Dataset Information

PolyTB: a genomic variation map for Mycobacterium tuberculosis.

ABSTRACT: Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is the second major cause of death from an infectious disease worldwide. Recent advances in DNA sequencing are leading to the ability to generate whole genome information in clinical isolates of M. tuberculosis complex (MTBC). The identification of informative genetic variants such as phylogenetic markers and those associated with drug resistance or virulence will help barcode Mtb in the context of epidemiological, diagnostic and clinical studies. Mtb genomic datasets are increasingly available as raw sequences, which are potentially difficult and computer intensive to process, and compare across studies. Here we have processed the raw sequence data (>1500 isolates, eight studies) to compile a catalogue of SNPs (n = 74,039, 63% non-synonymous, 51.1% in more than one isolate, i.e. non-private), small indels (n = 4810) and larger structural variants (n = 800). We have developed the PolyTB web-based tool (http://pathogenseq.lshtm.ac.uk/polytb) to visualise the resulting variation and important meta-data (e.g. in silico inferred strain-types, location) within geographical map and phylogenetic views. This resource will allow researchers to identify polymorphisms within candidate genes of interest, as well as examine the genomic diversity and distribution of strains. PolyTB source code is freely available to researchers wishing to develop similar tools for their pathogen of interest.

SUBMITTER: Coll F

PROVIDER: S-EPMC4066953 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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PolyTB: a genomic variation map for Mycobacterium tuberculosis.

Coll Francesc F Preston Mark M Guerra-Assunção José Afonso JA Hill-Cawthorn Grant G Harris David D Perdigão João J Viveiros Miguel M Portugal Isabel I Drobniewski Francis F Gagneux Sebastien S Glynn Judith R JR Pain Arnab A Parkhill Julian J McNerney Ruth R Martin Nigel N Clark Taane G TG

Tuberculosis (Edinburgh, Scotland) 20140215 3

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is the second major cause of death from an infectious disease worldwide. Recent advances in DNA sequencing are leading to the ability to generate whole genome information in clinical isolates of M. tuberculosis complex (MTBC). The identification of informative genetic variants such as phylogenetic markers and those associated with drug resistance or virulence will help barcode Mtb in the context of epidemiological, diagnostic and clini ...[more]

PMID: 24637013

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Project description:Mycobacterium tuberculosis H37Ra is an attenuated tubercle bacillus closely related to the virulent type strain M. tuberculosis H37Rv. Despite extensive study, the reason for the decreased virulence of M. tuberculosis H37Ra has not been determined. A genomic approach was therefore initiated to identify genetic differences between M. tuberculosis H37Rv and M. tuberculosis H37Ra as a means of pinpointing the attenuating mutation(s). Digestion with the rare-cutting restriction endonuclease DraI revealed two polymorphisms between the strains: a 480-kb fragment in M. tuberculosis H37Rv was replaced by two fragments of 220 and 260 kb in M. tuberculosis H37Ra, while there was a approximately 7.9-kb DraI fragment in M. tuberculosis H37Ra that had no counterpart in M. tuberculosis H37Rv. As the M. tuberculosis insertion sequence IS6110 contains a single DraI restriction site, it was considered possible that these polymorphisms were the result of IS6110 transposition events in M. tuberculosis H37Ra, events that may have inactivated virulence genes. The 7.9-kb polymorphism was found to be due to the presence of the previously described H37Rv RvD2 deletion in M. tuberculosis H37Ra, with sequence analysis suggesting an IS6110-mediated deletion mechanism for loss of RvD2. Three other IS6110-catalyzed deletions from the M. tuberculosis H37Rv chromosome (RvD3 to RvD5) were also identified, suggesting that this mechanism plays an important role in genome plasticity in the tubercle bacilli. Comparative mapping and sequencing revealed that the 480-kb polymorphism was due to an IS6110 insertion in M. tuberculosis H37Ra near oriC. Complementation of M. tuberculosis H37Ra with a 2.9-kb restriction fragment from M. tuberculosis H37Rv that encompassed the IS6110 insertion did not increase the survival of recombinant M. tuberculosis H37Ra in mice. In conclusion, this study describes the presence and mechanisms of genomic variation between M. tuberculosis H37Ra and M. tuberculosis H37Rv, although the role that they play in the attenuation of M. tuberculosis H37Ra is unclear.

