Russian "successful" clone B0/W148 of Mycobacterium tuberculosis Beijing genotype: a multiplex PCR assay for rapid detection and global screening

Abstract

We describe a multiplex PCR assay to detect the Mycobacterium tuberculosis Beijing genotype variant B0/W148, which is considered a "successful" clone of M. tuberculosis, widespread in Russia. Specificity and sensitivity of the assay were 100% based on the analysis of a collection of 516 M. tuberculosis isolates of different genotypes and origins. This assay may be used for accurate and simple detection and surveillance of this clinically and epidemiologically important variant of M. tuberculosis.

Fig 1

Schematic view of the genome region containing the IS6110 insertion specific of strain W148 (not to scale). Short arrows indicate the primers. PvuII site-flanked fragments are shown by dashed lines.

Fig 2

Southern hybridization of PvuII-digested DNA of M. tuberculosis isolates with probes. (a) Internal IS6110 fragment (primers INS1 and INS2); (b) Rv2664-Rv2665 fragment (primers W139F2 and W139R). Lanes: 1 to 5, Beijing B0/W148 isolates; 6, strain H37Rv; M, strain Mt14323 used as a molecular mass marker. The double-headed arrow indicates fragments corresponding to IS6110 RFLP profiles and the PvuII digest spanning the Rv2664-Rv2665 region in the Beijing B0/W148 isolates.

Fig 3

Agarose gel electrophoresis of the multiplex PCR products of M. tuberculosis isolates. Lanes: 3 to 5, 7, and 9, Beijing B0/W148 cluster isolates; 1 and 2, isolates of other Beijing variants; 6, H37Rv; 7, BCG; M, 100-bp DNA ladder (GE Healthcare). Arrows show specific bands for Beijing B0/W148 (1,018 bp) and other genotypes (410 bp).