Packaging of Rift Valley fever virus pseudoviruses and establishment of a neutralization assay method

Li, Yuetao; Zhao, Yongkun; Wang, Cuiling; Zheng, Xuexing; Wang, Hualei; Gai, Weiwei; Jin, Hongli; Yan, Feihu; Qiu, Boning; Gao, Yuwei; Li, Nan; Yang, Songtao; Xia, Xianzhu

Currently, pseudovirus packaging systems are primarily developed based on HIV lentiviral vectors and the murine leukemia virus [1, 9, 15] , which have the advantages of broad suitable host ranges and high packaging efficiencies. The present study used a lentiviral packaging system and recombinant expression plasmids encoding RVFV structural proteins to construct RVFV pseudoviruses. The electron microscopy observations and western blotting results all indicated the successful construction of RVFV pseudoviruses. Additionally, the infective activity of the obtained RVFV pseudoviruses was inhibited by specific antibodies, indicating that the pseudoviruses could be used to detect RVFV-neutralizing antibody titers. Moreover, the RVFV pseudovirus neutralization assay established in this study effectively evaluated antibody titers in positive mouse serum. The pseudovirus packaging experiment showed that the passage time and status of the 293T cells and the transfection reagents, plasmid purity, gene features of the structural proteins, and efficiency of the eukaryotic expression plasmids all greatly influenced the pseudovirus packaging efficiency and affected the titer levels and stability of the packaged pseudoviruses. Therefore, the pseudovirus neutralization assay established in the present study can be used for clinical monitoring and diagnosis of RVFV infection and for assessment of the effects of RVFV vaccines. Furthermore, the pseudovirus can be used to replace the natural virus in infection mechanism studies. However, the packaging conditions should be optimized to increase the lentiviral packaging efficiency and infection titers.