Research Of Actinobacillus Pleuropneumoniae Bacterial Ghost Loading Of Pasteurella Multocida Outer Membrane Protein (Omp) H DNA Vaccine

Porcine Contagious Pleuropneumonia and Swine Lung Plague as common bacterial infections of pigs, often have mixed or secondary infection at respiratory tract, making great damage to aquaculture. Vaccination is the best way to combat the disease in production practice. Commonly used inactivated vaccine has no protection for heterologous infection and can not prevent bacterial colonization.These bring some difficulties for production practice. Bacterial ghost as a new type of vaccine retaining intact membrane antigen in the gentle preparation process, can effectively induce humoral and cellular immunity. Therefore, whether as a vaccine or carrier, bacterial ghost has extensive value of use.In the experiment, lysis protein E gene and nuclease A gene (SN) were amplified by PCR, using phage PhiX174 and S. aureus genome as templates. The fusion gene E-15aalinker-SN was achieved by touchdown PCR through linker primers. Insert E gene and E-15aalinker-SN gene into pBV220 plasmid to construct pBV-E and pBV-E-15aalinker-SN. Amplify dual-gene lysis system (DLS) using pBV-E-15aalinker-SN plasmid as template. On the basis of pMD18-T and pMC-Express, pMD-WK and pMC-WK were completed. Cracking tests showed E-15aalinker-SN gene was better than E gene in the lysis activity, having a lysis rate at 99.999%. In the control of sodC promoter, DLS expressed lysis activity in App and finally got App bacterial ghosts.Using Pasteurella multocida genome as template, ompH gene was amplified by PCR. Construct pCDNA-ompH eukaryotic expression system as DNA vaccine. By soaking, the plasmid pCDNA-ompH was loaded into App bacterial ghosts. Animal tests showed that the App bacterial ghosts loading pCDNA-ompH plasmid vaccine had protection rate up to 100%.