【摘要 】 目的 了解我国结核分支杆菌耐利福平(RFP)分离株rpoB基因突变情况，建立快速检测结核分支杆菌耐药基因型的分子药敏试验方法。方法 通过聚合酶链反应-单链构象多态性(PCR-SSCP)和PCR-直接测序法(PCR-DS)分析50株结核分支杆菌临床分离株的rpoB基因。结果 3株结核分支杆菌药物敏感株和2株非RFP耐药株中，仅1株rpoB SSCP图谱异常的药物敏感株出现531位密码子TCG→TTG突变。45株RFP耐药株中，10株SSCP和DS分析均未见rpoB突变，35株SSCP和DS分析异常，其中14株为531位密码子TCG→TTG或TGG或TAC突变，14株为526位CAC→TAC或GAC或CCC或CTC或GTC突变，2株为516位GAC→GTC或TAC突变，2株为516位和526位及515位和516位密码子双点突变，3株rpoB序列与结核分支杆菌不同。结论 大多数结核分支杆菌耐RFP是由于其rpoB基因突变所致，采用PCR-SSCP和PCR-DS方法可快速测定结核分支杆菌RFP耐药基因型。

Wu Xueqiong, Zhang Junxian,Zhuang Yuhui, et al. Tuberculosis Research Laboratory, The 309th Hospital, PLA, Beijing100091

【Abstract 】 Objective To understand mutation of rifampicin-resistant genes in M.tuberculosis clinical isolates, and to develop a new method for detecting drug resistance.Method Analyzing the rpoBgenes in 50 M. tuberculosis clinical isolates with polymerase chainreaction-single-stranded conformation polymorphism (PCR-SSCP) and PCR-direct sequencing(PCR-DS) techniques. M. tuberculosis strain H₃₇ Rv was used ascontrol. Result Of 3drug-sensitive isolates and 2 non-rifampicin-resistant isolates selected, only 1drug-sensitive isolate with abnormal rpoB SSCP profile had TCG→TTG mutation at codon 531. Of 45 rifampicin-resistant isolates, 35isolates with abnormal SSCP profiles had mutations in rpoB sequences, in which 14 isolatesdisplayed TCG→TTG or TGG or TAC mutationsat codon 531, 14 had CAC→TAC or GAC or CCCor CTC or GTC mutations at codon 526, 2 had GAC→GTC or TAC mutations at codon 516, 2 had two point mutations, and 3had different rpoB sequences from that of M. tuberculosis. Conclusion Resistance to rifampicin in most M. tuberculosis isolatesis due to the mutations on the genes encoding the RNA polymerase subunit (rpoB).PCR-SSCP and PCR-DS techniques might become simple, rapid and reliable diagnostic testsfor rifampicin resistance in clinical isolates.

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