Project description:Mycobacterium tuberculosis complex (MTBC) species are classic examples of genetically monomorphic microorganisms due to their low genetic variability. Whole-genome sequencing made it possible to describe both the main species within the complex and M. tuberculosis lineages and sublineages. This differentiation is based on single nucleotide polymorphisms (SNPs) and large sequence polymorphisms in the so-called regions of difference (RDs). Although a number of studies have been performed to elucidate RD localizations, their distribution among MTBC species, and their role in the bacterial life cycle, there are some inconsistencies and ambiguities in the localization of RDs in different members of the complex. To address this issue, we conducted a thorough search for all possible deletions in the WGS data collection comprising 721 samples representing the full MTBC diversity. Discovered deletions were compared with a list of all previously described RDs. As with the SNP-based analysis, we confirmed the specificities of 79 regions at the species, lineage, or sublineage level, 17 of which are described for the first time. We also present RDscan (https://github.com/dbespiatykh/RDscan), an open-source workflow, which detects deletions from short-read sequencing data and correlates the results with high-specificity RDs, curated in this study. Testing of the workflow on a collection comprising ∼7,000 samples showed a high specificity of the found RDs. This study provides novel details that can contribute to a better understanding of the species differentiation within the MTBC and can help to determine how individual clusters evolve within various MTBC species. IMPORTANCE Reductive genome evolution is one of the most important and intriguing adaptation strategies of different living organisms to their environment. Mycobacterium offers several notorious examples of either naturally reduced (Mycobacterium leprae) or laboratory-reduced (Mycobacterium bovis BCG) genomes. Mycobacterium tuberculosis complex has its phylogeny unambiguously framed by large sequence polymorphisms that present unidirectional unique event changes. In the present study, we curated all known regions of difference and analyzed both Mycobacterium tuberculosis and animal-adapted MTBC species. For 79 loci, we have shown a relationship with phylogenetic units, which can serve as a marker for diagnosing or studying biological effects. Moreover, intersections were found for some loci, which may indicate the nonrandomness of these processes and the involvement of these regions in the adaptation of bacteria to external conditions.

Project description:BackgroundDue to excessive antibiotic use, drug-resistant Mycobacterium tuberculosis has become a serious public health threat and a major obstacle to disease control in many countries. To better understand the evolution of drug-resistant M. tuberculosis strains, we performed whole genome sequencing for 7 M. tuberculosis clinical isolates with different antibiotic resistance profiles and conducted comparative genomic analysis of gene variations among them.ResultsWe observed that all 7 M. tuberculosis clinical isolates with different levels of drug resistance harbored similar numbers of SNPs, ranging from 1409-1464. The numbers of insertion/deletions (Indels) identified in the 7 isolates were also similar, ranging from 56 to 101. A total of 39 types of mutations were identified in drug resistance-associated loci, including 14 previously reported ones and 25 newly identified ones. Sixteen of the identified large Indels spanned PE-PPE-PGRS genes, which represents a major source of antigenic variability. Aside from SNPs and Indels, a CRISPR locus with varied spacers was observed in all 7 clinical isolates, suggesting that they might play an important role in plasticity of the M. tuberculosis genome. The nucleotide diversity (Л value) and selection intensity (dN/dS value) of the whole genome sequences of the 7 isolates were similar. The dN/dS values were less than 1 for all 7 isolates (range from 0.608885 to 0.637365), supporting the notion that M. tuberculosis genomes undergo purifying selection. The Л values and dN/dS values were comparable between drug-susceptible and drug-resistant strains.ConclusionsIn this study, we show that clinical M. tuberculosis isolates exhibit distinct variations in terms of the distribution of SNP, Indels, CRISPR-cas locus, as well as the nucleotide diversity and selection intensity, but there are no generalizable differences between drug-susceptible and drug-resistant isolates on the genomic scale. Our study provides evidence strengthening the notion that the evolution of drug resistance among clinical M. tuberculosis isolates is clearly a complex and diversified process.

Dataset Information

PolyTB: a genomic variation map for Mycobacterium tuberculosis.

Publications

PolyTB: a genomic variation map for Mycobacterium tuberculosis.

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