Usefulness of ligation-mediated PCR genotyping in tracking outbreak-associated Mycobacterium tuberculosis strains.

Abstract

With the emergence of a multidrug resistant tuberculosis (MDR-TB) outbreak, the availability of a rapid typing method to carry out a nationwide prospective survey for the tracking of newly emerging MDR-TB foci became a priority. For this purpose, we have applied the IS6110 PCR-based genotyping assay, namely, LM-PCR (ligation-mediated PCR). The latter relies on ligation of a synthetic oligonucleotide priming site to a restriction site flanking IS6110. Sequences between the IS element and the restriction site are then amplified using an IS6110 specific outward primer and an oligonucleotide specific to the ligated priming site. Although it was found slightly less discriminative than the standard IS6110 restriction fragment length polymorphism analysis (IS6110 RFLP), LM-PCR allowed for the rapid and prospective identification of new outbreak-related cases within a large pool of circulating M. tuberculosis isolates. In comparison to IS6110 RFLP LM-PCR was found simple enough to justify its implementation in laboratories involved in MDR-TB surveillance at a nationwide scale.