Tick-borne encephalitis (TBE) is a severe disease affecting thousands of people throughout Eurasia. Despite the use of formalin-inactivated vaccines in endemic areas, an increasing incidence of TBE emphasizes the need for an alternative vaccine that will induce a more durable immunity against TBE virus (TBEV). The chimeric attenuated virus vaccine candidate containing the structural protein genes of TBEV on a dengue virus genetic background (TBEV/DEN4) retains a high level of neurovirulence in both mice and monkeys. Therefore, attenuating mutations were introduced into the envelope (E315) and NS5 (NS5654,655) proteins, and into the 3′ non-coding region (Δ30) of TBEV/DEN4. The variant that contained all three mutations (vΔ30/E315/NS5654,655) was significantly attenuated for neuroinvasiveness and neurovirulence, and displayed a reduced level of replication and virus-induced histopathology in the brains of mice. The high level of safety in the central nervous system indicates that vΔ30/E315/NS5654,655 should be further evaluated as a TBEV vaccine.

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In an effort to derive an efficacious live attenuated vaccine against tick-borne encephalitis, we generated a chimeric virus bearing the structural protein genes of a Far Eastern subtype of tick-borne encephalitis virus (TBEV) on the genetic background of recombinant dengue 4 (DEN4) virus. Introduction of attenuating mutations into the TBEV envelope protein gene, as well as the DEN4 NS5 protein gene and 3 0 noncoding region in the chimeric genome, results in decreased neurovirulence and neuroinvasiveness in mice, and restricted replication in mouse brain. Since TBEV and DEN4 viruses are transmitted in nature by ticks and mosquitoes, respectively, it was of interest to investigate the infectivity of the chimeric virus for both arthropod vectors. Therefore, parental and chimeric viruses were tested for growth in mosquito and tick cells and for oral infection in vivo. Although all chimeric viruses demonstrated moderate levels of replication in C6/36 mosquito cells, they were unable to replicate in ISE6 tick cells. Further, the chimeric viruses were unable to infect or replicate in Aedes aegypti mosquitoes and Ixodes scapularis tick larvae. The poor infectivity for both potential vectors reinforces the safety of chimeric virus-based vaccine candidates for the environment and for use in humans.