Overexpression of a virus-like particle influenza vaccine in Eri silkworm pupae, using Autographa californica nuclear polyhedrosis virus and host-range expansion

Abstract

Ecological investigations of silkworms have revealed that Eri silkworms (Samia cynthia ricini) possess useful morphological and ecological characteristics for virus-like particle (VLP) production, namely non-seasonal breeding, longer lengths, and heavier weights than Bombyx mori silkworms. Furthermore, when vector DNA from Bombyx mori nuclear polyhedrosis virus (BmNPV), which is unable to replicate in Sf9 cells from Eri silkworms, was replaced with the Autographa californica nuclear polyhedrosis virus (AcNPV) vector, three improved AcNPV influenza virus recombinants capable of replication in Sf9 cells were obtained. Although VLP antigens produced previously in silkworms were not evaluated individually, the present recombinant Fukushima (FkH5) and Anhui (AnH7) VLP antigens were detected in tissue fluids and fat bodies of Eri silkworms. Here, we aimed to determine the function of the AcNPV vector and P143 gene by expressing recombinants in Sf9 cells and eri silkworm pupae. The FkH5 recombinant produced high yields of haemagglutinin (HA)-positive VLPs, showing a mean HA titre of 1.2 million. Similarly, high production of H7 HA VLPs was observed in the fat bodies of eri silkworm pupae. Antigenic analysis and electron microscopy examination of Eri-silkworm-produced H5 HA VLPs showed characteristic antigenicity and morphology similar to those of the influenza virus. Although FkH5 recombinants possessing the AcNPV vector did not replicate in Bm-N cells, the introduction of the helicase p143 gene from BmNPV resulted in their production in Bm-N and Sf9 cells.

Interleukin 12-containing influenza virus-like-particle vaccine elevate its protective activity against heterotypic influenza virus infection

“…We previously established a recombinant AcNPV system in Eri silkworms that produces abundant recombinant proteins in pupae [ 13 ]. In this study, we generated three recombinant AcNPV viruses: FkH5-AcNPV, AnH7-AcNPV, and IL-12-AcNPV.…”

“…The generated plasmids were transformed into DH10Bac cells. Subsequently, recombinant Ac-NPV DNA was purified and transfected into Sf9 cells as described previously [ 13 ]. After 1 week of culture, the supernatant, containing recombinant Ac-NPV viruses, was harvested and used in subsequent experiments.…”

“…B. mori and Eri silkworm pupae were used to produce VLP vaccines by infecting the silkworms with BmNPV and AcNPV recombinant viruses, respectively ( Figure 1 ). Baculovirus inoculation and preparation of VLP vaccines were performed as previously described [ 13 ].…”

“…With an aim to provide inexpensive, effective vaccines against avian influenza, we developed a recombinant baculovirus system for the large-scale production of H5 and H7 HA-containing VLP vaccines in silkworms [ 8 , 12 ]. We also established a similar system in Eri silkworms [ 13 ] that allows the production of HA titres greater than 30 million per pupa. H7-containing VLP vaccines produced using different systems elicited strong immune responses in murine, ferret, and avian models [ 14 , 15 , 16 , 17 ].…”

Virus-Like Particles as an Instrument of Vaccine Production

“…This point will be addressed further in the review, we will just observe here that even complex epitopes represented by trimers may be obtained using heterologous expression systems. This has been demonstrated, for instance, for the trimeres of the human immunodeficiency virus (HIV-1) envelope glycoprotein [18,19] and influenza virus haemagglutinin [20].…”