Characterization of genotype V Japanese encephalitis virus isolates from Republic of Korea.

Japanese encephalitis (JE), caused by the Japanese encephalitis virus (JEV) infection, continues to pose significant public health challenges worldwide despite efficient vaccines. The virus is classified into five genotypes, among which genotype V (GV) was not detected for a long period after its initial isolation in 1952, until reports emerged from China and the Republic of Korea (ROK) since 2009. The characteristics of the virus are crucial in estimating its potential epidemiological impact. However, characterization of GV JEVs has so far been limited to two strains: Muar, the original isolate, and XZ0934, isolated in China. Two additional ROK GV JEV isolates, NCCP 43279 and NCCP 43413, are currently available, but their characteristics have not been explored. Our phylogenetic analysis revealed that GV virus sequences from the ROK segregate into two clades. NCCP 43279 and NCCP 43413 belong to different clades and exhibit distinct in vitro phenotypes. NCCP 43279 forms larger plaques but demonstrates inefficient propagation in cell culture compared to NCCP 43413. In vivo, NCCP 43279 induces higher morbidity and mortality in mice than NCCP 43413. Notably, NCCP 43279 shows more severe blood-brain barrier damage, suggesting superior brain invasion capabilities. Consistent with its higher virulence, NCCP 43279 displays more pronounced histopathological and immunopathological outcomes. In conclusion, our study confirms that the two ROK isolates are not only classified into different clades but also exhibit distinct in vitro and in vivo characteristics.

Seroprevalence of Japanese encephalitis virus in pig populations of Tamil Nadu, India: Exploring the tropical endemic link of virus.

Japanese encephalitis virus (JEV) is a major cause of encephalitis in Southeast Asia. Tamil Nadu, a state located in the southern part of India, contributes substantially to the national burden of human JE cases every year. However, limited information is available on the epidemiology of JE in pig populations of Tamil Nadu. A cross-sectional study was conducted to assess JEV prevalence in pig populations of Tamil Nadu. A total of 710 pigs reared in 118 farms across 10 districts of Tamil Nadu were sampled using multistage cluster random sampling. Serum samples were analyzed for their JEV status using Immunoglobulin M (IgM) and Immunoglobulin G (IgG) Enzyme-Linked Immunosorbent Assay (ELISA). At the animal-level, the apparent JEV seroprevalence was 60.4% (95% CI: 56.8% - 64.0%) and the true seroprevalence was 50.1% (95% CI: 47.0% - 53.2%). The herd-level apparent seroprevalence was 94.1% (95% CI: 88.1% - 97.5%) and the true seroprevalence was 93.3% (95% CI: 89.5% - 96.2%). The intensity of JEV circulation was high in all the districts, with seroprevalence ranging between 43% and 100%. Pigs across all age categories were seropositive and a high overall seroprevalence of 95.2% (95% CI: 76.2% - 99.9%) was recorded in pigs older than 12 months. JEV seropositivity was recorded in all the seasons but the prevalence peaked in the monsoon (67.9%, 95% CI: 61.1% - 74.2%) followed by winter (65.1%, 95%CI: 57.4% - 72.2%) and summer (53.3%, 95% CI: 47.8% - 58.8%) seasons. The results indicate that JEV is endemic in pigs populations of the state and a one health approach is essential with collaborative actions from animal and public health authorities to control JE in Tamil Nadu, India.

Development of immunochromatographic strip assay to detect recent infection of Japanese encephalitis virus in swine population.

Japanese Encephalitis (JE) is a mosquito borne re-emerging viral zoonotic disease. Sero-conversion in swine occurs 2-3 weeks before human infection, thus swine act as a suitable sentinel for predicting JE outbreaks in humans. The present study was undertaken with the objective of developing immunochromatographic strip (ICS) assay to detect recent infection of Japanese Encephalitis virus (JEV) in swine population. The two formats of ICS assay were standardized. In the first format, gold nanoparticles (GNP) were conjugated with goat anti-pig IgM (50 µg/ml) followed by spotting of recombinant NS1 protein (1 mg/ml) of JEV on NCM as test line and protein G (1 mg/ml) as control line. In the format-II, GNP were conjugated with rNS1 protein (50 µg/ml) followed by spotting of Goat anti-pig IgM (1 mg/ml) as test line and IgG against rNS1 (1 mg/ml) as control line. To decrease the non- specific binding, blocking of serum and nitrocellulose membrane (NCM) was done using 5% SMP in PBS-T and 1% BSA, respectively. Best reaction conditions for the assay were observed when 10 µl of GNP conjugate and 50 µl of 1:10 SMP blocked sera was reacted on BSA blocked NCM followed by reaction time of 15 mins. Samples showing both test and control line were considered positive whereas samples showing only control line were considered negative. A total of 318 field swine sera samples were screened using indirect IgM ELISA and developed ICS assay. Relative diagnostic sensitivity and specificity of format-I was 81.25% and 93.0% whereas of format-II was 87.50% and 62.93%, respectively. Out of 318 samples tested, 32 were positive through IgM ELISA with sero-positivity of 10.06% while sero-positivity with format-I of ICS was 8.1%. Owing to optimal sensitivity and higher specificity of format-I, it was validated in three different labs and the kappa agreement ranged from 0.80 to 1, which signifies excellent repeatability of the developed assay to test field swine sera samples for detecting recent JEV infection.

