Aujeszky's disease (AD), also called pseudorabies, is an infectious viral disease caused by an alpha herpes virus and has domestic and wild pigs, as well as a wide range of domestic and wild animals, as the natural host. Aujeszky's disease virus (ADV) virions contain several envelope glycoproteins. Among them, glycoproteins gB, gC and gD are regarded as the major immunogenicity proteins and the antibodies induced by them can neutralize virus in vitro or in vivo. In this study, we investigated expression of these glycoproteins using the bacterial and baculovirus expressionn system. Successful expression of ADV glycoproteins in E. coli was confirmed by SDS-PAGE and Western blot analysis and their optimal expression condition was determined. However, the recombinant proteins generated in the bacterial expression system which lacks glycosylation process frequently lose their biological activity. We tried to express the ADV glycoproteins using the baculovirus expression vector system. The recombinant gB, gC and gD were detected at approximately 100, 60 and 50 kDa on SDS-PAGE and Western blotting, respectively. The optimal expression conditions were determined for MOI(multiplicity of infection) and post-infection days. One MOI and 4 or 5 days post-infection were the best conditions for the expression of the ADV glycoproteins in Sf21 cells. We are currently investigating the antigenicity of recombinant proteins using experimental animals.

Pseudorabies virus (PRV), a member of the Alphaherpesviridae, is the causative agent of Aujeszky’s disease in pigs. Glycoprotein B (gB) of PRV, a major constituent of the viral envelope, consists of 916 amino acids. We continuously combined three gB epitopes, E1 (aa 62-129), E2 (aa 217-282), and E3 (aa 543-737). The DNA fragment containing the PRV gB epitopes was fused with polyhedrin gene in order to generate recombinant baculovirus that expresses the recombinant polyhedra with PRV gB epitopes under the control of the Bombyx mori nucleopolyhedrovirus polyhedrin promoter. Recombinant baculoviruses were injected into fifth-instar B. mori larvae. SDS-PAGE and Western blot analyses revealed that recombinant polyhedra constitute polyhedrin and PRV gB epitopes, and that the recombinant PRV gB epitopes showed cross-reactivity against antiserum of PRV gB produced from pig. To examine the immunogenicity of recombinant PRV gB epitopes, we injected into mice as model animals. ELISA results indicated that antibody production is increased in a similar manner in the injection of recombinant polyhedra with PRV gB epitopes, either injected recombinant polyhedra as a granule form antigen without adjuvant or injected recombinant polyhedrin as a soluble form antigen with adjuvant. Taken together, these data show that PRV gB epitopes were produced as a granule form antigen by fusing recombinant polyhedra in baculovirus-infected silkworm larvae and displayed the immunogenicity in mice, indicating the efficacy of the granule form antigen as a PRV gB vaccine.