Separation, Identification And Pathogenicity In Mice Of EHEC O157:H7

Enterohemorrhagic Escherichia coli (EHEC) O157:H7is a kind of important animal pathogens, it cause diarrhea, hemorrhagic colitis (HC),thrombotic thrombocytopenic purpura (TTP)and hemolytic uremic syndrome (HUS) and other serious complications, some HUS cases cause fatalities. The bacteria long dwell in cattle, sheep, pigs, chickens and other livestock and poultry gut, cattle are a principal reservoir, causing the animal diarrhea, and the pollution of livestock and poultry products and water and crop farming, to bring enormous loss and pose a huge threat to human health.1Detection and separation of Escherichia coli O157:H7420Fresh samples were collected from all regions of Nanjing:the faecal samples350,vegetables50,meat10, milk10, After cultured at37℃, enriched with immunomagnetic beads and spread onto selective agar. sorbitol-MacConkey agar culture medium (CT-SMAC) after cultured colony is circular, moist surface like, translucent, were suspicious colonies were rfbE/fliC PCR, while O157and H7diagnostic serum agglutination test for identification. Results:successful isolation of5strains of enterohemorrhagic Escherichia coli O157:H7. the positive rates of Samples of enterohemorrhagic Escherichia coli0157:H7is8%,2%,0%,2%,2%. The gram stain is negative bacilli,and lstrain delayed fermentation of sorbitol MacConkey medium.2Identification of isolated strains of Escherichia coli O157:H7According to isolated O157:H7strains design the stxl, stx2, eae, hlyA, and tccp primers to amplify the fragments, PCR identification,and biochemical test and drug sensitivity test.Virulence genes show:one of the4strains of phenotypic virulence factors were stxl-stx2+eae+hlyA+tccp+, otherwise a phenotype of stxl+stx2+eae+hlyA+tccp+, the bacterial tccp gene were positive, but contains different numbers of repeat fragments. Different isolates resistance difference.6#has strong resistance to ampicillin,kanamycin, pioneer V, streptomycin.3. Pathogenicity of Mouse of Escherichia coli O157:H7strain BALB/C mices were inoculated intragastrically and we studies the pathogenicity of Escherichia coli O157:H7, selection of eight week old mice, use streptomycin-treated, with1×1010CFU with oral dose of inoculated with PBS of five0157:H7strains isolated respectively, the clinical symptoms after infection of mice and survival were observed, the results showed:in mice infected with the bacteria in mice bacteria discharge time is7,9,13,13,15days respectively, the survival rate is40%,50%,60%,20%,50%, It’s established the foundation for animals infection model.