Sub-fragments of the envelope gene are highly protective against the Japanese encephalitis virus lethal infection in DNA priming--protein boosting immunization strategies.

The envelope (E) gene of Japanese encephalitis virus (JEV) plays a major protective role against JEV infection. In order to locate the part of E gene that is responsible for this protection, an N-terminal fragment EA (nucleotide number 933-1877 bp of JEV genome) and a C-terminal fragment EB (nucleotide number 1851-2330 bp of JEV genome) from E gene were prepared. Both of these fragments were used in the form of recombinant proteins (rEA and rEB) and plasmid DNA (pEA, pM15EA and pEB) for immunizations. Recombinant EA protein (rEA) was previously found to be non-protective because it was expressed in an insoluble form. Plasmid EA (pEA) was also found to be non-protective unless it is preceded by a 15 mer signal peptide derived from the very C-terminal of the membrane gene (M) of JEV to form pM15EA plasmid indicating the importance of the signal peptide in the expression of EA immunogenicity. Although pM15EA and pEB are both immunogenic and protective against JEV lethal infection, the protection by both fragments however is not optimal. Even when pM15EA and pEB were used together for immunization, maximum protection as those induced by control vaccine was not achieved. However, if individual fragments (EA or EB) were used in a DNA priming-protein boosting or protein priming-DNA boosting strategy, high levels of protection were achieved by both fragments. This was especially true for EA fragment where the level of protection against JEV lethal infection was equal to that induced by commercially available vaccine alone. The protection correlated very well with the neutralizing antibody titers and the T helper cell involved in this process in mainly the Th1 type.