摘要
目的快速检测痰中耐药结核分枝杆菌katG基因及了解耐异烟肼的分子机制。方法用PCR方法直接从耐药肺结核患者痰及临床分离菌株扩增katG基因片段,对扩增获得的katG基因片段进行序列测定,比较分析耐多药、全耐、多耐药但对异烟肼敏感、全敏感或标准结核分枝杆菌株之间的基因序列差异。结果于6 h内从耐药肺结核痰中快速检测到katG基因。从临床分离菌株中随机扩增药敏试验证实的耐多药3株,全耐、包括对异烟肼耐药的多耐、对异烟肼敏感的多耐药、全敏感及H37Rv标准结核分枝杆菌株各1株,均扩增获得273bp片段,序列测定比较发现,2株耐多药菌、1株全耐菌及包括异烟肼耐药的多耐药菌株均检测有katG基因点突变,突变位点均为第2066位碱基C突变为G,1例MDR-TB、全敏感株、对异烟肼敏感的多耐药菌株和标准株均无基因突变。结论PCR及序列分析方法可快速、准确检测结核分枝杆菌katG基因及其突变,结核分枝杆菌对异烟肼的耐药性与katG基因点突变有关,但可能还存在其它值得探讨的机制。
Objective To rapidly detect katG gene of Mycobacterium tuberculosis in sputum and to explore isoniazid（INH）-resistant mechanism. Methods katG fragment was amplified from M. tuberculosisstrains from sputum specimens of drug-resistant inpatients or clinic isolates by PCR. The fragments of katG were sequenced. The difference of katG gene fragments sequences of clinical isolates among multi drug-resistant, all drugs-resistant, multiple drug-resistant but INH-susceptible,all drugs-susceptible isolates of M. tuberculosis, or standard strain were compared. Results Within six hours, katG gene of M. tuberculosisstrains from sputum specimen of drug-resistant pulmonary tuberculosis was detected. A 273bp fragment of katG was amplified from three multidrug-resistant strains, two all drugs-resistant strains, one multiple drug-resistant but INH-susceptible isolate, or all drugs-susceptible clinical isolates and H37Rv standard strain of M. tuberculosisstrains. By comparative analysis of gene sequence, spot mutation was found from two multidrug-resistant, two all drugs-resistant strains in M. tuberculosisstrains. Nucleotide C at 2066 was mutated to G. No mutation in H37Rv standard strain, one multidrug-resistant strain,one multiple drug-resistant but INH-susceptible isola or all drugs-susceptible isolate was detected. Conclusion katG gene and its mutation in M. tuberculosis strains could be rapidly and accurately identified by polymerase chain reaction and sequence analysis. INH-resistance was related with spot mutation of katG gene in M. tuberculosisstrains, but other mechanisms may need explored.
出处
《江西医学院学报》
2008年第4期19-21,共3页
Acta Academiae Medicinae Jiangxi
基金
福建省青年科技人才创新基金项目(No.2005J081)
关键词
耐药结核分枝杆菌
异烟肼
KATG基因
聚合酶链式反应
drug-resistant Mycobacterium tuberculosis
isoniazid
catalase-peroxidase enzyme（katG） gene
polymerase chain reaction
