Isolation And Identification Of Infectious Bursal Disease Virus (IBDV) And Establishment Of The Method Of Immunohistochemistry Assay For The Detection Of IBDV

Infectious bursal disease (IBD) is a highly contagious disease of young chickens caused by infectious bursal disease virus (IBDV), characterized by immunosuppression and high mortality generally at 3 to 6 weeks of age. Chickens once infected with IBDV are susceptible to other diseases such as New Castle disease, Marek disease and so on due to vaccination failure.In recent years, there was an increasing incidence of IBD in Anhui province, which brought economic devastation to the poultry industry due to interference with effective vaccination. A survey showed that some vaccinated chickens could also be infected with IBDV. In order to make a more effective vaccination and minimize the losing caused by IBD, it is important to investigate the occurrence of IBD in Anhui province.In this dissertation, IBD occurred in Anhui province was studied systematically.First, bursals and spleens collected from chickens that suspected suffered IBD in Hefei and peripheral areas where IBD broke out were grinded thoroughly, frozen and melted for several times, then centrifuged and the supernatant was collected. Virus was isolated after passeged in chicken embryos then was subjected to agar diffusion test and colloidal gold stripes test, and four virus strains were identified, one of them was named IBD -1 for further study.Second, IBD -1 was used as antigen for the preparation of rabbit anti-chicken antibody. The titer of the antibody was at a high level of 1: 128. The serum was purified with the method of salting out and dialysis, and the protein concentrations was detected by the Folin-phenol method and ultraviolet spectrophotometry. The protein concentrations can reach 12 mg/mL after being purified. The purified IgG reached electrophoretic pure after the SDS- PAGE electrophoresis detection., and the refining effect was excellent. Therefore the purified IgG had two bands: the heavy chain and the light chain, furthermore, the molecular weight (MW) of chicken IgG heavy chain was 53KD, and the molecular weight of light chain was 23KD. The clinical suspected IBD cases in Anhui area were detected by the antibody, and compared with the standard positive serum, the positive coincidence rate can reach a level higher than 90%, offering the essential antibody for further immunohisto chemical study.Last, IBD-1 was inoculated into the chicken embryo fibrocyte with the gotten materials and the antibody, and the optimal concentration was experimented by cellular immunity. Then the virus was inoculated into the 28-day-old non-immunized chickens and the collecting virus samples were killed after the immunization respectively 7d and 14d, by which the immunohistochemical method for detecting the IBDV was experimented and the test conditions were optimized. Scientific data were gotten by this experiment for the antigen localization, dynamic distribution regular pattern and the pathogenesis of IBDV when the chickens were infected by IBDV.