Rapid Detection Of Gene Mutation In Multiple-drug Resistant Mycobacterium Tuberculosis By PCR And Single-strand Conformation Polymorphism Analysis

Objective: To establish PCR-SSCP for rapid analyzing resistant gene in multiple drug resistance of mycobacterium tuberculosis and to evaluate its use in clinics. Methods:30 mycobacterium tuberculosis strains in multiple drug resistance were analyzed by PCR-SSCP and classic drug susceptibility tests. A rpoB, rpsL and katG fragment were amplified from Mycobacterium tuberculosis trains and sputum specimens by PCR with a pair of primers designed according to the RFP, INH and SM resistant determinant of that three genes. The fragments of rpoB, rpsL and katG were analyzed by single-stranded conformation polymorphism (SSCP). Different PCR-SSCP patterns were obtained from RFP, INH, SM resistant stains and their susceptible control. Results: A 258bp and 282,267 bp fragments of rpoB and katG,rpsL were amplified from all mycobacterium tuberculosis strains. H₃₇R_V was used as a control,15 susceptible strains showed the same SSCP pattern. Respectively the gene mutation rate of rpoB, rpsL and katG resistance of isolated strains were 90% (27/30),53% (16/30),63%(19/30). There are total 8(26.7%) strains with three genes mutation and 18(60%) strains with two genes mutation. Among the MDR-TB strains,there are 2 strains with single gene mutation and 2 strains without gene mutation.