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目的 研究结核分支杆菌耐利福平 (RFP)分离株rpoB基因突变情况并评价其应用价值。方法 采用聚合酶链反应一单链构象多态性 (PCR -SSCP)方法对 3 5 5株结核分支杆菌临床分离株 (其中敏感株 10 9株 ,耐RFP或含耐RFP株 2 46株 )rpoB基因进行检测 ,并对其中 3 1株耐药株用双脱氧末端终止法进行测序分析。结果 以结核分支杆菌标准株H3 7Rv为对照 ,所有 10 9株药物敏感株rpoB基因均无突变 ;2 46株耐药株中 ,2 2 5株rpoB基因SSCP图谱异常 ,其敏感性为 91 5 %。 3 1株耐药株 (其中SSCP异常 2 5株 ,正常 6株 )PCR扩增产物经测序证实 ,19株在 5 3 1位密码子TCG -TTG ,7株 5 2 6位密码子CAC -TAC ,3株在 5 16位密码子GAC -GTC ,2株未改变。结论 rpoB基因突变是耐RFP结核分支杆菌耐药性的主要分子机制 ;其最常见的突变位点是 5 3 1位色氨酸和 5 2 6位组氨酸 ,PCR -SSCP可快速检测结核分支杆菌RFP耐药性
Objective To study the mutation of rpoB gene of Mycobacterium tuberculosis (rpf) isolate and to evaluate its value. Methods PCR-SSCP was used to analyze the clinical isolates of 3 5 5 Mycobacterium tuberculosis isolates (10 9 sensitive strains, RFP-resistant or 466-resistant RFP-containing strains) rpoB Genes were detected, and 31 strains were resistant to sequencing by dideoxy terminator method. Results There was no mutation in the rpoB gene of all 10 9 drug-sensitive strains against the standard strain of Mycobacterium tuberculosis H3 7Rv. The SSCP patterns of rpoB gene were found to be abnormal in 225 strains of 2 46 strains, with a sensitivity of 91.5% . The PCR amplification products of 1 drug-resistant strain (including 25 SSCP abnormalities and 6 normal ones) were confirmed by sequencing. Among the 19 strains, TCG-TTG at 531 and CTC-TAC at nucleotide 526 , Three strains at codon 5-16 of GAC-GTC, two strains did not change. Conclusion The mutation of rpoB gene is the main molecular mechanism of resistance to M. tuberculosis. The most common mutation sites are tryptophan at 531 and histidine at 526, PCR-SSCP can detect tuberculosis branch rapidly Bacillus RFP resistance