Spatio-temporal epidemiology of Japanese encephalitis virus infection in pig populations of eastern Uttar Pradesh, India, 2013-2022.

AIMS: Japanese encephalitis (JE) is endemic in India. Although pigs are considered important hosts and sentinels for JE outbreaks in people, limited information is available on JE virus (JEV) surveillance in pigs. METHODS AND RESULTS: We investigated the spatio-temporal distribution of JEV seroprevalence and its association with climate variables in 4451 samples from pigs in 10 districts of eastern Uttar Pradesh, India, over 10 years from 2013 to 2022. The mean seroprevalence of IgG (2013-2022) and IgM (2017-2022) was 14% (95% CI 12.8-15.2) and 10.98% (95% CI 9.8-12.2), respectively. Throughout the region, higher seroprevalence from 2013 to 2017 was observed and was highly variable with no predictable spatio-temporal pattern between districts. Seroprevalence of up to 60.8% in Sant Kabir Nagar in 2016 and 69.5% in Gorakhpur district in 2017 for IgG and IgM was observed, respectively. IgG seroprevalence did not increase with age. Monthly time-series decomposition of IgG and IgM seroprevalence demonstrated annual cyclicity (3-4 peaks) with seasonality (higher, broader peaks in the summer and monsoon periods). However, most variance was due to the overall trend and the random components of the time series. Autoregressive time-series modelling of pigs sampled from Gorakhpur was insufficiently predictive for forecasting; however, an inverse association between humidity (but not rainfall or temperature) was observed. CONCLUSIONS: Detection patterns confirm seasonal epidemic periods within year-round endemicity in pigs in eastern Uttar Pradesh. Lack of increasing age-associated seroprevalence indicates that JEV might not be immunizing in pigs which needs further investigation because models that inform public health interventions for JEV could be inaccurate if assuming long-term immunity in pigs. Although pigs are considered sentinels for human outbreaks, sufficient timeliness using sero-surveillance in pigs to inform public health interventions to prevent JEV in people will require more nuanced modelling than seroprevalence and broad climate variables alone.

Short Communication: Mosquito Histone 2A Protein Facilitate Japanese Encephalitis Virus Infection in the Mosquito.

Japanese encephalitis virus is mainly prevalent in the tropical and subtropical regions of Asia and Oceania. Through immunoprecipitation-mass spectrometry analysis using monoclonal antibodies targeting JEV E protein, we found that mosquito Histone 2A protein could bind to JEV particles. The binding of H2A and JEV was detected in the salivary gland and supernatant of mosquito cells. Furthermore, RNA interference experiments in vitro and in vivo confirmed that H2A protein promotes JEV infection in mosquitoes. In summary, we found that mosquito H2A is a factor that supports JEV infection and can potentially facilitate cross-species transmission of JEV.

Development of Colloidal Gold-Based Immunochromatographic Strips for Rapid Detection and Surveillance of Japanese Encephalitis Virus in Dogs across Shanghai, China.

Japanese encephalitis virus (JEV) causes acute encephalitis in humans and is of major public health concern in most Asian regions. Dogs are suitable sentinels for assessing the risk of JEV infection in humans. A neutralization test (NT) or an enzyme-linked immunosorbent assay (ELISA) is used for the serological detection of JEV in dogs; however, these tests have several limitations, and, thus, a more convenient and reliable alternative test is needed. In this study, a colloidal gold immunochromatographic strip (ICS), using a purified recombinant EDIII protein, was established for the serological survey of JEV infection in dogs. The results show that the ICSs could specifically detect JEV antibodies within 10 min without cross-reactions with antibodies against other canine viruses. The test strips could detect anti-JEV in serum with dilution up to 640 times, showing high sensitivity. The coincidence rate with the NT test was higher than 96.6%. Among 586 serum samples from dogs in Shanghai examined using the ICS test, 179 (29.98%) were found to be positive for JEV antibodies, and the high seropositivity of JEV in dogs in China was significantly correlated with the season and living environment. In summary, we developed an accurate and economical ICS for the rapid detection of anti-JEV in dog serum samples with great potential for the surveillance of JEV in dogs.

