Japanese encephalitis is prevalent throughout the temperate and tropical regions of Asia and is caused by the Japanese encephalitis virus (JEV), a mosquito-borne viral pathogen. The plaque reduction neutralization test (PRNT) is currently recommended as the gold standard test for detecting human antibodies against JEV. The plaque assay is the most widely used method for detecting infectious virions and involves counting discrete plaques in cells. However, it is time-consuming, and results can be subjective (owing to analyst variability during manual plaque counting). The focus reduction neutralization test (FRNT), which is based on an immuno-colorimetric assay, can be used to automatically count foci formed by the JEV. Here, we compared the efficacy of PRNT and FRNT in measuring the neutralizing antibody titers using 102 serum samples from vaccinated and unvaccinated individuals. We observed positive correlations between these neutralization assays against the Nakayama and Beijing strains (R² = 0.98 and 0.77, respectively). Thus, FRNT may be preferable to PRNT for evaluating the efficacy of JEV vaccines in large-scale serological studies.

Mumps virus (MuV) causes an acute contagious human disease characterized by swelling of the parotid glands. Despite the near elimination of mumps in many countries, the disease has recurred, even in vaccinated populations, especially adolescents. Immunization effectivity of the genotype A vaccine strain Jeryl Lynn (JL) is declining as genotype A is no longer predominant; therefore, a new vaccine strain and booster vaccine are required. We generated a cell culture-adapted MuV genotype F called F30 and evaluated its immunogenicity and cross-protective activity against diverse genotypes. F30 genome nucleotide sequence determination revealed changes in the NP, L, SH, and HN genes, leading to five amino acid changes compared to a minimally passaged stock (F10). F30 showed delayed growth, smaller plaque size in Vero cells, and lower neurotoxicity in neonatal mice than F10. Furthermore, F30 was immunogenic to other genotypes, including the JL vaccine strain, with higher efficacy than that of JL for homologous and heterologous immunization. Further, F30 exhibited cross-protective immunity against MuV genotypes F and G in Ifnar^−/− mice after a third immunization with F30 following two doses of JL. Our data suggest that the live-attenuated virus F30 could be an effective booster vaccine to control breakthrough infections and mumps epidemics.

Mumps is an acute infectious disease caused by the mumps virus (MuV). Despite high global vaccination coverage, mumps outbreaks continue to occur, even in vaccinated populations. Therefore, we aimed to identify candidate vaccines that can induce an immunogenic response against diverse MuV genotypes with greater efficacy than the currently available options. Vaccine candidates were sourced using formalin-inactivated viral strains. The inactivated vaccines were administered to BALB/c mice (through a primer and booster dose administered after a three-week interval). We tested the neutralizing antibodies of the candidate vaccines against various MuV genotypes to determine their overall efficacy. The formalin-inactivated F genotype vaccine was found to have higher cross-neutralizing titers against genotypes F, H, and G as well as significant Th1 cytokines responses, IFN-γ, TNF-α, and IL-2 than the Jeryl Lynn (JL) vaccine. Our findings suggest that the inactivated F genotype mumps vaccine has higher immunogenicity than the JL vaccine against diverse circulating MuVs.

The mumps virus (MuV) causes a highly contagious human disease characterized by swelling of the parotid glands. Although the administration of an attenuated Jeryl Lynn (JL) MuV vaccine shows efficacy in reducing the incidence of MuV infection, sporadic mumps outbreaks still occur in vaccinated populations. We have previously established that an inactivated F genotype mumps vaccine has a higher neutralizing antibody titer against diverse circulating mumps viruses in mice. Here, we aimed to develop a vaccination strategy to enhance the immune response for MuV and assess the effects of heterologous vaccination compared with homologous approaches. We administered an inactivated F genotype mumps vaccine booster following a homologous prime-boost regime and compared its efficacy with three doses of homologous JL vaccine in mice. We demonstrated robust stimulation of neutralizing antibodies and cellular immune response of interferon-γ-secreting cytotoxic T cells following administration of an inactivated F genotype mumps vaccine booster after a homologous prime-boost regime with JL. Compared with the homologous prime-boost regime, this heterologous prime-boost regime showed protective efficacy against the F genotype of MuV. These findings suggest that the heterologous vaccination strategy based on the administration of an inactivated F genotype mumps vaccine provides more effective cross-protection against circulating wild-type mumps viruses than homologous vaccination.