阴沟肠杆菌染色质ampD基因定点突变致AmpC β-内酰胺酶由非持续高产型转变为持续高产型  

Specific mutations of ampD gene leading to the emergence of derepressed high-production of AmpC β-lactamase

作　　者：涂婉[1] 赵虎[1] 

机构地区：[1]复旦大学附属华东医院检验科,上海200040

出　　处：《国际检验医学杂志》2010年第1期4-7,10,共5页International Journal of Laboratory Medicine

基　　金：上海市科委重点课题资助项目(No.054119509)

摘　　要：目的了解ampD基因特异突变对阴沟肠杆菌AmpCβ-内酰胺酶由非持续高产型转变为持续高产型的影响因素。方法筛选染色质介导的持续高产型AmpCβ内酰胺酶的阴沟肠杆菌,扩增其ampD基因并测序,确定其有义突变位点。采用定点突变的方法对野生型阴沟肠杆菌进行上述特异位点的突变,用头孢西丁三维试验测定突变前后菌株产酶类型的改变。结果 121株阴沟肠杆菌中15株为染色质介导的持续高产型,共筛选出8个有义突变位点,形成7株定点突变菌株。Ecl MA(274位插入A)、Ecl MC(327位缺失C)和Ecl MF(27位插入G)由非持续高产型转变为持续高产型,而Ecl MB(371位插入T)、Ecl MD(515位缺失C)、Ecl ME(324C→A)、Ecl MG(两点同时的突变,238C→A,302T→A)的产酶类型未发生变化。结论能够引起产酶类型发生改变的ampD基因特异突变均为可造成大片段氨基酸改变的框移突变,并且该改变至少从124位之前的氨基酸开始。但并不排除部分持续高产型AmpCβ内酰胺酶的产生存在其他调控机制。Objective To reveal the influence factor of ampD specific mutation on the transformation of AmpC β-1actamase from non-high-productive type to derepressed the high productive type in Enterobacter cloacae. Methods The selected chromatinmediated derepressed high-productive strains of Enterobacter cloacae and genornic DNA were extracted, and genomie ampD was amplified by using PCR, and then sequenced and confirmed by specific mutation. Use site-directed mutagenesis to mutate the wildtype Enterobacter cloacae for the above site. The changes of AmpCβ-1actamase types were detected by the improved Cefotaxime three-dimensional test. Results Of the 121 strains of Enterobacter cloacae, 15 strains were chromatin-mediated derepressed highproductive type,8 strains had significant mutations and 7 strains of site-directed mutagenesis were established. Ecl MA（274 inserting A）, Ecl MC（327 missing （7）and Ecl MF（27 inserting G）were changed from non-high-productive type to derepressed-high-produetive type,but Ecl MB（371 inserting T）,Ecl MD（515 missing C）,Eel ME（324 C→A）,Ecl MG（238C→A 302T→A）did not change in AmpC β-lactamase types. Conclusion The significant mutations that could lead to the transformation of AmpC β-1actamase were the Frame-shift mutations that could create the change of large region of amino acid. However, other mechanisms for the derepressed high-productive AmpC β-lactamase were not excluded in the study.

关 键 词：Β内酰胺酶类 肠杆菌 阴沟 基因 突变 

分 类 号：R378[医药卫生—病原生物学]