Isolation and identification of a duck-derived Japanese encephalitis virus and evaluation of its virulence in mice.

Birds including domestic and wild birds, as the amplifying or reservoir hosts of JEV, were sensitive to JEV infection and could develop a sufficiently high viremia to infect mosquitoes. However, most of JEV positive reports in birds were based on molecular detection, with few viruses isolated from clinical cases. In this study, one JEV strain, designated duck/2022-SD-1, was first isolated and identified from blood samples of ducks in 2022 in Shandong province of China. The JEV duck/2022-SD-1 strain was classified into genotype I cluster and shared 96.5 to 99.5 % nucleotide sequence identity with other GI JEV strains. Biological characteristics revealed that duck/2022-SD-1 possessed similar replication ability to a virulent strain Beijing/2020-1. Based on the amino acid identity comparison of E protein, amino acid sites responsible for JEV virulence were conserved between duck/2022-SD-1 and other virulence strains. Through virulence assays in mice, we further determined that duck/2022-SD-1 was a highly virulent JEV strain with highly neuroinvasive in mice, which is similar to the virulence of another virulent strain Beijing/2020-1. Thus, the potential threat of JEV strains originating from domestic birds should be brought to people's attention.

Potential for emergence of Japanese encephalitis in the European Union.

BACKGROUND AND OBJECTIVE: No autochthonous human cases of Japanese encephalitis (JE) have been reported to date in the European Union (EU). In this study, we assess the likelihood of Japanese encephalitis virus (JEV) introduction and transmission within the EU and propose outbreak response measures. RISK ASSESSMENT: Given the global geographical distribution of JEV, the probability of virus introduction into the EU is currently very low, with viremic bird migration being the most plausible pathway of introduction. However, this likelihood would significantly increase if the virus were to become established in the Middle East, Caucasus, Central Asia or Africa. Considering the environmental conditions that are expected to be conducive for virus circulation, there is a high likelihood of virus transmission within the EU after its introduction in environmentally suitable areas. The spread of the virus within the EU would likely occur through the movement of wild birds, pigs and mosquitoes. MITIGATION: To mitigate or potentially contain the emergence of JE in the EU, early detection of both human and animal cases will be crucial.

m6A modification associated with YTHDF1 is involved in Japanese encephalitis virus infection.

N6-methyladenosine (m6A), the most common modification in mammalian mRNA and viral RNA, regulates mRNA structure, stability, translation, and nuclear export. The Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus causing severe neurologic disease in humans. To date, the role of m6A modification in JEV infection remains unclear. Herein, we aimed to determine the impact of m6A methylation modification on JEV replication in vitro and in vivo. Our results demonstrated that the overexpression of the m6A reader protein YTHDF1 in vitro significantly inhibits JEV proliferation. Additionally, YTHDF1 negatively regulates JEV proliferation in YTHDF1 knockdown cells and YTHDF1 knockout mice. MeRIP-seq analysis indicated that YTHDF1 interacts with several interferon-stimulated genes (ISGs), especially in IFIT3. Overall, our data showed that YTHDF1 played a vital role in inhibiting JEV replication. These findings bring novel insights into the specific mechanisms involved in the innate immune response to infection with JEV. They can be used in the development of novel therapeutics for controlling JEV infection.

CD4 is an important host factor for Japanese encephalitis virus entry and replication in PK-15 cells.

