Isolation And Identification Of Guangxi Field Strains Of Marek's Disease Virus And Its Co-Infection With Other Viruses

Marek's disease (MD) is a highly contagious lymphoproliferative diseasecaused by Marek's disease virus (MDV), which cans lesion neural and viscerallymphomas, bursa and thymus atrophy, splenomegaly and skin neoplasms.Recently MD outbreaks could also be a problem in the industry even though thebirds had been vaccined with CVI988/Rispens.To investigate the prevalence, distribution, virulent variation and genevariety of epidemic field strains of MDV in Guangxi, the combination of cellculture, indirect immunofluorescrence assay (IFA) and polymerase chainreaction (PCR) was carried out. Three local MDV field isolates (named GXY1,GXY2, GXYL1) were isolated from different Three-Yellow chicken flocks withthe peripheral blood lymphocytes (PBLs) and feather follicles of birdsexperensed MD which had been vaccined with CVI988 vaccine.In the cell cultures of these isolates, GXYL1 infected DEFs showedpositive reaction when IFA was run with monoclonal antibodies (McAbs)11B118 of reticuloendotheliosis virus (REV), and REV LTR can amplified byPCR. That meant this MDV isolate was mixed the infectious REV, and the casewas co-infected by both MDV and REV. Infected DEFs of isolate GXY1showed negative reaction with REV McAbs but REV LTR, REV env and fowlpoxvirus (FPV) 4b were amplified. When DEF monolayer was infected with thecell culture supernatant of GXY1, IFA was negative with REV McAbs, whichmean this MDV isolate didn't contain free REV, but a free FPV with theintegrated REV LTR, REV env was co-infected in the sample. However, DEFsinfected with GXY2 showed negative with REV McAbs but REV LTR wasamplified, indicated that isolate GXY2 was a MDV field strain with an REVLTR integrated into the genome. To our knowledge this is the first report ofrecombinant field strain of MDV isolated from the feather follicles of MD chicken. These results manifested that the co-infection phenomena of MDV andother viruses especially with REV was common, and some MDV field strainshad been integrated with the REV genome fragments.Earlier studies have determined that the repeat region of MDV-1 Eco-Qgene (meq), which structurally resembles the fos/jun family of transcriptionalactivators was an oncogene. By compared with the sequences of MDV referencestrains GA and RB1B, the Guangxi MDV isolates possesd six unique sitemutations within the open reading frame (ORF) of meq gene on the 77, 80, 115,139, 176, 217 amino acids, and the 80, 115, 217 amino acids all of the Guangxiisolates mutated to Y, A and A respectively different from the two reference andvaccine strains, which showed off invariably regularity. And for the 139 aminoacid mutation was present in the most Chinese isolates except for only one fieldisolate on abroad, which may be correlated with virulence. While for the 176amino acid mutation was only found in the Chinese isolates that located in arepeat sequence of PPPP, which was reported to be correlated with virulence ofMDV and resulted in PRPP. It seems the some regularities exist between suchmutations in our Chinese MDV isolates and virulence of MDV, the mutationsare very significance for us to research the MDV-1 isolates from China.The research implies that the recently commercial flocks carried a mixedMDV and retrovirus-infection or FPV, and occurred to various recombinationevents. The sequence analysis of meq oncogene indicated, for the first time, thatit occurs a special site mutation in Chinese MDV field isolates. These results ofresearch provided useful informations to us to know and master the mechanismof virulent variation and biological characteristics of MDV.