临床常见产ampC β-内酰胺酶菌株中染色质ampD基因突变位点的研究

Exploration of the mutations of genomic ampD from the clinical isolates producing ampC β-lactamases

目的了解临床常见产ampCβ-内酰胺酶菌株中染色体ampD的分布及其突变位点。方法41株临床常见产ampCβ-内酰胺酶菌株(阴沟肠杆菌,肺炎克雷伯菌,鲍曼不动杆菌,铜绿假单胞菌)分离自医院感染样本,抽提其染色质DNA,PCR法扩增ampD基因并连接入pMD19-T载体后,再行PCR确定其存在。双链测序后比对染色体ampD基因的突变位点。结果41株细菌的基因组中,使用PCR法扩增出染色体ampD基因有36株,并成功测定了其ampD基因的序列;比对后发现阴沟肠杆菌、肺炎克雷伯菌、鲍曼不动杆菌和铜绿假单胞菌的染色体ampD基因的核苷酸序列差异较大。染色质ampD易突变位点分析统计表明,这些细菌的ampD基因的平均突变率分别为:64.5%、100.0%、97.0%和95.0%。结论大部分临床常见产ampCβ-内酰胺酶菌株含有ampD基因,并且有较高的突变率。

Objective To explore the mutations of the genomic ampD from the clinical isolates producing ampC β-lactamases. Methods Forty-one bacilli produced ampC β-lactamases were isolated from the nosocomial infection. The genomic DNA was extracted, genomic ampD was amplified using PCR, ligated into pMD19-T vector, and sequenced. Mutation sites were analyzed. Results Of the 41 clinical isolates, 36 were identified to contain the genomic ampD sequences, and their sequences were successfully measured. The results of the mutation analysis showed that the average mutation rates of Enterobacter cloacae, Klebsiella pneumonia, Acinetobacter baumannii and Pseudomonsa aeruginosa were 64. 45 %, 100.00 %, 97.00%, and 95.00%, respectively. Conclusions A large proportion of the clinical isolates producing ampC lactamases contained genomic ampD which is highly mutated.