Study On Duplex PCR Diagnosis Method For Escherichia Coli Causing Edema Disease Of Swine And Trivalent Inactivated Bacterin

Edema disease of Swine (EDS) which causes an acute lethal infectious disease, is induced by some special serotypes Shiga-like toxin Escherichia coli(SLTEC). It commonly takes place in the large appetite, quick growth, physiclal status muscularity piglets. There are several symptoms with clinic test, such as ataxia, anaesthetization and eclampsia, which would be observed in organisms such as enterotoxemia, greater gastric curvature, mesentery scrofula and encephalic. Its morbility ranges from 10%-35%, while mortality reaches 90%. Even the survivor may grow slowly, hence this disease induces enormous lost in stockbreeding.In this experiment, we focused on establishing the PCR detecting method for SLTEC and inactivated vaccine.1. According to the sequences of B subunit of SLT-â…¡e and its cadherin F18ab pilus FedA from genbank, two primers were designed. We developed a method to detect the two kinds of nosogenesises by duplex PCR amplification. The two PCR products, one is about 230bp and the other is 510bp .Using the Duplex PCR detecting method we established before, combined with the results of microscope examination and biochemistry assay, 6 Shiga-like toxin Escherichia coli(SLTEC) isolates were identified from clinic samples. Six isolates were separated into 3 serotypes in the isolated E. coli. Three strains are O₁₃₉, and two strains are O₁₃₈, and another one is O₇₄, by stereotyping.2. SPF Kunrning mice was involved in estimating bacterial virulence and immunocompetence. Piglet assay was performed to identify the most-virulent and most-immunocompetence strain to develop porcine edema disease inactivated accine3. Protective efficacy test and antibody variety test were employed to monitor antibody titer of SPF Kunming mices inoculated porcine edema disease inactivated vaccine. The result indicated that the protective efficacy reached to 87.5%.4. Protective efficacy test and antibody variety test were used to monitor antibody titer of piglets inoculated porcine edema disease inactivated vaccine. The results showed that the protective efficacy reached to 80.0%. The piglets could obtain protective antibody in 14 days, the antibody reached highest titer in 21 days and persisted above protective titer in 180 days.In this reserch, a rapid method based on duplex PCR was developed to detect two pathogenicity factors of Escherichia Coli Causing Edema disease at the same time. 6 strains were identified and separated by this method. 3 strains which have different serotypes, were selected to develop tervalent inactivated vaccine. Every exprement was finished according to the New Veterinary Drugs Standard. These results will provide a base for the further clinical test.