Abstract
Because of its portable data, discriminatory power, and recently proposed standardization, mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing has become a major method for the epidemiological tracking of Mycobacterium tuberculosis complex (MTBC) clones. However, no public MIRU-VNTR database based on well-characterized reference strains has been available hitherto for easy strain identification. Therefore, a collection of 186 reference strains representing the primary MTBC lineages was used to build a database, which is freely accessible at http://www.MIRU-VNTRplus.org. The geographical origin and the drug susceptibility profile of each strain were stored together with comprehensive genetic lineage information, including the 24-locus MIRU-VNTR profile, the spoligotyping pattern, the single-nucleotide- and large-sequence-polymorphism profiles, and the IS6110 restriction fragment length polymorphism fingerprint. Thanks to flexible import functions, a single or multiple user strains can be analyzed, e.g., for lineage identification with or without the use of reference strains, by best-match or tree-based analyses with single or combined marker data sets. The results can easily be exported. In the present study, we evaluated the database consistency and various analysis parameters both by testing the reference collection against itself and by using an external population-based data set comprising 629 different strains. Under the optimal conditions found, lineage predictions based on typing by 24-locus MIRU-VNTR analysis optionally combined with spoligotyping were verified in >99% of the cases. On the basis of this evaluation, a user strategy was defined, which consisted of best-match analysis followed, if necessary, by tree-based analysis. The MIRU-VNTRplus database is a powerful tool for high-resolution clonal identification and has little equivalent in terms of functionalities among the bacterial genotyping databases available so far.
Highlights
The clonal population structure of Mycobacterium tuberculosis complex (MTBC), which comprises distinct phylogenetic lineages characterized by differences in their geographical distributions, immunogenicities, virulence, and associations with multidrug-resistant TB [10, 14, 19, 32, 37, 42, 47]
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Best matching is a method which, depending on the distance coefficient chosen, identifies reference genotypes with the closest distance to the test isolate. It is a simple parsimonylike method because it uses an optimality criterion, defined as a minimal number of differences in allelic profiles reflected by the calculated distances. This analysis was performed by using the categorical distance with 24-locus MIRU-VNTR typing data alone or in combination with spoligotyping data, while the SNP, LSP, and IS6110 RFLP profiles were used only as confirmatory markers
Summary
For the present database evaluation, all the different strains based on 24-locus MIRU-VNTR typing and spoligotyping, corresponding to unique or clustered genotypes, were retained (n ϭ 629) This panel was separated into two groups, with one containing preidentified lineage isolates (n ϭ 442) and the other containing nonpreidentified lineage isolates or phylogenetically poorly informative spoligotypes (i.e., T spoligotypes) (n ϭ 187). Dicted on the basis of the congruence of the MIRU-VNTR groupings and the corresponding spoligotypes determined by using the existing nomenclatures [9, 14] These predictions were tested as described in Results by comparison with the reference strains in the MIRU-VNTRplus database, whose lineages were identified by their RD/LSP and SNP profiles
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