Isolation And Partial Sequencing Of Avian Leucosis Virus Subgroup J In Layer Flocks

Avian leucosis virus subgroup J in chickens ( ALV-J) is a tumor disease induced by virus, namely Myelocytomatosis. Since ALV-J was discovered, it has just been reported in meat-type chickens, resulting in considerable losses of chickens industry all over the world. In 2002, Xu Binrui, professor of China Agricultural University, discovered ALV-J in layer chickens. Naturally infected layer flocks showed the symptom of tabes and serious anemia, decrease of egg production, and the morality was as high as 10%. Maldevelopment was present in testes, ovaries, and oviducts. It caused considerable losses of layer chickens industry. Specific protein gp85 of ALV-J was detected through pathology, immunohistochemistry and PCR method. The layer chickens natural infected with ALV-J were first dicovered and reported in the world.In the study isolation of virus was performed using the samples of layer chickens naturally infected with ALV-J. The tissue samples (hearts, livers, spleens etc) of illed chickens were filtrated and inoculated into chicken embryo fibroblast (CEF). The CEFs were detected using the indirect fluorescence assay (I FA) with specific monoclonal antibody against ALV-J gp85 at 7 day after inoculation. Specific fluorescence was detected in the cytoplasm of CEFs and the caryon was unstained. Specific envelope protein gp85 was detected by using immunohistochemistry. For further detection RT-PCR was performed on the CEF inoculated with the samples. A 545bp specific product was amplified using primer set H5 and H7 that was designed in 3' region of pol gene and 5' region of the gp85 encoding gene respectively. The product was as same size as expected and was sequenced by TaKaRa company. The sequence data showed that the identity of the nucleotide sequence was 97.4% and that of the amino acid sequence was 96.1% compared with that of the prototypical strain of ALV-J HPRS-103. The results of RT-PCR illuminated that the gp85 gene of ALV-J isolated from egg-type chickens have some variation compared with that of prototype HPRS-103 strain.The results of this study demonstrated that ALV-J was successfully isolated from naturally infected egg-type chickens for the first time. The isolation of ALV-J in layer chickens will develope a new member for previously known diseases that affected the productivity of chickens, meanwhile, it will also provide theory and practice basis for better preventing, controlling the occurrence of ALV-J and decreasing the economic losses.