Japanese Encephalitis Virus Challenge Research Articles

The protective role of Mertk in JEV-induced encephalitis by maintaining the integrity of blood–brain barrier

Japanese encephalitis is an acute infectious disease of the central nervous system caused by neurotropic Japanese encephalitis virus (JEV). As a member of TAM (Tyro3, Axl and Mertk) family, Mertk has involved in multiple biological processes by engaging with its bridging ligands Gas6 and Protein S, including invasion of pathogens, phagocytosis of apoptotic cells, inflammatory response regulation, and the maintenance of blood brain barrier (BBB) integrity. However, its role in encephalitis caused by JEV infection has not been studied in detail. Here, we found that Mertk−/− mice exhibited higher mortality and more rapid disease progression than wild-type mice after JEV challenge. There were no significant differences in viral load and cytokines expression level in peripheral tissues between Wild type and Mertk−/− mice. Furthermore, the absence of Mertk had little effect on the inflammatory response and immunopathological damage while it can cause an increased viral load in the brain. For the in vitro model of BBB, Mertk was shown to maintain the integrity of the BBB. In vivo, Mertk−/− mice exhibited higher BBB permeability and lower BBB integrity. Taken together, our findings demonstrate that Mertk acts as a protective factor in the development of encephalitis induced by JEV infection, which is mainly associated with its beneficial effect on BBB integrity, rather than its regulation of inflammatory response.

Shift in dominant genotypes of Japanese encephalitis virus and its impact on current vaccination strategies.

Japanese encephalitis (JE) is a zoonotic ailment from the Japanese encephalitis virus (JEV). JEV belongs to the flavivirus genus and is categorized into a solitary serotype consisting of five genetically diverse genotypes (I, II, III, IV, and V). The JEV genotype III (GIII) was the prevailing strain responsible for multiple outbreaks in countries endemic to JEV until 1990. In recent years, significant improvements have occurred in the epidemiology of JE, encompassing the geographical expansion of the epidemic zone and the displacement of prevailing genotypes. The dominant genotype of the JEV has undergone a progressive shift from GIII to GI due to variations in its adaptability within avian populations. From 2021 to 2022, Australia encountered an epidemic of viral encephalitis resulting from infection with the GIV JEV pathogen. The current human viral encephalitis caused by GIV JEV is the initial outbreak since its initial discovery in Indonesia during the late 1970s. Furthermore, following a time frame of 50 years, the detection and isolation of GV JEV have been reported in Culex mosquitoes across China and South Korea. Evidence suggests that the prevalence of GIV and GV JEV epidemic regions may be on the rise, posing a significant threat to public safety and the sustainable growth of animal husbandry. The global approach to preventing and managing JE predominantly revolves around utilizing the GIII strain vaccine for vaccination purposes. Nevertheless, research has demonstrated that the antibodies generated by the GIII strain vaccine exhibit limited capacity to neutralize the GI and GV strains. Consequently, these antibodies cannot protect against JEV challenge caused by animal GI and GV strains. The limited cross-protective and neutralizing effects observed between various genotypes may be attributed to the low homology of the E protein with other genotypes. In addition, due to the GIV JEV outbreak in Australia, further experiments are needed to evaluate the protective efficiency of the current GIII based JE vaccine against GIV JEV. The alteration of the prevailing genotype of JEV and the subsequent enlargement of the geographical extent of the epidemic have presented novel obstacles in JE prevention and control. This paper examines the emerging features of the JE epidemic in recent years and the associated problems concerning prevention and control.

Nanoparticle vaccine based on the envelope protein domain III of Japanese encephalitis virus elicitsrobust protective immune responses in mice.

Aim: To develop a vaccine candidate for Japanese encephalitis virus (JEV), for which an effective and safe vaccine is urgently needed. Materials & methods: A vaccine candidate based on domain III of the JEV envelope protein and lumazine synthase (EDIII-LS) was prepared by coupling multivalent ED III to a self-assembling nanoparticle of LS through genetic fusion and self-assembly. Results: High enrichment of ED III was achieved based on the self-assembly of an EDIII-LS polymer. EDIII-LS strongly promoted dendritic cells' internalization and presentation compared with ED III monomer. The cellular and humoral immune responses provoked by EDIII-LS were remarkably higher than those caused by ED III in mice, and conferred complete protection against JEV challenge. Conclusion: The study of ED III-based nanoparticles suggests an effective approach against JEV.

