Development Of Agglutination Antigen Of Avibacterium Paragallinarum And Evaluation Of Immunoprotective Effect Of An Inactivated Vaccine On Epidemic Strains

Infectious coryza is an acute respiratory disease caused by Avibacterium paragallinarum(Apg).It is characterized by facial swelling,nasal discharge and conjunctivitis,and can lead to growth retardation and decreased egg production.In recent years,there are still frequent outbreaks of infectious coryza in chickens vaccinated with inactivated vaccine,which has caused huge losses to the poultry industry.Vaccine immunization is the main strategy to prevent chicken infectious coryza.Hemagglutination inhibition antibody titer is an important index to evaluate the immune effect of vaccine,but there is still a lack of commercial agglutination antigen.In this study,on one hand,the hemagglutination inhibition test antigen of Apg was developed.On the other hand,the immunoprotective effect of inactivated vaccine against current epidemic strains was evaluated with the prepared agglutination antigen.1.Development of hemagglutination inhibition test antigen of Avibacterium paragallinarum.In this study,the development methods of hemagglutination inhibitory antigens against types A,B and C were optimized and established by standard strains 221(type A),Spross(type B)and H-18(type C).Serum type A strain can agglutinate chicken erythrocytes fixed with glutaraldehyde without special treatment,and there was no difference in HA titer before and after treatment.The best treatment condition of s type B was 5U/mL hyaluronidase at 37℃ for 2 h.Furthermore,the strains with high hemagglutination and good stability were selected from the epidemic strains with serotypes A,B and C.Among them,the HA titer of 2019/JS15(type A)was 28,the HA titer of JS37(type B)was 27,and the HA titer of JS80(type C)was 27.At the same time,we prepared the freeze-dried Apg hemagglutination inhibition test antigen.The HA titer of serum type A Apg freeze-dried hemagglutination inhibition test antigen could be maintained stably for more than 4 months at-20℃.The preservation conditions of serum type B and type C Apg freeze-dried hemagglutination inhibition test antigens need to be further explored.2.Evaluation of immunoprotective effect of inactivated vaccine on epidemic strains.The purpose of this study is to verify immunoprotective effect of trivalent inactivated vaccine of infectious coryza(Indonesia Medion.)on the epidemic strains of Apg through animal experiments.One hundred chickens were divided into four groups:single immunity group(A1,B1,C1),double immunity group(A2,B2,C2),challenge control group(A3,B3,C3)and blank control group(D).The double immunization group received the first immunization at the age of 60 days and the second immunization at the age of 110 days.The single immunization group was immunized at the age of 110 days.The antibody titer of the vaccine was detected by HI test using the agglutination antigen prepared in the previous chapter.At the age of 130 days,groups A1,A2 and A3 were challenged with 2019/HB65(type A),B1,B2 and B3 with 2019/HB68(type B)and C1,C2 and C3 with 2020/JS80(type C)at the dose of 1 × 10 7 CFU/0.2 mL.The HI titer of type A antibody was higher than that of type B and C.The titer of type A antibody was 13.1 log2 in the third week after the twice immunization.Antibody of type B could not be detected after the once immunization,but the antibody titer was positive in two weeks after the twice immunization,and the titer was 6.1 log2.The antibody titer of C was 10.2 log2 in the third week after the twice immunization.The infection of chickens in C3 group was the most serious.After challenge,the clinical symptom score in the double immunity group was lower than that in the single immunity group,and significantly lower than that in the black control group.Especially after 2020/JS80(type C)challenge,the double immunity group did not have any clinical symptoms.The egg production rate of the single immunity group and the double immunity group was higher than that of the challenge control group,and there was no significant difference between the single immunity group and the blank control group.The egg production rate of experimental chickens in A3 and C3 groups decreased significantly,but there was no significant change in all immunity groups.Both the single and double immunization of the vaccine could reduce bacteria load after challenge,especially in the double immunity group.The results of isolation of Apg from suborbital sinus swabs showed that bacteria were all reisolated in control groups A3,B3 and C3 on the 7th day after challenge,and Apg could be partly reisolated in groups A1,A2,B1 and B2(4/5).However,Apg were not reisolated in C2 group at all and chickens achieved complete protection.On the 14th day after challenge,isolated rate of all groups decreased,and there was no difference among groups except group C.In summary,agglutination antigens with high HA titer and good stability were prepared in this study,and the protective effect of inactivated vaccine on isolates was evaluated.After immunization,the level of HI antibody in the double immunity group was significantly higher than that in the single immunization group.After the epidemic strain was challenged,the protective effect of double immunization was better than that of single immunization.it could significantly reduce the clinical symptoms,pathological changes and the amount of bacteria load,especially the complete protection of type C isolates.Although the disease still occurred in the immunization group,the body weight and egg production rate remained unchanged.