-- ivr locus allele determination from genomic data --
The ivr locus, also known as cod locus, is a random six-phase switch in a Type I restriction-modification system. It has been shown to be the responsible underlying mechanism for phase-variation in streptococcus pneumonia (Manso et al, 2014).
There are six possible alleles, identified from A to E:
- Allele A (modules 1.1 and 2.1)
- Allele B (modules 1.2 and 2.2)
- Allele E (modules 1.2 and 2.3)
- Allele D (modules 1.2 and 2.1)
- Allele C (modules 1.2 and 2.2)
- Allele F (modules 1.2 and 2.3)
Due to the high variation rate and structural rearrangement, this allele cannot reliably be determined using assembly or standard mapping approaches.
Therefore, we present ivrTyper, a standalone tool for the determination of the ivr locus allele from paired-end genomic data.
We've adapted the algorithm described by Lee et al (2017). The reads are mapped to the full iver locus and the mates of reads mapping to the reverse strand of the conserved 5' region are extracted for each sample and mapped with bowtie2 to the 1.1 and 1.2 modules. The representative module of the hsdS gene is chosen if the number of reads mapping is greater than the --proportionCutOff (default is 80%). The 3' allele is determined using the chosen 1.x module as target, and mapping the mates of reads to the module 2.x reference sequences.
For a module to be considered present it needs to have a number of reads mapped higher than --minCoverage.
The number of mates that map to the 1.x module is obtained though the conserved 5' region. The number of mates mapping to the 2.x module is obtained though the selected 1.x module. The calculation of the proportion takes into consideration the reads mapping to all 2.x possibilities only.
Required to download sequence data from ENA database:
- Aspera Connect 2 >= v3.6.1
Required to run analysis:
- Bowtie2 >= v2.2.9
- Samtools = v1.3.1
(these three executables are provided, but user's own executables can be used by providing --doNotUseProvidedSoftware option)
- pysam >= 0.11.2.2 (available though pip)
- python 2.7
git clone https://github.com/cimendes/ivrTyper.git
usage: ivrTyper.py [-h] [--version] -w /path/to/workdir/directory/ [-j N] [-u]
[-k] [-m N] [-c N] [-gt N] [-rf /path/to/run.*.log]
[-a /path/to/asperaweb_id_dsa.openssh] [-kd] [-ip ILLUMINA]
[-pm HiSeq] [-ls GENOMIC]
[-l /path/to/list_IDs.txt | -t "Streptococcus pneumoniae"]
Reads mapping against pneumococcal ivr locus for typing.
optional arguments:
-h, --help show this help message and exit
--version Version information
-l /path/to/list_IDs.txt, --listIDs /path/to/list_IDs.txt
Path to list containing the IDs to be downloaded (one
per line) (default: None)
-t "Streptococcus pneumoniae", --taxon "Streptococcus pneumoniae"
Taxon name for which fastq files will be downloaded
(default: None)
General options:
-w /path/to/workdir/directory/, --workdir /path/to/workdir/directory/
Path to the directory containing the read files in
subdirectories, one per sample (fastq.gz or fq.gz and
pair-end direction coded as _R1_001 / _R2_001 or _1 /
_2) or to be downloaded (default: None)
-j N, --threads N Number of threads to use (default: 1)
-u, --skipProvidedSoftware
Do not use provided software (default: False)
-k, --keepFiles Keep alignment files (default: False)
ivrTyper module facultative options:
-m N, --minCoverage N
minimumcoverage depth a module to be present in the
sample (default: 5)
-c N, --proportionCutOff N
Proportion cut off for the module 1.x to be chosen as
target (default: 0.8)
-gt N, --greaterThan N
proportion cut off for the "greater than" column in
the final report. (default: 0.5)
-rf /path/to/run.*.log, --reportFile /path/to/run.*.log
Logfile to append run information to. (default: None)
Download facultative options:
-a /path/to/asperaweb_id_dsa.openssh, --asperaKey /path/to/asperaweb_id_dsa.openssh
Download fastq files from ENA using Aspera Connect.
With this option, the path to Private-key file
asperaweb_id_dsa.openssh must be provided (normaly
found in
~/.aspera/connect/etc/asperaweb_id_dsa.openssh).