Japanese Encephalitis DNA Vaccines with Epitope Modification Reduce the Induction of Cross-Reactive Antibodies against Dengue Virus and Antibody-Dependent Enhancement of Dengue Virus Infection

Infection with viruses belonging to the genus Flavivirus, such as Japanese encephalitis virus (JEV) and dengue virus (DENV), is a worldwide health problem. Vaccines against JEV and DENV are currently available. However, the dengue vaccine possibly increases the risk of severe dengue due to antibody-dependent enhancement (ADE). Moreover, the Japanese encephalitis (JE) vaccine reportedly induces cross-reactive ADE-prone antibodies against DENV, potentially leading to symptomatic dengue. Therefore, it is necessary to eliminate the risk of ADE through vaccination. In this study, we attempted to develop a JE vaccine that does not induce ADE of DENV infection using an epitope modification strategy. We found that an ADE-prone monoclonal antibody cross-reactive to DENV and JEV recognizes the 106th amino acid residue of the E protein of JEV (E-106). The JE DNA vaccine with a mutation at E-106 (E-106 vaccine) induced comparable neutralizing antibody titers against JEV to those induced by the wild-type JE DNA vaccine. Meanwhile, the E-106 vaccine induced 64-fold less cross-reactive ADE-prone antibodies against DENV. The mutation did not compromise the protective efficacy of the vaccine in the lethal JEV challenge experiment. Altogether, the modification of a single amino acid residue identified in this study helped in the development of an ADE-free JE vaccine.

Japanese encephalitis virus induces vasodilation and severe lethality in adult and aged AG129 mice lacking alpha, beta and gamma interferon receptors

Japanese encephalitis virus (JEV) is a single-stranded positive-sense RNA virus belonging to the Flaviviridae family. The JEV is the leading cause of viral encephalitis in children and the elderly which is spread by mosquitoes. JEV infection has been established in different animal models such as mouse, hamster, guinea pig, swine, rat, monkey, rabbit by using the different routes of inoculations. Here, we have shown that the alpha/beta and gamma -receptor deficient AG129 mouse induces fatal encephalitis in both young and aged old mice, when challenged with high titer JEV Indian clinical isolate by both intraperitoneal and intradermal route. The JEV infected AG129 mouse have shown neurological symptoms, JEV-induced pathological features and supported high level viral replication. Additionally, administration of JEV in AG129 mice resulted in the induction of severe peripheral vascular permeability, which is a major hall mark of Dengue infection but not shown in JEV. Taken together, our results demonstrate interferon α/β and γ receptors knock out AG129 mouse does not need adaptation of JEV clinical isolates and could be is a promising JEV challenge mouse model by mimicking the natural intradermal route of administration for rapid screening of novel antivirals and vaccines.

Re-burying Artificially Exposed Surface of Viral Subunit Vaccines Through Oligomerization Enhances Vaccine Efficacy.

Viral subunit vaccines often suffer low efficacy. We recently showed that when taken out of the context of whole virus particles, recombinant subunit vaccines contain artificially exposed surface regions that are non-neutralizing and reduce their efficacy, and thus these regions need to be re-buried in vaccine design. Here we used the envelope protein domain III (EDIII) of Japanese encephalitis virus (JEV), a subunit vaccine candidate, to further validate this important concept for subunit vaccine designs. We constructed monomeric EDIII, dimeric EDIII via a linear space, dimeric EDIII via an Fc tag, and trimeric EDIII via a foldon tag. Compared to monomeric EDIII or linearly linked dimeric EDIII, tightly packed EDIII oligomers via the Fc or foldon tag induce higher neutralizing antibody titers in mice and also protect mice more effectively from lethal JEV challenge. Structural analyses demonstrate that part of the artificially exposed surface areas on recombinant EDIII becomes re-buried in Fc or foldon-mediated oligomers. This study further establishes the artificially exposed surfaces as an intrinsic limitation of subunit vaccines, and suggests that re-burying these surfaces through tightly packed oligomerization is a convenient and effective approach to overcome this limitation.

