Rescue of duck-origin virulent Newcastle disease virus from cloned cDNA and stable expression of the red fluorescent protein

Ducks are generally considered as potential reservoirs for different genotypes of Newcastle disease virus (NDV) and to be resistant even to velogenic NDV strains. However, outbreaks of highly virulent genotype VII NDV lethal to ducks have been frequently reported in China in recent years. But until now, the pathogenesis and potential vaccine of duck-origin genotype VII NDV are not known. In this study, a reverse genetics system using the prevalent high virulence genotype VIId isolate SS1 was constructed. Based on this system, a red fluorescent protein (RFP)-expressing virus was generated by inserting an additional transcription cassette coding for the RFP between the noncoding region of P and M genes. The rescue of the recombinant viruses was confirmed by western blotting, fluorescence microscopy and genetic marker detection. In addition, the replication kinetics, biological characteristics and pathogenicity of the rescued viruses were indistinguishable from the parental wild-type virus. Moreover, the recombinant virus rSS1-RFP could efficiently replicate in most of the duck tissues, especially in duck immune organs. The results obtained suggest that this reverse genetics system will provide a useful tool for the analysis of duck-origin NDV pathogenesis and dissemination, as well as preparation for novel vaccine vector or genotype-matched NDV attenuated vaccines used in ducks.