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Virion and soluble antigens of japanese encephalitis virus.

Japanese encephalitis virions contain a 58 X 10-3-molecular-weight envelope glycoprotein antigen that can be solubilized with sodium lauryl sulfate and separated from other virion structural polypeptides and viral ribonucleic acid by gel filtration chromatography. The 58 X 10-3-molecular-weight envelope protein is the major antigen responsible for cross-reactivity of the virion in complement fixation tests with other closely related arboviruses. A naturally occurring soluble complement-fixing antigen is found in Japanese encephalitis mouse brain preparations after removal of particulate antigens. After partial purification by gel filtration and isoelectric focusing, the 53 X 10-3-molecular weight soluble complement-fixing antigen is more type specific than the Japanese encephalitis envelope antigen in complement fixation tests. Further, the Japanese encephalitis soluble complement-fixing antigen is stable to treatment with sodium lauryl sulfate and 2-mercaptoethanol, whereas virion complement-fixing antigens are unstable after this treatment.

Amino Acids

Intracellular distribution of virus-specific RNA in chick embryo cells infected with Japanese encephalitis virus.

Japanese encephalitis virus (JEV) infected chick embryo (CE) cells were treated with 4 mug actinomycin D/ml and 5 mM-D-glucosamine at 2 or 3 h before harvesting. Production of JEV was not affected by the short-time treatment of these drugs. The radioactivity in virus-specific RNA in the glucosamine-treated cells was a-parently higher than in non-treated cells. Nuclear and cytoplasmic extracts were prepared from the JEV-infected cells pulse-labelled with 3H-uridine at 15 h after infection. Analysis of virus RNA in nuclear extracts on sucrose density gradients showed that most of the radioactivity was in 23S RNA, 26S RNA and 8 to 12S RNA. The radioactivity of virus RNA in cytoplasmic extracts was found in 42S RNA and RNA fragments sedimenting at less than 8S.

Animals

[Chronic infection of VERO cell cultures with Japanese encephalitis virus. 2. Certain characteristics of variability of persistent viruses].

Japanese encephalitis virus contained in the medium of chronically infected VERO-K43 cell cultures was shown to be sensitive to the effect of RNA-ase, resistant to treatment with urea, to have no hemagglutinating activity. After 2 passages in suckling mouse brain the virus lost its sensitivity to RNA-ase, was inactivated by urea, and its hemagglutinating activity was restored. The importance of Japanese encephalitis virus variability in the mechanism of chronic infection of cell cultures is discussed.

Animals

Experimental double infection of Japanese encephalitis virus and herpes simplex virus in mouse brain.

In our laboratory, 64 autopsy cases of Japanese encephalitis patients were examined by immunofluorescence. Three patients showed some evidences of double infection of Japanese encephalitis virus and herpes simplex virus. Experiments were done to see the mechanisms of double infection. In doubly infected mice, Japanese encephalitis virus antigen was localized in the herpes simplex virus infected areas in the brain. These results suggested that "blood brain barrier" was broken by herpes simplex virus infection and Japanese encephalitis virus gained access to the susceptible cells.

Animals

Bergamottin, a bioactive component of bergamot: dual inhibition of Japanese encephalitis virus internalization and genome replication.

Japanese encephalitis virus (JEV) is associated with high mortality and severe neurological sequelae, and existing prevention and control strategies remain insufficient. Therefore, the development of novel antiviral agents is of critical public health importance. This study systematically evaluated the antiviral activity and underlying mechanism of bergamottin, a natural product. Bergamottin exhibited significant dose-dependent inhibitory effects against JEV in multiple cell lines, including BHK-21, HuH-7, and Vero cells, demonstrating potent antiviral efficacy. Mechanistic investigations revealed that bergamottin primarily targeted the internalization and replication stages of the JEV life cycle, thereby effectively suppressing viral proliferation. Additionally, adaptive mutation screening indicated that the D389G mutation in envelope protein E confers drug resistance by potentially changing E protein conformation or reducing endocytic efficiency. In vivo experiment, bergamottin significantly reduced viral loads in mouse brain tissue and effectively improved the survival rate of infected mice. Our findings indicated that bergamottin exerted antiviral activity by dual targeting of key steps in the viral life cycle, making it a highly promising candidate for anti-JEV therapy. Further exploration of the antiviral properties of bergamottin is expected to facilitate its clinical development as a treatment for JEV infection.

Animals

Neutralization tests for dengue and Japanese encephalitis viruses by the focus reduction method using peroxidase-anti-peroxidase staining.

Neutralization tests were made on 4 types of dengue (DEN) virus and Japanese encephalitis (JE) virus by incubation of serially diluted antisera and constant amounts of the viruses and then focus assay of surviving virus infectivity with peroxidase-anti-peroxidase (PAP) staining. Neutralization reactions were virtually completed in 2 hr on incubation of serum-virus mixtures at 28 C. A straight regression line was obtained on a probit chart by plotting the focus reduction rates at various dilutions of a given serum against the logarithm of the serum dilution used in the test. The slopes of the probit regression lines for the neutralization for DEN types 1 and 3 were similar, but differed somewhat from those for DEN type 2 and type 4. The slope of the line for JE virus was quite different from those for DEN viruses. Using these relations, the fifty percent focus reduction titer (FR50) of neutralizing antibodies of a given serum could be estimated from the focus reduction rates at several dilutions of the test serum when the latter was between 25-75% of the value of the control.

