The Pathogenicity Of Three Duck Reovirus On Duck Embryo Fibroblasts

In this study, duck embryo and Duck Embryo fibroblast(DEF) were infected with three virus strains(HP09318strain、HP080421strain. HP081122strain) isolated in our laboratory. The proliferation of the virus, pathological changes and TNF-a content changes were studied using cell culture technology, light microscopy, electron microscopy, real-time RT-PCR and ELISA. The results are as follows:The result of artificial infection of duck embryo:9-11day old duck embryo were given allantocherion injection of three DRV strains. The result showed that embryo proper died in three to six days with symptoms of dysplasia, hemorrhage, and yellowish-white necrotic spots on the surface of liver.Pathological changes of DEF infected with three strains: The infection of monolayers of DEF with DRV causes a cytopathic effect(CPE) at24hour post infection(hpi). the infected cells manifest rounding, vacuoles in the cytoplasm and some of the cells were lysed. Then, CPE became more serious at48hpi. There were a great quantity of died cells floating in the medium, including multinucleated giant cells. HE staining was performed on sections, the infected cells showed reticular formation, fragmented nucleus and some of them has lysed, giant cells with multiple nucleus were also seen. Eosinophilic intranuclear inclusions appeared in cytoplasm. Cryosections from infected cells were prepared, and the CPE was monitored by electron microscopy. There were some vesicles in various size appeared in infected cells, and lamellar structures and Virus particles were also observed.Proliferation of DRV in DEF:cells, supernatant and cells supernatant mixture were collected at12,24,48,72,96hpi. The results of RT-PCR showed that there were a spot of virus at12hpi. After12hpi, there was a gradual increase. Virus in cells and mixture arrived peak at24hpi, but the virus in supernatant appeared peak at48hpi. Then the virus decreases gradually after arrival of the peak.TNF-a changes of DEF infected with three strains:the result of ELISA indicated that before12hpi TNF-a in infected cells was lower compared to control, then TNF-a increased gradually. There was a significant increase in TNF-a production in infected cells compared to uninfected control at24hpi, but TNF-a in infected cells was lower compared to control at96hpi. The result of real-time RT-PCR showed that expression of TNF-a mRNA increased significantly from24to48hpi, and decreased after96hpi.Conclusion: three virus strains(HP09318strain, HP080421strain, HP081122strain) has concurrent morphology, and could infect duck embryo and DEF, cause similar pathological changes. The proliferation of the virus, the extent of the CPE and changes of TNF-a levels were positively correlated.