Autophagy inhibitors alleviate Japanese encephalitis virus-induced cerebral inflammation in mice.

Japanese encephalitis (JE) is a zoonotic epidemic disease caused by Japanese encephalitis virus (JEV), and currently, no medicines are available to treat this disease. Autophagy modulators play an important role in the treatment of tumors, heart disease, and some viral diseases. The aim of this study was to investigate the effects of autophagy modulators on JEV infection and the host response in mice. The experimental mice were grouped as follows: DMEM (control), JEV, JEV+rapamycin (JEV+Rapa), JEV+wortmannin (JEV+Wort), JEV+chloroquine (JEV+CQ), Rapa, Wort, and CQ. The control group was treated with DMEM. The mice in other groups were infected with 105 PFU of JEV, and Rapa, Wort, and CQ were administered 2 h prior to JEV challenge and then administered daily for 10 consecutive days. All mice were monitored for neurological signs and survival. The damage of subcellular structures in the mouse brain was evaluated by transmission electron microscopy. The distribution of virus in the mouse brain was determined by RNAScope staining and immunohistochemical staining. The neuroinflammatory responses in the brain were examined via quantitative real-time PCR, and the signal pathways involved in neuroinflammation were identified by Western blot. The mice in the JEV+Wort and JEV+CQ groups showed milder neurological symptoms, less damage to the mitochondria in the brain tissue, and a higher survival rate than those in the JEV+Rapa and JEV groups. Compared with the JEV+Rapa and JEV groups, the distribution of JEV in the brain of mice in the JEV+Wort and JEV+CQ groups was lower, and the inflammatory response was weaker. No significant difference was observed in the expression of the PI3K/AKT/NF-κB pathway in mouse brain among the different groups. Our study suggests that the autophagy inhibitors Wort and CQ reduce JEV infection and weaken the inflammatory response, which does not depend on the PI3K/AKT/NF-κB pathway in mouse brain.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00705-021-05283-9.

A VLP-Based Vaccine Candidate Protects Mice against Japanese Encephalitis Virus Infection.

Japanese encephalitis virus (JEV) is the leading cause of epidemic encephalitis in Asia, and vaccination is the most effective way to prevent JE. Although several licensed vaccines were widely used, there is still a demand for developing safer, cheaper, and more effective JE vaccines. In the current study, a virus-like particle (VLP) vaccine candidate containing the envelope structural protein of JEV expressed by the Pichia pastoris was assembled in vitro. It elicited a robust humoral and cellular immune response in mice model, conferring immunodeficient mice complete protection against lethal doses of JEV challenge. Furthermore, pigs immunized with VLP alone without adjuvant via intramuscular produced high neutralizing antibodies against JEV. Consequently, this study showed a new design of JEV subunit vaccine based on VLP strategy and demonstrated the potential for clinical application.

A Novel Recombinant Virus-Like Particles Displaying B and T Cell Epitopes of Japanese Encephalitis Virus Offers Protective Immunity in Mice and Guinea Pigs.

Virus-like particles (VLPs) are non-replicative vectors for the delivery of heterologous epitopes and are considered one of the most potent inducers of cellular and humoral immune responses in mice and guinea pigs. In the present study, VLP-JEVe was constructed by the insertion of six Japanese encephalitis virus (JEV) envelope protein epitopes into different surface loop regions of PPV VP2 by the substitution of specific amino acid sequences without altering the assembly of the virus; subsequently, the protective efficacy of this VLP-JEVe was evaluated against JEV challenge in mice and guinea pigs. Mice immunized with the VLP-JEVe antigen developed high titers of neutralizing antibodies and 100% protection against lethal JEV challenge. The neutralizing and hemagglutination inhibition (HI) antibody responses were also induced in guinea pigs vaccinated with VLP-JEVe. In addition, immunization with VLP-JEVe in mice induced effective neutralizing antibodies and protective immunity against PPV (porcine parvovirus) challenge in guinea pigs. These studies suggest that VLP-JEVe produced as described here could be a potential candidate for vaccine development.

