肠出血性大肠杆菌O157:H7外膜蛋白A敲除菌株的构建及其黏附作用研究  

Knockout of gene encoding outer membrane protein A of enterohaemorrhagic Escherichia coli O157:H7 and study on its function

作　　者：王海光[1] 顾江[1] 方瑶[1] 于波[1] 李倩[1] 张卫军[1] 毛旭虎[1] 

机构地区：[1]第三军医大学检验系临床微生物及免疫学教研室,重庆400038

出　　处：《免疫学杂志》2012年第5期420-423,共4页Immunological Journal

基　　金：国家重大新药创制专项课题(2008ZXJ09014-007)

摘　　要：目的利用Red同源重组技术构建肠出血性大肠杆菌(EHEC)O157∶H7外膜蛋白A(ompA)基因敲除菌株,研究敲除菌株的黏附能力变化,阐明OmpA在细菌黏附以及致病中的作用。方法利用Red同源重组技术对EHEC O157∶H7 ompA基因进行敲除,同时构建EHEC O157:H7 ompA回复突变菌株,最后通过细胞试验对EHEC O157∶H7野生型菌株、敲除菌株及回复突变菌株的黏附能力进行比较。结果成功构建了EHEC O157:H7 ompA基因敲除菌株及其回复突变菌株。细胞试验结果表明,与野生型菌株相比,敲除菌株的黏附能力明显下降,而将ompA基因进行回复突变后其黏附能力又得到回复。结论 OmpA在EHEC O157∶H7黏附HeLa细胞过程中发挥重要作用,ompA基因敲除菌株的获得为进一步研究其功能提供了帮助。In this study,gene encoding outer membrane protein A（ompA） of enterohaemorrhagic Escherichia coli（EHEC） O157：H7 was knocked out（ompA-） using λ red homologous recombination system.To clarify the roles of OmpA in cell adhesion and pathogenesis,ompA-EHEC O157：H7 was then complemented with a plasmid containing the ompA gene and its promoter（ompA＋）.Meanwhile,the ability of ompA＋ EHEC O157∶H7 and ompA-EHEC O157∶H7 strains to attach to HeLa cells were compared with that of wild type EHEC O157∶H7 through cell adhesion assay.Results showed that the cell adhesion ability of ompA-EHEC O157：H7 strain decreased significantly,when compared with that of wild type EHEC O157：H7 and ompA＋ EHEC O157：H7.These data demonstrated that the OmpA of EHEC O157：H7 may play an important role in HeLa cell adhesion.In addition,ompA-EHEC O157∶H7 strain obtained in this study would provide a useful tool for the further study of EHEC O157∶H7 pathogenesis.

关 键 词：肠出血性大肠杆菌O157∶H7 外膜蛋白A 基因敲除 细胞黏附 

分 类 号：R378.21[医药卫生—病原生物学]