Use of Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeat Typing To Examine Genetic Diversity ofMycobacterium tuberculosisin Singapore

Abstract

Strain typing using variable-number tandem repeats of mycobacterial interspersed repetitive units (MIRU-VNTR) is a powerful tool for studying the epidemiology and genetic relationships of Mycobacterium tuberculosis isolates. For this study, isolates from 291 patients in Singapore were genotyped by this method. One hundred sixty-six distinct MIRU-VNTR patterns were detected. One hundred sixty-two strains were grouped into 1 of 35 different MIRU-VNTR clusters and 131 isolates were unique. In this sample collection, 9 of the 12 MIRU-VNTR loci were moderately or highly discriminative according to their allelic diversities. The HunterGaston discriminatory index was 0.975, indicating the high power of discrimination of MIRU-VNTR typing. By direct comparisons with previously typed MIRU-VNTR patterns and by genetic relationship analyses, we could identify and clearly define four epidemic groups of M. tuberculosis in our sample, corresponding to the W/Beijing, East-Africa-Indian, Haarlem, and Delhi genotype families. Furthermore, MIRU-VNTR typing was able to clearly distinguish ancestral and modern M. tuberculosis strains as defined by TbD1 genomic deletion analysis. These results indicate that MIRU-VNTR typing can be a useful first-line tool for studying the genetic diversity of M. tuberculosis isolates in a large urban setting such as Singapore.Tuberculosis remains a major infectious disease and causes high morbidity and mortality worldwide. Mycobacterium tuberculosis genotyping is a useful tool for the epidemiological surveillance and control of disease transmission. Although a large number of DNA-fingerprinting methods for typing M. tuberculosis isolates have been developed (15), only IS6110 restriction fragment length polymorphism (RFLP) typing (5) and spacer oligonucleotide typing (spoligotyping) (14) have gained wide acceptance.IS6110 typing is the most commonly used strain-typing method for M. tuberculosis. This method is based on variations among IS6110 insertion elements between strains. IS6110 typing has been shown to be reproducible and highly discriminatory (15). However, there are several drawbacks of IS6110 typing. Firstly, it has poor discriminatory power for M. tuberculosis strains with low copy numbers of IS6110 (7,15,17). Secondly, it requires relatively large quantities of purified DNA. Thirdly, it is time-consuming to perform and analyze. Finally, although a standardized methodology has been recommended (29), it is difficult to compare IS6110 RFLP results between laboratories. This hinders global studies of M. tuberculosis epidemiology and transmission. Spoligotyping is a PCRbased method which detects the presence or absence of 43 spacers in the direct repeat locus of the M. tuberculosis genome. This has advantages over IS6110 typing in that it is faster and easier to perform, it is reproducible, and it requires only small quantities of DNA. However, the major drawback of spoligotyping is that it has a lower discriminatory power than IS6110 typing. In particular, this method is not informative in are...

Comparison of spoligotyping, mycobacterial interspersed repetitive units typing and IS6110-RFLP in a study of genotypic diversity of Mycobacterium tuberculosis in Delhi, North India

“…However, in our study, locus 31 was equally discriminatory with an allelic diversity of 0.76. Unlike studies by other groups (Cowan et al 2002, Sola et al 2003, Sun et al 2004, MIRU locus 24 was polymorphic in the current study, with an allelic diversity of 0.6 and four alleles. Valcheva et al (2008) found MIRU locus 24 to have only a single allele, whereas Sun et al (2004) found a maximum of two alleles at this locus.…”

“…Unlike studies by other groups (Cowan et al 2002, Sola et al 2003, Sun et al 2004, MIRU locus 24 was polymorphic in the current study, with an allelic diversity of 0.6 and four alleles. Valcheva et al (2008) found MIRU locus 24 to have only a single allele, whereas Sun et al (2004) found a maximum of two alleles at this locus. This reinforces the observation that first-line typing schemes for different countries may require the use of different loci because the population structures of circulating M. tuberculosis strains vary across different parts of the world .…”

Evaluation of the Selected 12-locus MIRU for Genotyping Beijing Family Mycobacterium tuberculosis in Korea

“…The allelic diversity (ℎ) at each MIRU locus was calculated using the index ℎ=1−∑x i 2 , where x i is the frequency of the ith allele at the locus, as used in other studies 18,19 . The Hunter-Gaston discriminatory index (HGDI) was calculated in accordance with a method ex- …”

Mycobacterium tuberculosis Isolates from Single Outpatient Clinic in Panama City Exhibit Wide Genetic Diversity

“…Thus, our 12 MIRU-VNTR loci showed high discriminatory power, similar to previous reports using this marker set. 14,20 This allelic diversity allowed us to reach an HGDI of 0.988. Thus, we confirmed the discriminatory power of this set of 12 MIRU-VNTR loci for analyzing M. tuberculosis isolate samples in Panama City.…”