浙江省O157大肠杆菌监测及其分子生物学特性

目的了解肠出血性大肠杆菌O157∶H7和其他O157大肠杆菌在浙江省动物、人群中的分布、流行以及PFGE分型、毒力基因携带状况。方法按全国O157∶H7监测方案在5-10月份肠道传染病高发季节,采集全省各地(市)肠道门诊腹泻病人粪便,进行O157大肠杆菌分离培养,并用免疫磁珠分离法对浙江省5个监测点的宿主动物进行O157∶H7分离培养、鉴定,可疑菌株以PCR法检测O157∶H7抗原、志贺样毒素(stx1和stx2)、粘附抹平因子(eaeA)及溶血素(hly)4种毒力基因。用脉冲场凝胶电泳(pulse field gel electrophoresis,PFGE)方法进行同源性分析,同时选择14种抗生素进行药敏试验,并将分析结果与本省首株患者粪便中分离的产志贺样毒素的O157∶H7菌株进行比较。结果全省5个监测点2006年共监测动物粪便标本2377份,分离到4株O157∶H7菌株,阳性率为0.17%;同时在绍兴、舟山肠道门诊腹泻病人粪便中分离到2株O157:H?菌株。4株O157∶H7菌株,stx2、Hly、eaeA均阳性,stx1均阴性;2株O157∶H?菌株仅1株携带eaeA毒力基因。脉冲场凝胶电泳分型显示,4株O157∶H7菌株分3个型,除金华地区2006年分离所得2株完全相似外,其它相同地区不同年代分离的O157∶H7菌株及相同年代不同地区分离的O157∶H7菌株则完全不相似。2株O157∶H?菌株1株PF-GE电泳条带降解,另1株与其它O157∶H7菌株电泳条带差异明显。结论浙江省大肠杆菌0157菌株在动物中以携带stx2毒力基因的O157∶H7菌株为主,但在腹泻患者中则以不带志贺样毒素的O157∶H?菌株为主。不同地区分离的O157∶H7菌株PFGE分型差异明显。羊、奶牛是携带stx2毒力基因的O157∶H7大肠杆菌的主要宿主。各级疾控应加强对宿主动物和腹泻病人大肠杆菌O157的分离监测和流行病学调查。

Surveillance and molecular characterization of Escherchia coli O157 in Zhejiang province

To understand the distribution of enterohemorrhagic Escherichia coli(EHEC)O157∶H7 and other E.coli O157 in animals and human in Zhejiang province,their prevalence of infections,the pulse field-gel electrophoresis(PFGE)patterns,and the condition of individuals carrying the virulence genes,according to the strategies of National Surveillance on E coli O157∶H7,the stool samples from patients at diarrhea clinics of different prefectures and cities in Zhejiang province were collected from May to October 2006 when the enteric infections were highly prevalent. From these samples,E coli O157 strains were isolated and the virulence genes including stx1,stx2,hly and eaeA were detected from these isolates by routine PCR assay. PFGE was used for the homology analysis,and the anti-microbial susceptibility tests were performed for 14 antibiotics. The results obtained from these analyses were compared with the O157∶H7 strain producing Shiga-like toxin firstly isolated from stool of patient in Zhejiang province. The results showed that 4 strains of E coli O157∶H7 were isolated from 2377 stool samples of animals in 5 surveillance area in Zhejiang province,with a positive detection rate of 0.17% ,meanwhile, 2 strains of E coli O157∶H? were isolated from stool samples of patients with diarrhea in Shaoxing and Zhoushan districts. As demonstrated by PCR assay,in these 4 strains of animal origin,the stx2,hly and eaeA genes were detected,but the stx1 gene was not found,while only one strain carried the eaeA gene in 2 strains of O157∶H7. Results of PFGE typing revealed that 3 types could be differentiated in 4 strains of animal origin,and except 2 strains isolated from Jinhua district in 2006 with the same types,other strains of O157∶H7 isolated in the same regions during different years or strains isolated at different regions at the same years were entirely different.Electrophoretic band of one O157∶H?isolate was showed to be degraded and the PFGE pattern of the other one showed apparent difference with others. From the above observations,it is apparent that the predominant strains of E coli O157 of animal origin isolated from Zhejiang province carries the stx2 virulence gene,but strains isolated from patients with diarrhea are predominantly the O157∶H?without Shiga-like toxin. It was also proved that sheep and milk cows were the main hosts of E. coli O157∶H7 carrying the stx2 gene. Disease controls at different levels should be strengthen for the surveillance and epidemiologic investigation on E.coli O157 strains in zooid hosts and patients with diarrhea.