Zika virus baculovirus-expressed envelope protein elicited humoral and cellular immunity in immunocompetent mice

Zika virus (ZIKV) is a mosquito-borne virus that has a high risk of inducing Guillain–Barré syndrome and microcephaly in newborns. Because vaccination is considered the most effective strategy against ZIKV infection, we designed a recombinant vaccine utilizing the baculovirus expression system with two strains of ZIKV envelope protein (MR766, Env_M; ZBRX6, Env_Z). Animals inoculated with Env_M and Env_Z produced ZIKV-specific antibodies and secreted effector cytokines such as interferon-γ, tumor necrosis factor-α, and interleukin-12. Moreover, the progeny of immunized females had detectable maternal antibodies that protected them against two ZIKV strains (MR766 and PRVABC59) and a Dengue virus strain. We propose that the baculovirus expression system ZIKV envelope protein recombinant provides a safe and effective vaccine strategy.

RG203KR mutations in SARS-CoV-2 Nucleocapsid: Assessing the impact using Virus-like particle model system

The emergence and evolution of SARS-CoV-2 is characterized by the occurrence of diverse sets of mutations that affect virus characteristics, including transmissibility and antigenicity. Recent studies have focused mostly on Spike protein mutations; however, SARS-CoV-2 variants of interest (VoI) or concern (VoC) contain significant mutations in the nucleocapsid protein as well. To study the relevance of the mutations at the virion level, recombinant baculovirus expression system based VLPs were generated for the prototype Wuhan sequence along with Spike mutants like D614G, G1124V and the significant RG203KR mutation in Nucleocapsid. All the four structural proteins assembled in a particle wherein the morphology and size of the particle confirmed by TEM closely resembles the native virion. The VLP harbouring RG203KR mutations in nucleocapsid exhibited augmentation of humoral immune responses and enhanced neutralization by the immunized mice sera. Results demonstrate a non-infectious platform to quickly assess the implication of mutations in structural proteins of the emerging variant.

Unique features of a recombinant haemagglutinin influenza vaccine that influence vaccine performance

The influenza vaccine field has been constantly evolving to improve the speed, scalability, and flexibility of manufacturing, and to improve the breadth and longevity of the protective immune response across age groups, giving rise to an array of next generation vaccines in development. Among these, the recombinant influenza vaccine tetravalent (RIV4), using a baculovirus expression vector system to express recombinant haemagglutinin (rHA) in insect cells, is the only one to have reached the market and has been studied extensively. We describe how the unique structural features of rHA in RIV4 improve protective immune responses compared to conventional influenza vaccines made from propagated influenza virus. In addition to the sequence integrity, characteristic of recombinant proteins, unique post-translational processing of the rHA in insect cells instills favourable tertiary and quaternary structural features. The absence of protease-driven cleavage and addition of simple N-linked glycans help to preserve and expose certain conserved epitopes on HA molecules, which are likely responsible for the high levels of broadly cross-reactive and protective antibodies with rare specificities observed with RIV4. Furthermore, the presence of uniform compact HA oligomers and absence of egg proteins, viral RNA or process impurities, typically found in conventional vaccines, are expected to eliminate potential adverse reactions to these components in susceptible individuals with the use of RIV4. These distinct structural features and purity of the recombinant HA vaccine thus provide a number of benefits in vaccine performance which can be extended to other viral targets, such as for COVID-19.

The potential application of Trichoplusia ni granulovirus En3 enhancin as a synergist in baculovirus-based insecticides

The increased costs associated with baculovirus mass-production urge the search for synergistic products that reduce the amount of active matter. In the present thesis, a synergistic factor with great potential for baculovirus-based formulations was expressed and produced using a baculovirus expression system. The main achievement of the present thesis is that the in vivo production of solubilized enhancins using baculovirus-based expression systems can be used to improve the efficacy of biological insecticides against lepidopteran pests, reducing the active matter of bioinsecticides and making them commercially competitive with chemicals.

Pigs Immunized with the Virus-like Particle Vaccine Are Protected against the Hepatitis E-3 Virus

In this study, we generated the HEV virus-like particle (VLP) vaccine expressing 239 amino acids (367–605 aa) of the HEV-3 ORF2 using the baculovirus expression system. The HEV-3-239-VLP vaccine efficacy was evaluated by dividing 12 pathogen-free pigs into four groups: negative control, positive control, 100 μg VLP-, and 200 μg VLP-vaccinated groups for 10 weeks. The pigs in either of the vaccinated groups were administered the corresponding first and booster doses on weeks 0 and 2. At week 4, the positive control and two vaccinated groups were challenged with 106 HEV-3 genomic equivalent copies; viremia and fecal shedding of the virus were identified in pigs in the positive control and 100 μg VLP-vaccinated pigs showed transient viremia and fecal viral shedding. However, no viruses were detected in the serum or fecal samples of the 200 μg VLP-vaccinated pigs. The 100 and 200 μg VLP-vaccinated pigs had significantly higher (p < 0.01) anti-HEV antibodies than the negative control pigs from weeks 6–10 with normal levels of liver enzymes. The 200 μg VLP-vaccinated pigs showed statistically less liver tissue fibrosis (p < 0.05) than that of the positive control pigs. Thus, the novel baculovirus expression system-generated VLP vaccine dose-dependently protects against HEV-3 challenge and may be useful in other animal species, including humans.

Studies of the Parasite-Midgut Interaction Reveal Plasmodium Proteins Important for Malaria Transmission to Mosquitoes

Malaria transmission relies on parasite-mosquito midgut interaction. The interactive proteins are hypothesized to be ideal targets to block malaria transmission to mosquitoes. We chose 76 genes that contain signal peptide-coding regions and are upregulated and highly abundant at sexual stages. Forty-six of these candidate genes (60%) were cloned and expressed using the baculovirus expression system in insect cells. Six of them, e.g., PF3D7_0303900, PF3D7_0406200 (Pfs16), PF3D7_1204400 (Pfs37), PF3D7_1214800, PF3D7_1239400, and PF3D7_1472800 were discovered to interact with blood-fed mosquito midgut lysate. Previous works showed that among these interactive proteins, knockout the orthologs of Pfs37 or Pfs16 in P. berghei reduced oocysts in mosquitoes. Here we further found that anti-Pfs16 polyclonal antibody significantly inhibited P. falciparum transmission to Anopheles gambiae. Investigating these candidate proteins will improve our understanding of malaria transmission and discover new targets to break malaria transmission.

Active Human and Murine Tumor Necrosis Factor α Cytokines Produced from Silkworm Baculovirus Expression System

The tumor necrosis factor α (TNFα) has been employed as a promising reagent in treating autoimmunity and cancer diseases. To meet the substantial requirement of TNFα proteins, we report in this study that mature types of recombinant human and murine TNFα proteins are successfully expressed in the baculovirus expression system using silkworm larvae as hosts. The biological activities of purified products were verified in culture murine L929 cells, showing better performance over a commercial Escherichia coli-derived murine TNFα. By comparing the activity of purified TNFα with or without the tag removal, it is also concluded that the overall activity of purified TNFα cytokines could be further improved by the complete removal of C-terminal fusion tags. Collectively, our current attempt demonstrates an alternative platform for supplying high-quality TNFα products with excellent activities for further pharmaceutical and clinical trials.