Automated high-throughput genotyping for study of global epidemiology of Mycobacterium tuberculosis based on mycobacterial interspersed repetitive units

Large-scale genotyping of Mycobacterium tuberculosis is especially challeng ing, as the current typing methods are labor-intensive and the results are difficult to compare among laboratories. Here, automated typing based on va riable-number tandem repeats (VNTRs) of genetic elements named mycobacteria l interspersed repetitive units (MIRUs) in 12 mammalian minisatellite-like loci of M. tuberculosis is presented. This system combines analysis of mult iplex PCRs on a fluorescence-based DNA analyzer with computerized automatio n of the genotyping. Analysis of a blinded reference set of 90 strains from 38 countries (K. Kremer et al., J. Clin. Microbiol. 37:2607-2618, 1999) de monstrated that it is 100% reproducible, sensitive, and specific for M. tub erculosis complex isolates, a performance that has not been achieved by any other typing method tested in the same conditions. MIRU-VNTRs can be used for analysis of the global genetic diversity of M. tuberculosis complex str ains at different levels of evolutionary divergence. To fully exploit the p ortability of this typing system, a website was set up for the analysis of M. tuberculosis MIRU-VNTR genotypes via the Internet. This opens the way fo r global epidemiological surveillance of tuberculosis and should lead to no vel insights into the evolutionary and population genetics of this major pa thogen.