Establishment of a Reverse Genetics System for Rotavirus Vaccine Strain LLR and Developing Vaccine Candidates Carrying VP7 Gene Cloned From Human Strains Circulating in China

Abstract

Human rotavirus A (RVA) causes acute gastroenteritis in infants and young children. The LLR RVA vaccine, which licensed in 2000 and widely used in China, significantly reduced rotavirus disease burden in China. With the changing of RV circulating strains and the emergence of new genotypes, the LLR vaccine against RVGE needed to be upgraded. In this study, we aimed to establish an RG system for the RVA vaccine strain LLR (G10P[15]). Transfection with plasmids expressing 11 genomic RNA segments of LLR along with the pCMV/868CP helper plasmid, resulted in rescue of the infectious virus with an artificially introduced genetic marker on its genome, indicating that an RG system for the LLR strain was successfully established. Furthermore, the plasmid-based reverse genetics system was used to generate lamb RVA reassortants with VP4 or VP7 genes derived from human RVA strains in China, which were not previously adapted to cell culture. We were able to rescue the six VP7 (G1, G2, G3, G4, G8, and G9) mono-reassortants, but no VP4 (P[4] or P[8]) mono-reassortant was rescued. The six VP7 reassortants covered all G-genotypes currently circulating in China and stably replicated in MA104 cells, which should be exploited as the next-generation rotavirus vaccines candidates in China. Furthermore, the LLR RG system in this study will be a useful vaccine vector for intestinal pathogens such as norovirus and Vibrio cholerae.