| H9 subtype avian influenza virus is the main subtype that endanger poultry industry in china, which characterizes as elevated body temperature of chickens,respiratory disease,egg production decline and the increasing abnormal eggs.In recent years, studies have shown that suspected cases continue to occur after making H9 subtype avian influenza virus immunization,resulting in a certain amount of economic loss and bringing a great challenge to epidemic prevention. The disease is also easily induced E. coli disease,mycoplasma disease, Newcastle disease and other disease.Therefore,in order to improve immune efficiency, united immunization on avian influenza and Newcastle disease and prevention in cross-infection become an important means of immunization in large-scale farms.In this study, the suspicious epidemic materials of the chicken was collected in a farm that may be infected with avian influenza,virus was isolated and purified and then its gene was cloned to identify it. H9 subtype avian influenza virus named XY-14 that identified by mang means was immune together with NDV IV, to learn about the molecular immunogenic and biological characteristics of H9 subtypes of avian influenza strains, the key point of the study is the laws of the body dynamics antibody levels.1、epidemic materials was respectively collected from different regions,virus was isolated and cultivated by chicken embryo and screened premilinary by hemagglutination and hemagglutination inhibition testing and the method of RT-PCR. The isolated virus was 50-fold concentrated and purified by sucrose density gradient centrifugation, and then amplified with universal primer and avian influenza H9 subtype of avian influenza universal primers, bright stipe in 229 bp and 732 bp appeared, which are consistent with the expected size of the target fragment. HA, NA, NP, M, NS gene fragments were cloned, bright bands in 1740 bp, 1470 bp, 1565 bp, 1027 bp and 890 bp were appeared,which were also consistent with the expected location,so it was concluded that the strain is H9 subtype avian influenza,named A / chicken / Xin Yang / 04/2014(H9 Subtype), referred to as the XY-14 strain.2、The titer of HA is 210 for XY-14,the titer of HI is 210,then after H9 subtype avian influenza serum neutralization test, the allantoic fluid collected has no hemagglutination.3、Toxicity test of the strain showed that the result of ICPI(1-day-old chicken brain index stastics) is 0.18;IVPI =0;which indicates that XY-14 is attenuated strain.4、Inactivate XY-14 virus strain; Immunization together with Newcastle disease IV vaccine with 0.3m L dose ND and AIV H9 antibodies will come up during the first week.ND antibodies will comes at its peak by 11log2 in the 3th week, and the antibody levels are still reach 6log2 in the 22 th week. AIV H9 antibody levels reach the peak in 4th week, reach up to 11.33log2. united immunization with 0.3m L dose can provide long-term high levels of antibody protection. Therefore, 0.3m L is the more appropriate immunization dose.The results showed that united immunization can provide higher level antibody protection than the single immunization with ND IV vaccine. and can continue for a long time with high antibody titers. |
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