Japanese encephalitis virus (JEV) is a flavivirus that is spread through mosquito bites and is the leading cause of viral encephalitis in Asia. JEV can infect a variety of cell types; however, crucial receptor molecules remain unclear. The purpose of this study was to determine whether porcine CD4 protein is a receptor protein that impacts JEV entry into PK15 cells and subsequent viral replication. We confirmed the interaction between the JEV E protein and the CD4 protein through Co-IP, virus binding and internalization, antibody blocking, and overexpression and created a PK-15 cell line with CD4 gene knockdown by CRISPR/Cas9. The results show that CD4 interacts with JEV E and that CD4 knockdown cells altered virus adsorption and internalization, drastically reducing virus attachment. The level of viral transcription in CD4 antibody-blocked cells, vs. control cells, was decreased by 49.1%. Based on these results, we believe that CD4 is a receptor protein for JEVs. Furthermore, most viral receptors appear to be associated with lipid rafts, and colocalization studies demonstrate the presence of CD4 protein on lipid rafts. RT‒qPCR and WB results show that virus replication was suppressed in PK-15-CD4KD cells. The difference in viral titer between KD and WT PK-15 cells peaked at 24 h, and the viral titer in WT PK-15 cells was 5.6 × 106, whereas in PK-15-CD4KD cells, it was only 1.8 × 106, a 64% drop, demonstrating that CD4 deficiency has an effect on the process of viral replication. These findings suggest that JEV enters porcine kidney cells via lipid raft-colocalized CD4, and the proliferation process is positively correlated with CD4.

Sero-molecular epidemiology of Japanese encephalitis virus in swine population of western Uttar Pradesh, India: Unraveling the geographical expansion of the virus.

BACKGROUND & OBJECTIVES: Swine is a good sentinel for forecast of Japanese encephalitis virus (JEV) outbreaks in humans. The present study was envisaged with objectives to know the sero-conversion period of JEV and to assess the prevalence of JEV in swine population of western Uttar Pradesh state of India. METHODS: A total of 252 swine serum samples were screened using IgM ELISA over the period of one year to determine the sero-conversion rate and compared seasonally to check the transmission peak of virus. Further, 321 swine blood and serum samples were collected from all seven divisions of western Uttar Pradesh to determine prevalence of JEV using real time RT-PCR and ELISA. RESULTS: Seasonal sero-conversion rate was high during monsoon and post-monsoon (32%) followed by winter (22.91%) and summer (10.71%) seasons. The sero-conversion was observed in all months indicating viral activity throughout the year in the region. The low degree of correlation was found between meteorological variables (day temperature, rainfall) and sero-conversion rate. A total of 52 samples (16.19%) were found positive by real time RT-PCR while sero-positivity of 29.91% was observed using IgG and IgM ELISA(s). The overall prevalence of JEV was 39.25%. INTERPRETATION & CONCLUSION: The presence of JEV was recorded throughout the year with peak occurrence during monsoon and post-monsoon season indicating that virus has spread its realm to western region of the state. The information generated in the present study will aid in initiating timely vector control measures and human vaccination program to mitigate risk of JEV infection in the region.

Mosquito surveillance on U.S military installations as part of a Japanese encephalitis virus detection program: 2016 to 2021.

Japanese encephalitis virus (JEV) continues to circulate throughout Southeast Asia and the Western Pacific where approximately 3 billion people in 24 countries are at risk of infection. Surveillance targeting the mosquito vectors of JEV was conducted at four military installations on Okinawa, Japan, between 2016 and 2021. Out of a total of 10,426 mosquitoes from 20 different species, zero were positive for JEV. The most abundant mosquito species collected were Aedes albopictus (36.4%) followed by Culex sitiens (24.3%) and Armigeres subalbatus (19%). Statistically significant differences in mosquito species populations according to location were observed. Changes in land use over time appear to be correlated with the species and number of mosquitoes trapped in each location. JEV appears to be absent from mosquito populations on Okinawa, but further research on domestic pigs and ardeid birds is warranted.

Infection of Feral Phenotype Swine with Japanese Encephalitis Virus.

Background: Japanese encephalitis virus (JEV) is a mosquito-borne zoonotic flavivirus and the leading cause of pediatric encephalitis in the Asian Pacific region. The transmission cycle primarily involves Culex spp. mosquitoes and Ardeid birds, with domestic pigs (Sus scrofa domestica) being the source of infectious viruses for the spillover of JEV from the natural endemic transmission cycle into the human population. Although many studies have concluded that domestic pigs play an important role in the transmission cycle of JEV, and infection of humans, the role of feral pigs in the transmission of JEV remains unclear. Since domestic and feral pigs are the same species, and because feral pig populations in the United States are increasing and expanding geographically, the current study aimed to test the hypothesis that if JEV were introduced into the United States, feral pigs might play a role in the transmission cycle. Materials and Methods: Sinclair miniature pigs, that exhibit the feral phenotype, were intradermally inoculated with JEV genotype Ib. These pigs were derived from crossing miniature domestic pig with four strains of feral pigs and were used since obtaining feral swine was not possible. Results: The Sinclair miniature pigs became viremic and displayed pathological outcomes similar to those observed in domestic swine. Conclusion: Based on these findings, we conclude that in the event of JEV being introduced into the United States, feral pig populations could contribute to establishment and maintenance of a transmission cycle of JEV and could lead to the virus becoming endemic in the United States.

