Proposal of a Consensus Set of Hypervariable Mycobacterial Interspersed Repetitive-Unit–Variable-Number Tandem-Repeat Loci for Subtyping of Mycobacterium tuberculosis Beijing Isolates

Abstract

q Mycobacterium tuberculosis Beijing strains represent targets of special importance for molecular surveillance of tuberculosis (TB), especially because they are associated with spread of multidrug resistance in some world regions. Standard 24-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing lacks resolution power for accurately discriminating closely related clones that often compose Beijing strain populations. Therefore, we evaluated a set of 7 additional, hypervariable MIRU-VNTR loci for better resolution and tracing of such strains, using a collection of 535 Beijing isolates from six world regions where these strains are known to be prevalent. The typeability and interlaboratory reproducibility of these hypervariable loci were lower than those of the 24 standard loci. Three loci (2163a, 3155, and 3336) were excluded because of their redundant variability and/or more frequent noninterpretable results compared to the 4 other markers. The use of the remaining 4-locus set (1982, 3232, 3820, and 4120) increased the number of types by 52% (from 223 to 340) and reduced the clustering rate from 58.3 to 36.6%, when combined with the use of the standard 24-locus set. Known major clonal complexes/ 24-locus-based clusters were all subdivided, although the degree of subdivision varied depending on the complex. Only five single-locus variations were detected among the hypervariable loci of an additional panel of 92 isolates, representing 15 years of clonal spread of a single Beijing strain in a geographically restricted setting. On this calibrated basis, we propose this 4-locus set as a consensus for subtyping Beijing clonal complexes and clusters, after standard typing.

A four-year nationwide molecular epidemiological study in Estonia: Risk factors for tuberculosis transmission

“…Несмотря на высокую дискриминирующую способность, 24-х локусное VNTR-типи-рование не обладает достаточным разрешением для точного разли-чения близко связанных клонов, составляющих популяцию штамма Beijing, по сравнению с IS6110-RFLP или с использованием дополни-тельных гипервариабельных VNTR локусов. 8,28 Таким образом наша оценка недавней трансмиссии может быть слегка завышенной.…”

“…Сполиготипирование, которое имеется, но в на-стоящее время не используется в Эстонии, и дополнительное исполь-зование VNTR-типирования гипервариабельных локусов изолятов Beijing позволило бы с большей точностью определить принадлеж-ность к семейству и обеспечило бы точную дискриминацию случаев внутри кластера. 28,35 Однако, как уже говорилось выше, проведенный нами анализ факторов риска формирования кластеров позволяет с большой долей уверенности судить о эпидемиологической значимо-сти выделенных кластеров.…”

“…Spoligotyping, which is available but not currently in use in Estonia, and additional use of VNTR hypervariable loci on Beijing isolates would have allowed a degree of independent confirmation of lineages and more precise discrimination of cases within clusters, respectively. 28,35 However, as indicated above, our results from the analysis of risk factors for clustering allows a good degree of confidence in the epidemiological relevance of the clusters identified.…”

“…Although highly discriminatory, 24-locus VNTR typing lacks resolution power for accurately discriminating closely related clones composing Beijing strain populations in comparison with IS6110-RFLP or with the use of additional hypervariable VNTR loci. 8,28 There may thus be some minimal overestimation in our assessment of recent transmission. Third, we found certain socio-demographic and clinical factors that were significantly associated with clustering, indicating that the clustering detected reflects recent transmission to a good extent.…”

Implementation of a Consensus Set of Hypervariable Mycobacterial Interspersed Repetitive-Unit–Variable-Number Tandem-Repeat Loci in Mycobacterium tuberculosis Molecular Epidemiology

“…It is considered the international standard for molecular typing (3,4), sometimes supplemented by spoligotyping (5). Despite its remarkable advantages in terms of technical simplicity, reproducibility, and portability compared with IS6110 restriction fragment length polymorphism (RFLP) analysis (6), several studies have highlighted the relative lack of resolution power of MIRU-VNTR analysis (7), particularly for highly clonal strain groups, such as the Beijing lineage (8,9).…”