[Direct, rapid detection for rifampin susceptibility to M. tuberculosis]

To evaluate the use of molecular biotechnology for direct, rapid detection of rifampicin-resistance mutations in M. tuberculosis.

45 M. tuberculosis clinical isolates and 70 sputum samples were tested by polymerase chain reaction-single stranded conformation polymorphism (PCR-SSCP) technique. M. tuberculosis strain H37Rv was used as control and compared with the result of susceptibility test. DNA sequencing was also performed in some of the strains.

All tested susceptible isolates displayed identical SSCP patterns. Of 29 RFP resistance strains, 26(90%) had distinct mobility shifts that can be discriminated from susceptible isolates. 9 sputum samples which were succeasfully evaluated by PCR-SSCP showed concordant result acquired from BACTEC 460 method. As the result of DNA sequencing, it was observed that seven RFP-resistance phenotype of M. tuberculosis strains had missense mutation, in which 5 isolates displayed TCG-->TTG or CAG mutations at codon 531.2 had CAC-->TAC mutation at codon 526. On the other hand, one strain which was susceptible to rifampin exhibited identical nucleotide alignment to the sequence of rpoB gene.

PCR-SSCP could be used as a method for simple, rapid, and reliable detection of rifampicin-resistance mutations in clinical samples of M. tuberculosis.