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The VP6, the group antigenic rotavirus (RV), is highly conserved and the most abundant, constituting about 39% of the viral structure proteins by weight. The high degree of identity (>87%–99%) in the primary amino acid sequences suggests VP6-based vaccines could potentially provide heterotypic protection. Although some efforts have been made toward producing recombinant rotavirus VP6 vaccines, the native VP6 is still unsatisfactory as an optimal vaccine. The major neutralizing antigenic epitopes that exist on VP4 or VP7 are not on the native VP6, and as a vector the native VP6 lacks insertion sites that can be used for insertion of foreign epitopes. In this study, a new foreign epitope presenting system using VP6 as a vector (VP6F) was constructed on the outer surface of the vector six sites that could be used for insertion of the foreign epitopes created. Using this system, three VP6-based VP4 epitope chimeric proteins were constructed. Results showed that these chimeric proteins reacted with anti-VP6 and -VP4 antibodies, and elicited antibodies against VP6 and VP4 in guinea pigs. Antibodies against VP6F or antibodies against the chimeric proteins neutralized RV Wa and SA11 infection in vitro. It is optimistic that the limitation for using the native VP6 as a vaccine candidate or vector will be solved with our proposed approach. It is expected that this VP6-based epitope presenting system and the VP6-based VP4 epitope chimeric proteins will be valuable for and contribute to the development of novel RV vaccines and vaccine vectors.

A G2P[4]/NSP4[A] rotavirus strain TB-Chen was isolated from a 2-year-old patient hospitalized with acute gastroenteritis in Kunming, China. The strain TB-Chen was demonstrated having group A-specific antigenicity, a “short” (subgroup II) electropherotype. To investigate its overall genomic relatedness and to determine which group it belonged, the complete genome of strain TB-Chen was determined. Genomic comparison based on amino acid sequence identity and phylogenetic analysis revealed that all 11 gene segments of strain TB-Chen were highly identical (>91.80%) with the representative G2P[4]/NSP4[A] human strains DS-1, S2, NR1 and IS2, suggesting that this rotavirus strain was derived from human host. Besides, almost all the available representative rotavirus gene segments among group A were analyzed and identified within 15 G-types, 28 P-types, and 6 NSP4 genotypes. This is the first report of group A rotavirus genomic analyses in China and the findings have important implications for rotavirus vaccine development.

Rotavirus capsid protein Vp4 plays an important role in the virus adhering and entering the cells. In this study, a Vp4 gene cloned from a rotavirus strain TB-Chen was highly expressed in E. coli BL21 (DE3). The results of the Western blot showed that the protein possesses specific immuno-reactivities and can be specifically recognized by guinea pig antibodies against rotavirus strain SA11 or Wa. Some Vp4 dimers were formed during renaturation. These data obtained from this study provide a strong basis for further study on the structure and function of the Vp4.

VP7 of group A rotavirus (RVA) contains major neutralizing epitopes. Using the antigenic protein VP6 as the vector, chimeric proteins carrying foreign epitopes have been shown to possess good immunoreactivity and immunogenicity. In the present study, using modified VP6 as the vector, three chimeric proteins carrying epitopes derived from VP7 of RVA were constructed. The results showed that the chimeric proteins reacted with anti-VP6 and with SA11 and Wa virus strains. Antibodies from guinea pigs inoculated with the chimeric proteins recognized VP6 and VP7 of RVA and protected mammalian cells from SA11 and Wa infection in vitro. The neutralizing activities of the antibodies against the chimeric proteins were significantly higher than those against the vector protein VP6F. Thus, development of chimeric vaccines carrying VP7 epitopes using VP6 as a vector could be a promising alternative to enhance immunization against RVAs. [Image: see text]

It has been demonstrated that the immunodominant region of the HCV core protein and the hepatitis B surface antigen (HBsAg) have high degree of reactivity. In order to construct a chimeric protein that carries HCV and HBV epitopes and possesses immunogenicity to both HCV and HBV, four epitopes derived from residues aa2-21 (epitope C1), aa22-40 (epitope C2) of the core protein, residues aa315-328 (epitope E) of E1 protein of HCV, and residues aa124-147 (epitope S) of HBsAg were chosen to be displayed in a conformation-specific manner on the outer surface of the Flock House virus capsid protein and expressed in E. coli cells. The reactivity of these epitopes with antisera from hepatitis C and hepatitis B patients and induction of immune response in guinea pigs were determined. The results showed that when displayed in this system, the chimeric protein carrying only epitope S could react with anti-HBsAg positive human sera, elicit an anti-HBsAg response in guinea pigs. The chimeric protein carrying epitopes C1, C2 and E could react with antibodies to different HCV genotypes, elicit an anti-HCV response in guinea pigs. The chimeric protein carrying epitopes C1, C2, E, and S could react with antibodies against HCV and HBV, elicit anti-HCV and anti-HBsAg responses in guinea pigs. The results suggested that these epitopes displayed in this form could be considered for development of epitope-based vaccines against HCV/HBV infections.