Novel Japanese encephalitis virus NS1-based vaccine: Truncated NS1 fused with E. coli heat labile enterotoxin B subunit.

BackgroundCurrent vaccines against Japanese encephalitis virus (JEV) of flaviviruses have some disadvantages, such as the risk of virulent reversion. Non-structural protein NS1 is conserved among flaviviruses and confers immune protection without the risk of antibody-dependent enhancement (ADE). Therefore, NS1 has become a promising vaccine candidate against flaviviruses.MethodsA NS1-based vaccine (LTB-NS1∆63) with a truncated NS1 protein (NS1∆63) fused to E. coli heat-labile enterotoxin B subunit (LTB) was expressed in E.coli and explored for its ability to induce immune responses. Safety of LTB-NS1∆63 was assessed by determining its toxicity in vitro and in vivo. Protective capability of LTB-NS1∆63 and its-induced antisera was evaluated in the mice challenged with JEV by analyzing mortality and morbidity.FindingsLTB-NS1∆63 induced immune responses to a similar level as LTB-NS1, but more robust than NS1∆63 alone, particularly in the context of oral immunization of mice. Oral vaccination of LTB-NS1∆63 led to a higher survival rate than that of NS1∆63 or live-attenuated JEV vaccine SA14–14–2 in the mice receiving lethal JEV challenge. LTB-NS1∆63 protein also significantly decreases the morbidity of JEV-infected mice. In addition, passive transfer of LTB-NS1∆63-induced antisera provides a protection against JEV infection in mice.InterpretationNS1∆63 bears JEV NS1 antigenicity. Besides, LTB-NS1∆63 could serve as a novel protein-based mucosa vaccine targeting JEV and other flaviviruses.FundingThis work was supported by the National Natural Science Foundation, Jiangxi Province Science and Technology Committee, Education Department of Jiangxi Province.

Identification of a protective epitope in Japanese encephalitis virus NS1 protein

Japanese encephalitis virus (JEV) is one of the most important culex transmitted-flaviviruses, which can cause encephalitis in humans. Although non-structural protein 1 (NS1) of JEV does not stimulate neutralizing antibodies, this protein can provide high immunoprotection in vivo. The protective epitopes and the protective mechanism of NS1 still remain unclear. In this study, we generated five different monoclonal antibodies (mAbs) targeting the NS1 protein of JEV. In vitro experiments revealed that none of these five antibodies neutralized the JEV infection. In mouse protection studies, one of these mAbs, designated 2B8, provided a therapeutic effect against JEV lethal challenge (70% survival rate). Using peptide mapping analysis, we found that mAb 2B8 reacted with the epitope 225PETHTLWGD233 in the NS1 protein, in which any mutations among amino acid residues T228, H229, L231 or W232 could cause binding failure of 2B8 to the NS1 protein. Furthermore, mice immunized with KLH-polypeptide (225PETHTLWGD233) showed reduced mortality following JEV challenge. Collectively, we found a new protective epitope in the JEV NS1 protein. These results may facilitate the development of therapeutic agent and subunit-based vaccines based on the NS1 protein.

A replication-defective Japanese encephalitis virus (JEV) vaccine candidate with NS1 deletion confers dual protection against JEV and West Nile virus in mice

In our previous study, we have demonstrated in the context of WNV-ΔNS1 vaccine (a replication-defective West Nile virus (WNV) lacking NS1) that the NS1 trans-complementation system may offer a promising platform for the development of safe and efficient flavivirus vaccines only requiring one dose. Here, we produced high titer (107 IU/ml) replication-defective Japanese encephalitis virus (JEV) with NS1 deletion (JEV-ΔNS1) in the BHK-21 cell line stably expressing NS1 (BHKNS1) using the same strategy. JEV-ΔNS1 appeared safe with a remarkable genetic stability and high degrees of attenuation of in vivo neuroinvasiveness and neurovirulence. Meanwhile, it was demonstrated to be highly immunogenic in mice after a single dose, providing similar degrees of protection to SA14-14-2 vaccine (a most widely used live attenuated JEV vaccine), with healthy condition, undetectable viremia and gradually rising body weight. Importantly, we also found JEV-ΔNS1 induced robust cross-protective immune responses against the challenge of heterologous West Nile virus (WNV), another important member in the same JEV serocomplex, accounting for up to 80% survival rate following a single dose of immunization relative to mock-vaccinated mice. These results not only support the identification of the NS1-deleted flavivirus vaccines with a satisfied balance between safety and efficacy, but also demonstrate the potential of the JEV-ΔNS1 as an alternative vaccine candidate against both JEV and WNV challenge.

