The Preparation And Analysis Of Immune Efficacy Of Bovine Pasteurella Multocida Ghosts

Bovine Pasteurella multocida, also known as bovine hemorrhagic septicemia, is an acute infection caused by P.multocida infection, acute hemorrhagic septicemia yielded rapid death; Multiple lesions of chronic cases confined to the local organ, usually through subcutaneous connective tissue, joints, organs purulent lesions. Currently primarily used to prevent bovine P.multocida disease vaccines for virulent strains of inactivated vaccine, attenuated live bacteria vaccines and subunit vaccines, because immune effect of traditional whole-cell bacterial inactivated vaccine is not ideal, It does not provide a strong protection force, attenuated live bacteria vaccine its inherent live P.multocida, in the host animal organism exist risk reversion of virulence of the organism. Therefore, the development of a safe and effective vaccine is important for the control of bovine pasteurellosis.In this study, the temperature-sensitive lysis cassettes containing phage Phi X174 lytic g ene E and start repressor system CI857-PL-PR was amplified from plasmid p HH43 by P CR, the lysis cassettes CI857-PL-PR-E was inserted into Shuttle vector p PBA1100. Cons truction of bacteriolysis plasmid p PBA1100-E, p PBA1100-E was transfered into bovine P.multocida through electric conversion method, bovine P.multocida harboring the bacterioly sis plasmidwas grown in a shaker at 37℃until OD600 nm value is 0.4, the culture tem perature is raised to 42 ℃induced prepare bovine P.multocida bacterial ghosts, Calculate d separately the number of bacteria before induction and post-induction,Its lysis efficienc y up to 99.997%. Ghost retained the basic structure of bacteria By electron microscopy a nd their contents were released to extracellular region. Leaving onlythe shell of bacteria, in the middle of the cells showed obvious holes.Using bacterial ghost vaccine immunization of mice, analysis of ghost vaccineefficacy. T he 5-week-old female BALB / c mice were randomly divided into four groups:ghost、g host+adjuvant、nactivated vaccine and phosphate buffer solution group. After laboratory a daptation one week, two weeks apart vaccinated twice in a row. The serum antibody tite rs were determined using indirect enzyme-linked immunosorbent assay(ELISA); Using cy tokine detection kit to detectserum levels of IL-4 and IFN-γ of content; CD4+and CD8+ weredetected by flow cytometry. And survival rates, the immune protection efficacy of va ccine were assessed. Mice was challenged by bovine P.multocida After second immunity two weeks. Record survival after challenge, humoral and cellular immunity analyzed the effectiveness of the immune bovine P.multocida ghostvaccine.The test results show that this test prepared bovine P.multocida bacteria ghosts vaccine can induce mice higher levels of humoral and cellular immunity, For mice against bovine P.multocida challenge provide higher protection force. The test laid a solid foundation. For the further study of bovine P.multocida bacterial ghost vaccine. This laid the foundation for the further development of the bovine P.multocida ghost vaccine.