Animals

Natural and experimental infections of Japanese tree sparrows with Japanese encephalitis virus.

20 to 37 per cent of wild Japanese tree sparrows had neutralizing antibodies to Japanese encephalitis virus (JEV). Sparrows free of antibodies were inoculated with 10,000 plaque-forming units of JEV. None of the birds became clinically ill. Virus could be detected in blood plasma during the first 3 days after inoculation but not thereafter. Neutralizing antibodies started to appear at 4 days after inoculation and the response was of variable duration, in some cases extending up to 200 days after inoculation.

Animals

Proteome Profiling in Cerebrospinal Fluid Reveals Increased Levels of Peroxiredoxin 2 Discriminating Japanese Encephalitis Virus and Scrub Typhus Infection.

Japanese encephalitis virus (JEV) and scrub typhus (ST) are major etiological agents of acute encephalitis syndrome (AES) in India and South Asia. The pathophysiological changes at the molecular level caused by JEV and ST have yet to be studied in detail. The cerebrospinal fluid (CSF) proteomic landscape is a critical indicator of CNS pathology. Here, we conducted label-free quantitative proteomics on CSF from AES patients (n = 15) to identify etiology-specific differentially expressed proteins (DEPs) linked to encephalitis. The key DEPs were validated via ELISA in CSF (n = 49) and serum (n = 33). Our findings revealed 50 proteins exhibited differential expression across JEV and ST groups, with a notable subset of three proteins, PRDX2, KLK6, and TTR. PRDX2 and KLK6 were markedly elevated in JEV CSF (AUC: 0.8933 and 0.9689) but not in ST or non-JEV AES, while TTR was reduced in JEV yet elevated in ST (AUC: 0.5619 vs. 0.8238). Further, PRDX2 upregulation was validated in JEV-infected mouse brains and cortical neurons. Overexpression of PRDX2 in human neuroblastoma cells correlated with enhanced antiviral gene expression, p-STAT1, p-AKT (ser473), and viral replication. Thus, our comprehensive proteomic analysis of CSF identifies PRDX2 as an important circulatory protein, differentially expressed between JEV and ST, with high specificity and enhancing viral propagation, underscoring its role in viral propagation and pathogenesis.

Humans

Dual infections of mice: visceral larva migrans and sublethal infection with Japanese encephalitis virus.

Experiments in 3 weeks old albino mice with Toxocara canis and sublethal infection with JE virus established a marked synergestic effect in dually infected mice. The results are discussed to indicate the possible role of visceral larva migrans in creating exploxive outbreaks of "acute encephalopathy syndrome" in individuals having simultaneous infection with a virus (es) which, alone, might produce only mild illness. The nature of the possible mechanisms involved yet remains to be understood.

Animals

Attenuation of Japanese encephalitis virus by brain passage of guinea pigs.

JE virus was difficult to keep alive in guinea pig brain beyond 3 days and the virus was lost after 3 days. Ten cross passages from guinea pig to mice attenuated JE virus (GP10) and the result indicated that 3 passages could induce illness in guinea pig (GP4). GP4 and GP10 did not induce viremia to mice. 0.1 ml of 1:100,000 and 1:10,000 mouse suspension of GP4 inoculated into the author subcutaneously did not cause any uncomfortable symptom.

Animals

Role of RNA G-Quadruplexes in the Japanese Encephalitis Virus Genome and Their Recognition as Prospective Antiviral Targets.

G-quadruplexes (GQs) have been primarily studied in the context of cancer and neurodegenerative pathologies. However, recent research has shifted focus to their existence and functional roles in viral genomes, revealing GQ-regulated key pathways in various human pathogenic viruses. While GQ structures have been reported in the genomes of emerging and re-emerging viruses, RNA viruses have been understudied compared to DNA viruses, including notable examples such as human immunodeficiency virus-1, hepatitis C virus, Ebola virus, Nipah virus, Zika virus, and SARS-CoV-2. The flavivirus family, comprising the Japanese encephalitis virus (JEV), poses a significant global threat due to recurring outbreaks yet lacks approved antivirals. In this study, we identified and characterized eight putative G-quadruplex-forming motifs within essential genes involved in genome replication, assembly, and internalization in the host cell, conserved across different JEV isolates. The formation and stability of these motifs were validated through a multitude of biophysical and cell-based assays. The interaction and binding affinity of these motifs with the known GQ-binding ligand BRACO-19 were supported by biophysical assays, confirming the capability of these motifs to form GQ structures. Notably, BRACO-19 also exerted antiviral properties through reduction of viral replication and infectious virus titers as well as inhibition of viral protein expression, as evaluated by the cell-based assays. This comprehensive molecular characterization of G-quadruplex structures within the JEV genome highlights their potential as promising antiviral targets for intervention strategies against JEV infection through GQ-specific ligands.

G-Quadruplexes