Immune responses and protective effects against Japanese encephalitis induced by a DNA vaccine encoding the prM/E proteins of the attenuated SA14-14-2 strain

Japanese encephalitis virus (JEV) is the causal pathogen of Japanese encephalitis (JE), which has become a severe public health problem and is one of the most rapidly spreading mosquito-borne diseases worldwide. Currently, there is no specific treatment for JEV. A vaccine would be an effective measure for reducing morbidity and mortality. Although the live attenuated vaccine SA14-14-2 has been approved in some countries, it is still necessary to develop safer, more effective, and less costly vaccines. In this study, a DNA vaccine candidate, pV-SA14ME, expressing the prM/E proteins of SA14-14-2 was inoculated into BALB/c mice via intramuscular electroporation, and the immunogenicity and degree of protection were evaluated. We found that administration of 50 μg pV-SA14ME via electroporation via three immunizations could induce persistent humoral and cellular immune responses and effectively protect mice against lethal JEV challenge. This study provides a basis for the subsequent promotion and use of the vaccine and lays the foundation for its further use in swine and humans.

Sub-lethal Dose of Atropine Gives Protection from Japanese Encephalitis Virus Infection in Chick Embryo Model

Japanese encephalitis (JE) is now a public health concern due to limitation of the available vaccine and nonexistence of specific antiviral therapy and thus demands new potential therapeutical strategies. The present study was conducted to evaluate the effect of atropine in controlling JE virus infection in chick embryo, which is already established as an ideal model for study of various host–pathogen interactions and screening of newer antiviral agents. In this experiment we pre-treated 12 day old embryos with atropine sulphate to see whether it can give protection from JE virus challenge. There were significant decrease in the viral loads in the brain, chorioallantoic membrane and amniotic fluid of the embryo in atropine sulphate pre-treated groups compared to the infection groups. Atropine sulphate pre-treated group showed significant up-regulation of interferon alpha and toll like receptor-3 mRNA, than other analogous groups. The histological changes showed significant reduction of necrosis, inflammation and other co-morbid pathology in the atropine sulphate pre-treated groups. Hence this study demonstrated the protective role of atropine in controlling the JE virus replication and its organ-tropic dissemination to brain which may pave way for a prospective therapy in the future treatment of JE.

Enhanced Immune Responses Against Japanese Encephalitis Virus Infection Using Japanese Encephalitis Live-Attenuated Virus Adjuvanted with Montanide GEL 01 ST in Mice.

Japanese encephalitis virus (JEV) is one of the major causes of acute encephalitis in human and animal. To prevent JEV infection, an effective live-attenuated vaccine is needed. In the article, JEV attenuated strain, SCYA201201 of GI genotype, which was mixed with 10% concentrate GEL 01 ST adjuvant (Montanide™ GEL 01 ST), was selected for a vaccine candidate and its immunogenicity was evaluated in mice. Our results showed that JEV mixed with GEL 01 ST elicited production of both IgG1 and IgG2a antibodies, and enhanced virus-specific crossprotective intergenotypic response in mice. Proliferation of splenocytes was observed in all immunized groups and a relatively higher proliferation activity was detected in JEV mixed with GEL 01 ST group (p < 0.05). In the JEV + 10% GEL 01 ST vaccinated group, the proportion of CD4+ T cells in spleen was significantly higher than that of control group (p < 0.05), and the yields of interleukin (IL)-2, IL-4, and interferon-γ in the splenocyte supernatant were also significantly higher than that of control group (p < 0.05). Moreover, complete protection was provided after JEV challenge in mice in JEV mixed with GEL 01 ST group, and early immunity was detected in those mice immunized with JEV mixed with GEL 01 ST. These findings confirm that GEL 01 ST can enhance JEV live-attenuated immunogenicity, and JEV +10% GEL 01 ST used as vaccine candidates provide protection against JEV infection in a mouse model, which could be used as potential vaccine candidates in pig.