(default: None)
-kd, --keepDownloadFiles
Keep downloaded read files (default: False)
-ip ILLUMINA, --instrumentPlatform ILLUMINA
Download files with specific library layout (available
options: ILLUMINA, ALL) (default: ALL)
-pm HiSeq, --platformModel HiSeq
Filter download by the model of the Illumina machine.
(default: None)
-ls GENOMIC, --librarySource GENOMIC
Filter download by library source (available options:
GENOMIC, ALL) (default: None)
To run ivrTyper in local fastq files, these need to be organized in sample folders.
It is advisable to use copied fastq files or symbolic links to the original ones. Then provide the directory containing sample folders to --workdir. The output report will be stored there.
workir/
sample_1/
fastq_file_a_1.fq.gz
fastq_file_a_2.fq.gz
sample_2/
fastq_file_b_R1_001.fastq.gz
fastq_file_b_R2_001.fastq.gz
ivrTyper.py --workdir /path/to/data/ --threads 8
To run ivrTyper in a specific set of ENA IDs, provide a file to --listIDs containing a list of ENA IDs that will be downloaded. This tool requires the genomic data to be paired-end and from Illumina technology.
ReMatCh will store the output files in the --workdir.
ivrTyper.py --listIDs /path/to/list_IDs.txt --workdir /path/to/output/directory/ --threads 8
To run ivrTyper in all ENA data of a given taxon, provide the taxon name to --taxon.
The ENA Run Accession numbers for the given taxon will be stored in IDs_list.seqFromWebTaxon.tab file.
Warning - This is option is in development.
ivrTyper.py --taxon "Streptococcus pneumoniae" --workdir /path/to/output/directory/ --threads 8
run.*.log ivrTyper running log file.
report_*.csv Report for the samples that ran the ivrTyper successfully.
- Sample - Sample name
- 1.1 - Number of reads that mapped to the 1.1 module
- 1.2 - Number of reads that mapped to the 1.2 module
- 2.1 - Number of reads that mapped to the 2.1 module, using the propper 1.x module as target
- 2.2 - Number of reads that mapped to the 2.2 module, using the propper 1.x module as target
- 2.3 - Number of reads that mapped to the 2.3 module, using the propper 1.x module as target
- pA - proportion of reads that map to the allele A
- pB - proportion of reads that map to the allele B
- pE - proportion of reads that map to the allele E
- pD - proportion of reads that map to the allele D
- pC - proportion of reads that map to the allele C
- pF - proportion of reads that map to the allele F
- Most Prevalent - Most prevalent ivr type in the sample
- gt0.5 - ivr type with a proportion greater than 0.5 (or whatever is set by
--greaterThan)
Lees, J.A., Kremer, P.H.C., Manso, A.S., Croucher, N.J., Ferwerda, B., Serón, M.V., Oggioni, M.R., Parkhill, J., Brouwer, M.C., Ende, A. Van Der, Beek, D. Van De, Bentley, S.D., 2017. Large scale genomic analysis shows no evidence for pathogen adaptation between the blood and cerebrospinal fluid niches during bacterial meningitis. Microb. Genomics 3, 1–12. doi:10.1099/mgen.0.000103
Li, J., Li, J.W., Feng, Z., Wang, J., An, H., Liu, Y., Wang, Y., Wang, K., Zhang, X., Miao, Z., Liang, W., Sebra, R., Wang, G., Wang, W.C., Zhang, J.R., 2016. Epigenetic Switch Driven by DNA Inversions Dictates Phase Variation in Streptococcus pneumoniae. PLoS Pathog. 12, 1–36. doi:10.1371/journal.ppat.1005762
Manso, A.S., Chai, M.H., Atack, J.M., Furi, L., De Ste Croix, M., Haigh, R., Trappetti, C., Ogunniyi, A.D., Shewell, L.K., Boitano, M., Clark, T.A., Korlach, J., Blades, M., Mirkes, E., Gorban, A.N., Paton, J.C., Jennings, M.P., Oggioni, M.R., 2014. A random six-phase switch regulates pneumococcal virulence via global epigenetic changes. Nat. Commun. 5, 5055. doi:10.1038/ncomms6055
Catarina Mendes [email protected]