Immunogenicity Studies On Outer Membrane Proteins In Strains Of Haemophilus Parasuis With Different Serotype

Haemophilus parasuis(HPs) can often invade and cause Glasser's disease which were pathogen. Outer-membrane proteins(OMP) is the virulence factor of Haemophilus parasuis, which have good immunogenicity. The experiment had extracted Haemophilus parasuis OMP and studied immunogenicity. It establish the foundation with next study of Haemophilus parasuis OMP subunit vaccine.1. Haemophilus parasuis OMP extraction and concentration determinationTo study Haemophilus parasuis OMP immunogenicity, this experiment selected Haemophilus parasuis strains serotype 1,4,5,12,13. To explore extraction and identification Haemophilus parasuis OMP, We had used ultrasound wave and Sodium Lauroyl sarcosine (SLS).We could determine contents of Haemophilus parasuis OMP with nucleic acid protein equipment, and also do SDS-PAGE electrophoresis and Western-blot test to analysis the immunoreactivity of Haemophilus parasuis OMP. the concentration of OMP of Haemophilus parasuis strains serotype 1,4,5,12,13, were respectively 58.3mg/ml, 17.1mg/ml, 34.7mg/ml, 63mg/ml, 28.7mg/ml by nucleic acid protein equipment. Western-blot of OMP found that Haemophilus parasuis have a good immunoreactivity.2. Study on antigenicity of the outer-membrane protein of Haemophilus parasuisPrepare the oil-emulsion OMP antigen and the whole-cell inactication vaccine to immunity subcutaneous inoculation with the 6-week-old Kunming mice. The concentration of OMP were 10ug/each, 20ug/each, 40ug/each, 80ug/each, and the whole-cell vaccine is 0.2ml/each. Detect the change of the level of antibody by ELISA. The results showed that: The whole-cell of Haemophilus parasuis vaccine and OMP vaccine have a strong immunogenicity in Kunming mice. After 7 days, the positive antibody of OMP can be detected firstly. Haemophilus parasuis OMP serotype 1 best immune doses was 80ug/each, while the rates of protection conteracting toxic substances reached 100%; the rates of protection conteracting toxic substances reached 70% with the whole-cell inactication vaccine. Haemophilus parasuis OMP serotype 4 best immune doses was 80ug/each, while the rates of protection conteracting toxic substances reached 80%; the rates of protection conteracting toxic substances reached 60% with the whole-cell inactication vaccine. Haemophilus parasuis OMP serotype 5 best immune doses was 40ug/each and 80ug/each, at this time the rates of protection conteracting toxic substances all reached 80%, indicating that serotype 5 OMP can provide better protection in mice when 40ug; the rates of protection conteracting toxic substances reached 80% with the whole-cell inactication vaccine. Haemophilus parasuis OMP serotype 12 best immune doses was 40ug, while the rates of protection conteracting toxic substances reached 90%; the rates of protection conteracting toxic substances reached 80% with the whole-cell inactication vaccine. Haemophilus parasuis OMP serotype 13 best immune doses was 80ug, while the rates of protection conteracting toxic substances reached 80%; the rates of protection conteracting toxic substances reached 80% with the whole-cell inactication vaccine. By testing the level of antibodies and the rates of protection conteracting toxic substances, we had found that OMP had good immunogenicity. The testing results will establish a good foundation for Haemophilus parasuis OMP's rapid diagnostic kit, the development of subunit vaccine and the effective control of the disease.