A Japanese Encephalitis Virus Vaccine Inducing Antibodies Strongly Enhancing In Vitro Infection Is Protective in Pigs.

The Japanese encephalitis virus (JEV) is responsible for zoonotic severe viral encephalitis transmitted by Culex mosquitoes. Although birds are reservoirs, pigs play a role as amplifying hosts, and are affected in particular through reproductive failure. Here, we show that a lentiviral JEV vector, expressing JEV prM and E proteins (TRIP/JEV.prME), but not JEV infection induces strong antibody-dependent enhancement (ADE) activities for infection of macrophages. Such antibodies strongly promoted infection via Fc receptors. ADE was found at both neutralizing and non-neutralizing serum dilutions. Nevertheless, in vivo JEV challenge of pigs demonstrated comparable protection induced by the TRIP/JEV.prME vaccine or heterologous JEV infection. Thus, either ADE antibodies cause no harm in the presence of neutralizing antibodies or may even have protective effects in vivo in pigs. Additionally, we found that both pre-infected and vaccinated pigs were not fully protected as low levels of viral RNA were found in lymphoid and nervous system tissue in some animals. Strikingly, the virus from the pre-infection persisted in the tonsils throughout the experiment. Finally, despite the vaccination challenge, viral RNA was detected in the oronasal swabs in all vaccinated pigs. These latter data are relevant when JEV vaccination is employed in pigs.

Enhanced immune responses against Japanese encephalitis virus using recombinant adenoviruses coexpressing Japanese encephalitis virus envelope and porcine interleukin-6 proteins in mice

Japanese encephalitis is a reproductive disorder caused by Japanese encephalitis virus (JEV) in swine. Previous studies have demonstrated that recombinant adenovirus serotype 5 (Ad5) may be a potential vaccine candidate because it can express JEV envelope epitopes and induce immune responses against JEV. Still, it will be necessary to develop an adjuvant that can enhance both humoral and cellular immune responses to the recombinant antigen delivered by non-replicating Ad5. In this study, we investigated the systemic immune responses of BALB/c mice immunized with recombinant adenovirus expressing JEV envelope epitopes in combination with porcine interleukin-6 (rAdE-IL-6).The rAdE-IL-6 immunized group had the highest titers of anti-JEV antibody as detected by an enzyme-linked immunosorbent assay (ELISA), as well as the highest levels of neutralizing antibody (1:75) as detected by a serum neutralization test. Similarly, higher concentrations of interferon-gamma (834.7pg/ml) and interleukin-6 (IL-6) (229.7pg/ml) were detected in the rAdE-IL-6 group using an ELISA assay. These data indicate that immunized BALB/c induce a strong cellular response against rAdE-IL-6. Furthermore, after challenge with the virulent JEV SCYA201201 strain, the rAdE-IL-6 group generated an immune protective response 70% greater than that of the control group, indicating that rAdE-IL-6 induced a protective immune response against JEV challenge in mice. The results from this study demonstrated that IL-6 is a strong adjuvant that can enhance both humoral and cellular immune responses in mice. Furthermore, a recombinant adenovirus coexpressing JEV envelope epitopes and porcine IL-6 protein may be an effective vaccine in animals.

Generation and characterization of a new mammalian cell line continuously expressing virus-like particles of Japanese encephalitis virus for a subunit vaccine candidate.

