Drug-resistant gene detection in clinically isolated Mycobacterium tuberculosis isolates

Objective:To research the molecular mechanisms of drug resistance in M. tuberculosis, and to develop a new method for detecting the drug resistance. Methods:The rpoB, rpsL, katG genes and inhA regulatory sequence in 62 M. tuberculosis clinical isolates were analysed with PCR, PCR-SSCP and PCR-RFLP. Results:The sensitivity of amplifying the drug-resistant genes with PCR was 1￣10 pg DNA. In comparison with conventional susceptibility testing methods, the sensitivity and specificity of detecting the drug-resistant genes in M. tuberculosis isolates with PCR and PCR-SSCP were 93.8% and 96.2% for rpoB, 81.6% and 91.7% for rpsL, 76.0% and 75.7% for katG, and 16% and 100% for inhA, respectively. 90% of the multidrug-resistant strains had mutations in more than two drug-resistant genes. We had analyzed the rpsL gene codon 43 in streptomycin-resistant isolates. It was found that the rpsL gene mutations all situated at codon 43.Conclusion:Alterations in rpoB, rpsL and katG gene may be the important mechanisms of M. tuberculosis resistance to rifampin, streptomycin and isoniazid. PCR, PCR-SSCP and PCR-RFLP are going to become the simple, rapid and reliable diagnostic tests for drug resistance.