广西地区仔猪致病性大肠杆菌菌毛基因多重PCR检测及质粒DNA指纹图谱分析.docVIP

广西地区仔猪致病性大肠杆菌菌毛基因多重PCR检测及质粒DNA指纹图谱分析 　　摘要对广西地区56株仔猪致病性大肠杆菌进行菌毛基因的多重PCR检测及质粒DNA指纹图谱分析。多重PCR检测结果证实该方法可用于各大肠杆菌野生菌株的检测,且适用于仔猪大肠杆菌性腹泻病的快速诊断和流行病学调查。质粒DNA指纹图谱分析发现这56个菌株质粒的获得率为100%,来源相同的菌株具有相同或相似的质粒图谱,来源不同的菌株的质粒图谱一般不同。所有菌株都含有1 条大小相同的质粒带,表明广西流行的猪大肠杆菌性腹泻病主要是由携带相同质粒的大肠杆菌引起。试验结果表明,质粒DNA指纹图谱法可作为大肠埃希氏菌分型的分子生物学方法。 　　关键词仔猪大肠杆菌;菌毛基因;多重PCR;质粒指纹图谱;广西壮族自治区 　　中图分类号S858.285.1+2;Q78文献标识码A文章编号 1007-5739(2010)07-0349-03 　　 　　DetectionofFimbrialGene ofPathogenicEscherichia colifromPigletsinGuangxiRegionbyMultiplexPCRAssayandItsPlasmidProfileAnalysis 　　XIE Yong-pingPENG HaoCHEN Ze-xiangYANG Wei 　　(Guangxi Veterinary Research Institute,Nanning Guangxi 530001) 　　AbstractThe fimbrial gene of fifty-six strains of pathogenic Escherichia coli from piglets in Guangxi were detected by multiplex PCR and analysised by plasmid profile.The results showed that the multiplex PCR assay could be used to detect the wild type E.coli strains and also had been proved to be the useful method for rapid diagnosis and epidemiological investigation of piglets diarrhoea caused by E.coli.Plasmid were discovered in all E.coli strains and the isolates from different sources had distinct plasmid patterns,while those from same sources held same plasmids.All strains contained a plasmid with the same size,which indicated that the pathogenic E.coli caused piglets diarrhoea in Guangxi had one common plasmid. The results of analysis suggested that Plasmid DNA Finger Printing method is useful to subtype the Escherichia coli. 　　Key wordspiglets;Escherichia coli;fimbrial gene;multiplex PCR;plasmid profile analysis;Guangxi zhuang Aotonomous Region 　　 　　致病性大肠杆菌所致的腹泻是仔猪腹泻最主要的原因,各地普遍存在[1]。随着养猪向规模化发展,仔猪黄、白痢在我国及世界各地均有发生,目前仍是我国养猪业健康发展的严重制约因素[2]。黄、白痢的大肠杆菌的致病性与菌株的粘附性菌毛、肠毒素、R 质粒等有关[3]。致病性大肠杆菌常具有K88、K99、987P等粘附性菌毛中的一种或几种[3]。粘附性菌毛的检测对致病性大肠杆菌的诊断具有重要意义。 　　而肠毒素及R质粒的基因主要由大肠杆菌的质粒所编码[4],笔者在对广西地区规模化猪场56株大肠杆菌分离株进行培养及生化鉴定的基础上,并对所有分离株的菌毛基因进行多重PCR检测及质粒指纹图谱分析,为研究仔猪致病性大肠杆菌腹泻病的快速诊断和分子流行病学调查奠定了基础。 　　1材料与方法 　　1.1供试菌株 　　56株大肠杆菌来源于广西壮族自治区不同规模化猪