BackgroundJapanese encephalitis virus (JEV) is the most important cause of epidemic encephalitis in most Asian regions. There is no specific treatment available for Japanese encephalitis, and vaccination is the only effective way to prevent JEV infection in humans and domestic animals. The purpose of this study is to establish a new mammalian cell line stably and efficiently expressing virus-like particle of JEV for potential use of JEV subunit vaccine.ResultsWe generated a new cell clone (BJ-ME cells) that stably produces a secreted form of Japanese encephalitis virus (JEV) virus-like particle (VLP). The BJ-ME cells were engineered by transfecting BHK-21 cells with a code-optimized cDNA encoding JEV prM and E protein expression plasmid. Cell line BJ-ME can stably produces a secreted form of Japanese encephalitis virus virus-like particle (JEV-VLP) which contains the JEV envelope glycoprotein (E) and membrane protein (M). The amount of JEV-VLP antigen released into the culture fluid of BJ-ME cells was as high as 15–20 μg/ml. JEV-VLP production was stable after multiple cell passages and 100% cell expression was maintained without detectable cell fusion or apoptosis. Cell culture fluid containing the JEV-VLP antigen could be harvested five to seven times continuously at intervals of 4–6 days while maintaining the culture. Mice immunized with the JEV-VLP antigen with or without adjuvant developed high titers of neutralizing antibodies and 100% protection against lethal JEV challenge.ConclusionThese results suggest that the recombinant JEV-VLP antigen produced by the BJ-ME cell line is an effective, safe and affordable subunit Japanese encephalitis vaccine candidate, especially for domestic animals such as pig and horse.

Inactivated genotype 1 Japanese encephalitis vaccine for swine.

PurposeJapanese encephalitis is a reproductive disorder caused by Japanese encephalitis virus (JEV) in swine. Recent genotype (G) shift phenomenon (G3 to G1) in the Asia-wide has posed a challenge for proper prevention by the current vaccine strain. Thus, new kinds of JEV G1 vaccines with enhanced immunogenicity have been required for pigs.Materials and MethodsRecombinant porcine granulocyte monocyte-colony stimulating factor (reporGM-CSF) protein was expressed in Spodoptera frugiperda (Sf-9) cells using baculovirus expression system. Two kinds of trials with inactivated JEV vaccines containing IMS1313 adjuvant (Seppic, France) were prepared with or without reporGM-CSF protein. Safety and immunogenicity of the pigs inoculated with the JEV vaccines via intramuscular route was evaluated for 28 days after inoculation.ResultsMice, guinea pigs, and fattening pigs inoculated with the inactivated vaccine showed no signs for 14 and 21 days. Both hemagglutination inhibition and plaque reduction neutralizing antibody titers were significantly higher in pigs immunized with the vaccine containing reporGM-CSF protein after boosting. However, on the side of vaccine efficacy, most mice (87%) immunized with the inactivated JEV vaccine survived after virulent JEV challenge. Whereas the group with the vaccine containing reporGM-CSF protein showed lower protective effects than the vaccine alone for the biological activity of the GM-CSF depending on species specific.ConclusionOur data indicate that animals inoculated with the JEV vaccines was safe and pigs inoculated with inactivated JEV vaccine containing reporGM-CSF protein showed higher humoral immune responses than that of inactivated JEV vaccine without reporGM-CSF protein.

Antibodies generated by immunization with the NS1 protein of West Nile virus confer partial protection against lethal Japanese encephalitis virus challenge

Japanese encephalitis virus (JEV) and West Nile virus (WNV) are two medically important flaviviruses that can cause severe hemorrhagic and encephalitic diseases in humans. Immune responses directed against the NS1 protein of flaviviruses can confer protection against lethal viral challenge. Previous studies have shown that the WNV NS1 protein harbors epitopes that elicit antibodies that cross react with JEV. Here we demonstrate that the WNV NS1 protein not only contains cross-reactive epitopes, but that the antibodies elicited by these cross-reactive epitopes provide partial protection against lethal JEV challenge in a mouse model. Mice immunized with WNV NS1 protein showed reduced morbidity and mortality following both intracerebral and intraperitoneal JEV challenge. WNV NS1 immunization attenuated the extent of lung pathology generated following JEV challenge, and delayed the appearance of other pathological findings including vascular cuffing. By screening and identifying the specific WNV NS1 protein-derived peptides recognized by serum antibodies elicited by immunization with WNV NS1 protein and by JEV challenge, we found after JEV challenge will induce several new epitopes, but which epitope primarily contribute to antibody-mediated cross protection need further evaluation. The knowledge and reagents generated in this study have potential applications in vaccine and subunit vaccine development for WNV and JEV.