耐多药结核分枝杆菌pncA基因变异研究

发布时间：2019-04-27 08:54

【摘要】：目的:了解pnc A基因变异与结核分枝杆菌耐多药表型、菌株基因型及初/复治结核病的关系,为结核分枝杆菌耐吡嗪酰胺的分子检测及耐药机制提供实验室数据。方法:收集肺结核患者痰标本,行抗酸杆菌涂片、结核分枝杆菌培养、菌型鉴定、药物敏感试验及基因分型鉴定,提取结核分枝杆菌DNA,行pnc A基因扩增及测序,与NCBI数据库中标准菌株H37Rv序列比对,最后整理分析结果。结果:筛选出耐多药结核分枝杆菌103株(耐多药组)及对一线四种抗结核药物(HRSE)均敏感的临床分离株110例(敏感组)。耐多药组中,初治28.2%,复治71.8%;北京型菌株70.9%,非北京型菌株29.1%;48株发生pnc A基因变异,突变率为46.6%(18/103),其中初治突变占25%,复治突变占75%,北京型突变菌株占75%,非北京型突变菌株占25%。共检测到52种pnc A基因突变类型,其中18种与结核分枝杆菌PZA耐药高信度相关、18种已有文献报道、16种未见文献报道目前暂定为pnc A基因新突变,包括FSC46(Del G)、CCG54GCG、TTC58GTC、CCG62TCG、TCC65CCC、GGT97CGT、GAA107CAA、GAT126AAT、GAT126CAT、GTC131TTC、ACC135GCC、GTG155GAG、GGT162CGT、GCC170CCC、GTC180CTC及GAG181AAG,已报送美国基因库进一步核实。敏感组中,初治79.1%,复治20.9%;北京型菌株52.7%,非北京型菌株47.3%;仅3例出现pnc A基因变异,变异率为2.7%(3/110),其中2株为初治,1株为复治,2株为北京型,1株为非北京型,CAT42AAT、CGC29TGC及FSC167(Ins C)为新突变类型,已报送美国基因库进一步核实。结论:结核分枝杆菌耐多药表型与pnc A基因变异存在相关性,结合耐多药表型及pnc A突变可以一定程度预测和提示可能存在吡嗪酰胺耐药;北京家族菌株是耐多药结核病的主要流行菌株,北京基因型菌株可能更容易对吡嗪酰胺耐药;复治结核病结核分枝杆菌pnc A基因突变率明显增高。
[Abstract]:Aim: to investigate the relationship between pnc A gene mutation and multidrug resistance phenotype, genotype of Mycobacterium tuberculosis and primary / retreatment of tuberculosis, and to provide laboratory data for molecular detection and mechanism of resistance to pyrazinamide in Mycobacterium tuberculosis. Methods: sputum samples of patients with pulmonary tuberculosis were collected. The smear of acid-fast bacilli, culture of Mycobacterium tuberculosis, identification of bacteria type, drug sensitivity test and genotyping were collected. The DNA, of Mycobacterium tuberculosis was amplified and sequenced by pnc A gene. The results were compared with that of standard strain H37Rv in NCBI database, and the results were analyzed at last. Results: 103strains of multidrug-resistant Mycobacterium tuberculosis (MDR group) and 110 clinical isolates (sensitive group) sensitive to the first-line four anti-tuberculosis drugs (HRSE) were screened. In the multi-drug resistant group, the initial treatment rate was 28.2%, the retreatment rate was 71.8%, the Beijing type strain was 70.9%, the non-Beijing type strain was 29.1%. The mutation rate of pnc A gene was 46.6% (18 / 103) in 48 strains, of which 25% were newly diagnosed, 75% were re-treated, 75% were Beijing-type, and 25% were non-Beijing-type. A total of 52 pnc A gene mutations were detected, 18 of which were associated with the high reliability of PZA resistance in Mycobacterium tuberculosis, 18 have been reported, and 16 have not been tentatively reported as new mutations in the pnc A gene, including FSC46 (Del G), CCG54GCG,TTC58GTC,. CCG62TCG,TCC65CCC,GGT97CGT,GAA107CAA,GAT126AAT,GAT126CAT,GTC131TTC,ACC135GCC,GTG155GAG,GGT162CGT,GCC170CCC,GTC180CTC and GAG181AAG, have been reported to the US gene bank for further verification. In the sensitive group, the first treatment rate was 79.1%, the re-treatment rate was 20.9%, the Beijing type strain was 52.7%, the non Beijing type strain was 47.3%. The mutation rate of pnc A gene was 2.7% in only 3 cases. Among them, 2 strains were newly diagnosed, 1 was retreated, 2 were Beijing type, 1 was non-Beijing type, and CAT42AAT,CGC29TGC and FSC167 (Ins C) were new mutation types. Reported to the U. S. gene bank for further verification. Conclusion: there is a correlation between multidrug resistance phenotype of Mycobacterium tuberculosis and mutation of pnc A gene. The combination of multidrug resistance phenotype and pnc A mutation can predict and suggest the existence of pyrazinamide resistance to some extent. Beijing family strains are the main strains resistant to multi-drug tuberculosis, and Beijing genotype strains may be more susceptible to pyrazinamide resistance, and the mutation rate of pnc A gene in retreated Mycobacterium tuberculosis is significantly increased.
【学位授予单位】：遵义医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R52

【相似文献】