Pathological Development Study On Ducklings Experimentally Infected With Duck Hepatitis Virus And Distribting Regularity Of Duck Hepatitis Virus In Infected Ducklings Using Indirect Immunoperoxidase Staining (IIS) And Indirect Fluorescent Antibody(IFA)

The dissertation revolves around Duck Hepatitis Virus(DHV) isolation and identification, pathology of experimentally infection disease model and localize the virus antigens using Immunoperoxidase Staining(fluorescent antibody), development series research:①Pathogen isolation and identification of outbreak and epidemic of Duck Virus,②Pathological Development study on ducklings experimentally infected with Duck Hepatitis Virus,③Detection of duck hepatitis virus in the paraffin wax tissue splice using indirect immunoperoxidase staining & indirect fluorescent antibody and its application at antigen localization of infection ducklings tissue.The result is as follows:1. Pathogen isolation and identification of outbreak and epidemic of Duck Virus Hepatitis:Three strains of virus were isolated from the liver of dead ducklings in the disease outbreak and epidemic farms of Sichuan,which had model clinic symptom and pathological changes of Duck Virus Hepatitis,and showed positive using indirect fluorescent antibody(IFA).The virus was shown spherical, no envelope, 30nm-40nm in diameter by electron microscopy. The virus was resistant to chloroform, heat(50℃,60min) and pH3.All duck embryos inoculated with the 3rd filial generation isolation virus were died in 24-60 hours, and duck embryos serious hyperemia and hemorrhage. 10-day-old duck embryos inoculated with the mixture of DHV typeⅠantiserum and the virus, no duck embryo dead. 1-day-old ducklings were experimentally infected with three strains of virus could arise the same clinical symptom and pathological changes as natural cases, and virus could be isolated again. Liver, spleen, kidneys from dead ducklings infected with 3 strains of virus and natural cases, detected by Indirect Immunoperoxidase Staining(IIS), were positive or strongly positive. These results demonstrate that the three strains are Duck Hepatitis typeⅠVirus. This dissertation can provide the useful testing data for effective preventing DVH. 2. Pathological Development study on ducklings experimentally infected with Duck Hepatitis Virus: 1-day-old ducklings were experimentally infected with DHV-XC from intramascular injection, orally, and intranasally inoculated,and the pathological mechanism model having copied duck virus hepatitis successfully.The result of the three groups pathological change showed that: Histopathological examination revealed the hemorrhagic and necortic inflammation of the liver at originally.then proliferative inflammation was observed. Granular degeneration in epithelia of renal tubule and more infiltration of neutrophilesin in the kidney. Heavy haemorrhage and necrosis of spleen. There was limitation coagulation necrosis and councilman's bodies in pancreas. Cardiac muscle vacuolation and diffuse hemorrhage,and the lymphocytes necrosis of thymus and bursa of Fabricius,and congestive capillaries of brain tissue at the time after inoculation,and the hyperemia and shedding some of the epithelium intestine and Harderian gland.3. Establishment of indirect immunoperoxidase staining & indirect fluorescent antibody technique to the detection for Duck Hepatitis Virus in the paraffin wax tissue splice and its application at antigen localization of infection ducklings tissue.①Detection of duck hepatitis virus in the paraffin wax tissue splice using indirect immunoperoxidase staining and antigen localization of infection ducklings tissue:Antibodies against DHV-Ⅰwere acquired from rabbits vaccinated with purified DHV-Ⅰby means of sucrose gradient centrifugation. An Indirect Immunoperoxidase Staining(IIS) were established for the detection of DHV-Ⅰantigen in the paraffin wax tissue splice. Detected DHV-Ⅰin each organ tissues of dead or dying ducklings experimentally infected with DHV-Ⅰ. Using the IIS and the IFA, the positive reaction appears in liver and so on of the DHV-Ⅰinfection dead or dying ducklings, but the negative reaction appears in liver of Duck Plague Virus, duck's Riemerella Anatipestifer, E. coli, Salmonella and Pasteurella multocida infection dead and arises duck.Ducklings detected by IIS of dynamic distribution of virus antigens in vivo results showed that: Liver, spleen, kidney, pancreas, thymus, intestine, muscle and Bursa of Fabricius of ducklings from 3 groups can be detected DHV antigen at different time 12HPI,and lungs, heart, brain tissue of some deed or dying ducklings were detected DHV antigens,and the DHV antigen-positive cells were located in the cytoplasm. The sequence of the virus antigen detection in the intramascular injection group: liver, spleen,kidney, duodenum, pancreas, muscle,jejunum,thymus,bursa of Fabricius,Harderian gland,ileum,rectum,cecum,cardiac muscle,cerebrum,lung.The sequence of the virus antigen detection in the orally inoculated group: liver, spleen,kidney,pancreas,thymus,Harderian gland,bursa of Fabricius,duodenum, muscle,jejunum,ileum,rectum,cecum,cardiac muscle,lung,cerebrum.The sequence of the virus antigen detection in the intranasally inoculated group:liver, spleen, kidney, lung, pancreas, bursa of Fabricius, duodenum, jejunum, thymus, Harderian gland, muscle, ileum,rectum,cecum,cardiac muscle,cerebrum.②The results of establishment IFA detection for DHV and localization of DHV antigens indicated that the distribution regularity of Duck Hepatitis Virus in artifically infected ducklings accorded with the results of IIS,although there were several differences in some individuality.