Abstract
BackgroundWhole-genome sequencing is increasingly used in clinical diagnosis of tuberculosis and study of Mycobacterium tuberculosis complex (MTC). MTC consists of several genetically homogenous mycobacteria species which can cause tuberculosis in humans and animals. Regions of difference (RDs) are commonly regarded as gold standard genetic markers for MTC classification.ResultsWe develop RD-Analyzer, a tool that can accurately infer the species and lineage of MTC isolates from sequence reads based on the presence and absence of a set of 31 RDs. Applied on a publicly available diverse set of 377 sequenced MTC isolates from known major species and lineages, RD-Analyzer achieved an accuracy of 98.14 % (370/377) in species prediction and a concordance of 98.47 % (257/261) in Mycobacterium tuberculosis lineage prediction compared to predictions based on single nucleotide polymorphism markers. By comparing respective sequencing read depths on each genomic position between isolates of different sublineages, we were able to identify the known RD markers in different sublineages of Lineage 4 and provide support for six potential delineating markers having high sensitivities and specificities for sublineage prediction. An extended version of RD-Analyzer was thus developed to allow user-defined RDs for lineage prediction.ConclusionsRD-Analyzer is a useful and accurate tool for species, lineage and sublineage prediction using known RDs of MTC from sequence reads and is extendable to accepting user-defined RDs for analysis. RD-Analyzer is written in Python and is freely available at https://github.com/xiaeryu/RD-Analyzer.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3213-1) contains supplementary material, which is available to authorized users.
Highlights
Whole-genome sequencing is increasingly used in clinical diagnosis of tuberculosis and study of Mycobacterium tuberculosis complex (MTC)
It has been suggested that one of the best strategies for MTC genotyping is using MIRU-VNTR combined with spoligotyping, which can differentiate between clinical isolates to identify disease transmission and outbreak, distinguish between disease relapse and re-infection, and identify contamination [8, 9]
Description of algorithm Region of difference (RD)-Analyzer is written in Python and can be used to accurately determine the presence or absence of 31 informative RD markers from raw sequence reads in order to infer the species and lineages of MTC isolates
Summary
Whole-genome sequencing is increasingly used in clinical diagnosis of tuberculosis and study of Mycobacterium tuberculosis complex (MTC). MTC consists of several genetically homogenous mycobacteria species which can cause tuberculosis in humans and animals. Mycobacterium tuberculosis complex (MTC) is the causal agent of TB, which comprises of several genetically homogenous mycobacteria species including human-adapted pathogens of Mycobacterium tuberculosis (Mtb), M. africanum, M. canettii, M. bovis and animal-adapted pathogens of M. caprae, M. microti and M. pinnipedii which have been reported to cause human. Several studies have shown that the rapidly evolving genetic markers used in MIRU-VNTR and spoligotyping, highly discriminatory, are prone to homoplasy or convergent evolution, where the same genetic profile could be obtained in distinct MTC strains that are phylogenetically unrelated, confounding strain classification and phylogenetic inference [10,11,12]
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