Virulence and Cross-Protection Conferred by an Attenuated Genotype I-Based Chimeric Japanese Encephalitis Virus Strain Harboring the E Protein of Genotype V in Mice.

Japanese encephalitis virus (JEV) genotype V (GV) emerged in China in 2009, then South Korea, and has since spread to other regions in Asia and beyond, raising concern about its pathogenicity and the cross-protection offered by JEV vaccines against different genotypes. In this study, we replaced the structural proteins (C-prM-E) of an attenuated genotype I (GI) SD12-F120 strain with those of a virulent GV XZ0934 strain to construct a recombinant chimeric GI-GV JEV (JEV-GI/V) strain to determine the role of the structural proteins in virulence and cross-protection. The recombinant chimeric virus was highly neurovirulent and neuroinvasive in mice. This demonstrated the determinant role of the structural proteins in the virulence of the GV strain. Intracerebral or intraperitoneal inoculation of mice with JEV-GI/V-E5 harboring a combination of substitutions (N47K, L107F, E138K, H123R, and I176R) in E protein, but not mutants containing single substitution of these residues, resulted in decreased or disappeared mortality, suggesting that these residues synergistically, but not individually, played a role in determining the neurovirulence and neuroinvasiveness of the GV strain. Immunization of mice with attenuated strain JEV-GI/V-E5 provided complete protection and induced high neutralizing antibody titers against parental strain JEV-GI/V, but partial cross-protection and low cross-neutralizing antibodies titers against the heterologous GI and GIII strains in mice, suggesting the reduced cross-protection of JEV vaccines among different genotypes. Overall, these findings suggested the essential role of the structural proteins in determination of the virulence of GV strain, and highlighted the need for a novel vaccine against this newly emerged strain. IMPORTANCE The GV JEV showed an increase in epidemic areas, which exhibited higher pathogenicity in mice than the prevalent GI and GIII strains. We replaced a recombinant chimeric GI-GV JEV (JEV-GI/V) strain to determine the role of the structural proteins in virulence and cross-protection. It was found that the essential role of the structural proteins is to determinethe virulence of the GV strain. It is also suggested that there is reduced cross-protection of JEV vaccines among different genotypes, which provides basic data for subsequent JEV prevention, control, and new vaccine development.