Development of a Japanese encephalitis virus genotype V virus-like particle vaccine in silkworms

Abstract

To counter the spread of multiple Japanese encephalitis virus (JEV) variants harboured in alternative host species and highly neurotoxic variants with new antigenicity, such as genotype V (Muar), methods for developing more effective and low-cost vaccines against a variety of epidemic JEV strains are required. Here, we successfully synthesized large amounts of a Muar virus-like particle (MVLP) vaccine for JEV in silkworm pupae by using a Bombyx mori nuclear polyhedrosis virus recombinant consisting of JEV codon-optimized envelope (E) DNA. In particular, histopathological examination suggested that MVLP was efficiently synthesized in body fat tissues as well as epithelial cells. Quantitative analysis indicated that one silkworm pupa produced 724.8 µg of E protein in the MVLP vaccine. Electron microscopic examination of purified MVLP vaccine defined a typical MVLP morphological structure. Detailed MVLP antigen assessment by immune-electron microscopy revealed that the majority of MVLPs were covered with approximately 10 nm projections. Boosted immunization with MVLP antigens in mice and rabbits tended to show improved plaque inhibition potency against homologous Muar and heterologous Nakayama, but less potency to Beijing-1 strains. Notably, mixed immune rabbit antisera against Nakayama and Muar VLP antigens led to an increase in the low antibody reaction to Beijing-1. Additionally, a stopgap divalent JEV vaccine consisting of MVLP and Nakayama VLP and its immune mouse serum significantly increased plaque inhibition titre against Muar, Nakayama and Beijing-1 strains. These findings suggested that low-cost MVLP vaccines prepared in silkworm pupae are suitable for providing simultaneous protection of individuals in developing countries against various JEV strains.

Mosquito Cell-Derived Japanese Encephalitis Virus-Like Particles Induce Specific Humoral and Cellular Immune Responses in Mice

“…Mosquito cell-derived JEV VLPs presented a density range of~35-45% sucrose ( Figure 4A). TEM and DLS analysis revealed that the average diameter of the JEV VLPs was roughly 30 nm ( Figure 4C), which is similar to the size of Zika [56] and JE GV VLPs [23,29] produced using a conventional baculovirus system or stable cell lines.…”

“…Nonetheless, JEV GI VLPs produced from stable mammalian cell lines have been shown to induce immunity against GI and GIII JEV in mice and swine [27]. The conventional baculovirus expression system has been employed to produce large quantities of JEV VLPs in insect cells [28,29]; however, the antigenicity of Lepidoptera-derived JEV VLPs has yet to be elucidated. In both mosquitos and vertebrates, arboviruses confer virion-specific properties to the host [30].…”

Solutions against emerging infectious and noninfectious human diseases through the application of baculovirus technologies

“…mosquitoes. The infection is spread mainly in Asian countries causing around 50,000 cases with some 15,000 deaths per year (Nerome et al 2018). Even though a vaccine against JEV has been approved for human use (IXIARO, Valneva), BEVS has been explored as an alternative platform to achieve more costeffective prototypes.…”

“…In addition, stably transformed insect cells were developed for the constitutive production of JEV VLPs (Yamaji and Konishi 2013). As a low-cost alternative, large amounts of VLPs were furthermore produced in silkworm pupae infected with rec-baculoviruses that subsequently induced protection in mice (Nerome et al 2018).…”

Recent Advances in the Development of Virus-Like Particle-Based Flavivirus Vaccines

“…Nerome et al developed a Nakayama JE VLP in silkworms using codon-optimized prM and E genes, and the VLPs showed protection against both homologous and heterologous strains (Beijing-1 and Muar) [110]. The same group successfully generated a Muar JE VLP vaccine in silkworm pupae through codon optimization of E gene [111]. A genotype I JE VLP, consisting of prM and E proteins, was used to establish a mouse model exhibiting neurological symptoms, and it showed potent protection against homologous or heterologous JEVs [112].…”