结核分枝杆菌耐异烟肼基因突变的快速检测

目的：探讨编码过氧化氢-过氧化物酶的katG基因突变与结核分枝杆菌异烟肼(INH)耐药性的相关关系。方法：根据结核分枝杆菌genebank中katG序列，自行设计特异性寡聚核苷酸引物，采用聚合酶链反应-单链构象多态性(polymerase chain reaction-single strand conformation polymorphism，PCR-SSCP)分析和直接测序法(direct sequencing，DS)分析结核分枝杆菌中katG基因突变情况．以H37，Rv标准株为对照。结果：所有23株敏感菌均未有SSCP结果异常；35株耐药菌中，有2株(5．7％)katG基因扩增阴性，且发生在高度耐药菌中．进一步分析发现，SSCP法突变检出23株(65．7％)，测序法突变检出24株(68．6％)，符合率为95．8％(23／24)．结论：参照测序法对耐药菌突变序列的分析结果，PCR-SSCP敏感、特异，可快速检测结核分枝杆菌katG耐药基因突变，有利于耐药结核分枝杆菌耐药性的快速检测。

Rapid detection of gene mutation in isoniazid-resistant M.tuberculosis

AIM: To investigate the relationship between the mutation of the katG gene and isoniazid resistance in M. Tuberculosis . METHODS: The specific oligonucleotide primers of the katG gene was designed according to the EMBL:X68081 in genebank. KatG gene mutations were detected by PCR SSCP and DS. H 37 Rv standard isolates were used as control. RESULTS: Compared with the control, no difference was found among the 23 sensitive isolates. Among the 35 resistant isolates, 2 (5.7%) lacked katG gene in those highly INH resistant isolates. The mutation of 23 (65.7%) isolates was detected by the PCR SSCP and that of 24 isolates was detected by DS. The coincidence rate for two the techniques was 95.8% (23/24). CONCLUSION: PCR SSCP is a sensitive and specific method for rapid detection of katG gene mutations in M. tuberculosis and its drug resistance.