Isolation And Identification Of Avian Leukosis Viruses Subgroup J,b And Preparation Of Nucleic Acid Probe

Avian leukosis virus (ALV) is the most common naturally occurring avian retrovirus that can cause a variety of neoplastic disease in chickens. In addition to causing neoplasia, ALV is known to be associated with reduced productivity and other production problems in affected flocks. ALV is one the most important pathogens and is widely distributed in the world in chiken industry. ALV causes transient severe aplastic anemia due to destruction of erythroblastoid cells and generalized lymphoid atrophy with a concomitant immunosuppression. It causes great economic losses in poultry industry as a result of co-infection and secondary infection. Avian leukosis viruses, isolated from chickens, are classified based on their host range, cross-neutralization, and viral interference into six groups (A, B, C, D, E, and J). So it is important to reaserch ALV systematically. In this study, we systematically tested Avian leukosis virus subgroup B (ALV-B) and Avian leukosis virus subgroup J (ALV-J) in aspects of dot blot detection methods, epidemiology and genetic variation and pathogenicity of a pandemic strain.Two strains of ALV-B were isolated for the first time in China Hy-line White on the cultured DF-1 cells which were inoculated tissue samples from by an ELISA assay, a histopathology examination and a PCR-based diagnosis. The results from the ELISA assay indicated that the positive rate of serum antibodies to ALV-B and ALV-J were 16.3% (15/92) and 13% (12/92), respectively. The histopathological examination indicated that two types of tumor cells existed at same focus in liver and spleen, which mainly were myelocytoma cells and lymphosarcoma cells. The PCR-based diagnosis were performed as follows: the cellular DNA was extracted from the infected DF-1 cells; the specific fragments of 1100bp and 924bp were obtained by a PCR system with the diagnostic primers of ALV-B and ALV-J; and the PCR results for ALV-A, MDV and REV were all negative. Then, the amplified fragments of the two ALV-B stains were partially sequenced and shown an identity of 93.2%and 94.8% with the prototype strain of ALV-B (RSV Schmidt-ruppin B). The identities of two ALV-J strains with the prototype strain HPRS-103 were 96.9%and 91.5%;The identities of two ALV-J strains with the American prototype strain at 85.9% and 81.5%. The study showed that ALV-B was isolated for the first time from the ALV-J infected commercial layer flocks in China. It also indicated that the chance of genetic recombination among various subgroups of ALV was increased.According to the genomic sequences of gag and env genes of ALV-B and ALV-J published in Genbank, the gp85 genes were synthesized directly. Then the genes were labelled by DIG with random primer method as probe for detection of ALV. The dot blot hybridization assay result of specificity showed ALV was positive, but other nucleotide extracted from MDV, REV were negative. So the DIG-labeled probe could be used to detect the ALV.