Complementation of recombinant baculoviruses by coinfection with wild-type virus facilitates production in insect larvae of antigenic proteins of hepatitis B virus and influenza virus (baculovlrus/hepatitis B surface antigen/influenza A virus neuramlnidase)

We describe the coinfection of insects with wild-type and recombinant baculoviruses in which the polyhe- drin gene promoter is used to express hepatitis B virus envelope protein (hepatitis B virus surface antigen; HBsAg) or influenza A virus neuraminidase (NA). Viruses were administered per os to larvae of the cabbage looper, Trichoplusia ni, causing an infection that within 5 days resulted in the production of 0O.15 mg of HBsAg per insect, representing 1.5% of the total extracted protein, or "'2.8 mg of NA per insect, representing 28% of the total extractable protein. The HBsAg and NA produced by infected larvae were purified from insect lysates. These proteins were antigenic as determined by conformation- dependent immunoassays. The NA was enzymatically active with conventional substrates. The method of infection de- scribed allows genetic complementation by wild-type virus of recombinant viruses lacking the polyhedrin gene essential for infection per os and has implications for the high-yield pro- duction in insect larvae of other recombinant proteins of baculoviruses. Infection of insect cells in vitro or insect larvae in vivo with baculovirus results in the production by virus of polyhedra, polymers of the 33-kDa polyhedrin protein (1, 2). The polyhedrin protein is overexpressed in vivo and in cultured cells can eventually account for up to 75% of the total cellular protein. The polyhedra enclose the newly synthesized virus particles and facilitate in vivo horizontal transmission of the virus, primarily by preventing virus inactivation in the environment and in the insect gut after ingestion. After the infection is initiated by entry of the virus into epithelial cells of the midgut region, the polyhedra are no longer necessary to perpetuate the infection (3). Because the polyhedrin protein is not required for in vitro infection (4), a foreign gene can be substituted for the structural gene for polyhedrin without affecting viral replication and thus can then utilize