Adaptability of Fowlpox Virus in Chicken Embryo Passage Cells

[Objective] To observe whether fowlpox virus （FPV） can proliferate in chicken embryo passage fibroblasts or not and then try to use chicken embryo passage fibroblasts to replace primary chicken embryo cells for FPV culture. [Method] Primary chicken embryo fibroblasts were prepared and subcultured. After FPV were inoculated on the 20th passage fibroblasts, cytopathy was observed. Then, the FPV culture was identified and determined quantificationally. [Result] Specific cytopathy appeared in the FPV-inoculated chicken embryo passage fibroblasts. The titer of the yielded FPV culture reached the standard for production of fowl pox vaccine. Further analysis reveals that the chorioallantoic membrane lesions were caused by FPV. [ Conclusion] FPV can reproduce in chicken embryo passage fibroblasts, and the Uter of FPV cell culture can meet the pro- duction requirements of fowl pox vaccine.

Antiviral Effects of Compound Traditional Chinese Medicine on Newcastle Disease Virus

[Objective] To investigate the mechanism of compound traditional Chinese medicine （TCM） on Newcastle disease virus （NDV） and to provide a scientific basis for the reasonable usage of antiviral drugs in clinic. [Method] The compound TCM was composed of Hedyotis diffusa, Lonicera japonica Thunb, Radix astragali and Glycyrrhiza uralensis. Different dilutions of fluid extract were prepared. Its antiviral effects on NDV were observed through three inoculation ways, first, inoculation with the medicine and NDV mixture which had been incubated at 37 ℃; second, incubating chicken embryo fibroblasts （CEF） with the medicine followed by inoculation with NDV; third, inoculation with N DV followed by incubating CEF with the medicine. The A,= was determined by M]-r [ 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide） ~ method. Therapeutic indexes were used to evaluate the antiviral effects. [ Result] The minimum effective concentration of the compound TCM which acted through the three ways was 1.0 × 2^-10 1.0 × 2^-8 and 1.0 × 2^-7 g/ml, respectively. The antiviral effects of the compound TCM were the best through inoculation with the incubated medicine and NDV mixture, followed by the second method and the third method. [ Conclusion] The compound TCM can not only kill NDV directly in vitro but also inhibit viral propagation.

shRNA-triggered RNAi inhibits expression of NDV NP gene in chicken embryo fibroblast

RNA interference(RNAi)technology is a powerful tool for identifying gene functions.Chicken embryo fibroblast(CEF)is an ideal model for studying the interaction between avian viruses and their hosts.To establish a methodological platform for RNAi studies in CEF,three plasmid vectors expressing short hairpin RNAs(shRNAs)targeted against the Newcastle disease virus(NDV)NP gene were constructed.One of them,ndv1,was proven effective on blocking viral replication in CEF and chicken embryos.Four hours prior to infection with NDV,the CEF was transfected with the plasmids by Silent-fect.An unrelated shRNA sequence(HK)was used in mock transfection.The expression of a potent shRNA resulted in up to 2.3,21.1 and 9.8 fold decreases in NP gene expression at 3,6 and 9 h post infection in CEF,respectively.The ndv1 was able to completely inhibit the replication of the virus in CEF within 48 post infection.Furthermore,the pathological changes in CEF caused by NDV were delayed,and the degree of pathological changes was lighter compared with the mock transfection in the presence of ndv1.When the complex of shRNASilent-fect and NDV was co-injected into the allantoic cavity of 10-day-old embryonated eggs with 10^(5) or 10^(6) ELD50 NDV,NDV replication was decreased by 94.14% and 62.15% after 17 h,respectively.These findings suggest that the newly synthesized NP protein is critical for NDV transcription and replication and provide a basis for identifying the functions of viral genes and screening for effective siRNAs against viruses in CEF and chicken embryo by RNAi.