemblies.

   progressiveMauve --output=full_alignment.xmfa data/saur_assemblies.fasta

2. Extract LCBs shared by all genomes. The first number "500" is the minimum length of the LCB; the second number "4" indicates the minimum number of genomes that share an LCB.

   stripSubsetLCBs full_alignment.xmfa full_alignment.xmfa.bbcols core_alignment.xmfa 500 4

3. Concatenate all the LCBs.

   perl xmfa2fasta.pl --file core_alignment.xmfa > core_alignment.fasta

4. Run hierBAPS to obtain a clustering with "1" level and a maximum of "10" clusters. The clustering is stored in the results.partition.txt file. Sequences ">1" and ">2" belong to cluster 1; sequences ">3" and ">4" to cluster 2.

   hierBAPS.sh exData core_alignment.fasta fasta

   hierBAPS.sh hierBAPS seqs.mat 1 10 results

5. Select sequences ">1" and ">3" to represent the clusters using fastagrep.sh from the BitSeq package.

   fastagrep.sh ">1 " core_alignment.fasta > ref_seqs.fasta

   fastagrep.sh ">3 " core_alignment.fasta >> ref_seqs.fasta

6. Remove gaps from the reference sequences.

   sed 's/-//g' ref_seqs.fasta > ref_seqs_gapless.fasta

7. Build the alignment index.

   python BIB_prepare_index.py ref_seqs_gapless.fasta reference_alignment_index

8. Perform the read alignment and abundance estimation.

   python BIB_analyse_reads.py SRR016122.fastq.gz ref_seqs_gapless.fasta reference_alignment_index SRR016122_abundances