The GTPase activity and isoprenylation of Swine GBP1 are critical for inhibiting the production of Japanese Encephalitis Virus.

Japanese encephalitis virus (JEV) is a flavivirus that cause severe neurological deficits. The guanylate-binding protein 1 (GBP1) gene is an interferon-stimulated gene and exerts antiviral functions on many RNA and DNA viruses via diverse mechanisms, however, the roles and the action modes of GBP1 in the antiviral effect on the production of JEV RNA and infectious virions remain to be clarified. In this study, we found that the RNA levels of swine GBP1 (sGBP1) in PK15 cells were up-regulated at the late stage of JEV infection. The overexpression of sGBP1 significantly inhibited the production of JEV while the knockdown of sGBP1 promoted the production of JEV. The GTPase activity and isoprenylation of sGBP1 both are critical for anti-JEV activity. The GTPase activity of sGBP1 is responsible for inhibiting the production of JEV genomic RNA. The isoprenylation of sGBP1 inhibited the expression and cleavage of JEV prM to decrease the yields of infectious virions, which may be associated with the interaction between sGBP1 and cellular proprotein convertase furin. Taken together, the study dissected the action modes of sGBP1with potent anti-JEV activity in more details.

Phylogeny and spatial distribution of Japanese encephalitis virus vector species in Cambodia.

In Southeast Asia, despite the use of Japanese encephalitis vaccines and vaccination coverage, Japanese encephalitis (JE) transmission is still a major public health issue. The main vectors of this virus are mosquitoes from the genus Culex, which diversity and density are important in Southeast Asia. The main vector species of Japanese encephalitis virus (JEV) in Cambodia belong to the Vishnui subgroup. However, their morphological identification solely based on the adult stage remains challenging, making their segregation and detection difficult. In order to identify and describe the distribution of the three main JEV vector species in Cambodia, namely Culex vishnui, Cx. pseudovishnui and Cx. tritaeniorhynchus, mosquito samplings were carried out throughout the country in different environments. Phylogenetic analysis of the cytochrome c oxidase subunit I (coI) gene using maximum-likelihood tree with ultrafast bootstrap and phylogeographic analysis were performed. The three main Culex species are phylogenetically separated, and represent two distinct clades, one with Cx. tritaeniorhynchus and the second with Cx. vishnui and Cx. pseudovishnui, the latter appearing as a subgroup of Cx. vishnui. The phylogeographic analysis shows a distribution of the Vishnui subgroup on the entire Cambodian territory with an overlapped distribution areas leading to a sympatric distribution of these species. The three JEV vector species are geographically well-defined with a strong presence of Cx. pseudovishnui in the forest. Combined with the presence of Cx. tritaeniorhynchus and Cx. vishnui in rural, peri-urban, and urban areas, the presence of JEV-competent vectors is widespread in Cambodia.

A Serosurvey of Japanese Encephalitis Virus in Monkeys and Humans Living in Proximity in Thailand.

Japanese encephalitis virus (JEV) is a member of the Flaviviridae family and one of Asia's most common causes of encephalitis. JEV is a zoonotic virus that is transmitted to humans through the bite of infected mosquitoes of the Culex species. While humans are dead-end hosts for the virus, domestic animals such as pigs and birds are amplification hosts. Although JEV naturally infected monkeys have been reported in Asia, the role of non-human primates (NHPs) in the JEV transmission cycle has not been intensively investigated. In this study, we demonstrated neutralizing antibodies against JEV in NHPs (Macaca fascicularis) and humans living in proximity in two provinces located in western and eastern Thailand by using Plaque Reduction Neutralization Test (PRNT). We found a 14.7% and 5.6% seropositive rate in monkeys and 43.7% and 45.2% seropositive rate in humans living in west and east Thailand, respectively. This study observed a higher seropositivity rate in the older age group in humans. The presence of JEV neutralizing antibodies in NHPs that live in proximity to humans shows the occurrence of natural JEV infection, suggesting the endemic transmission of this virus in NHPs. According to the One Health concept, regular serological studies should be conducted especially at the animal-human interface.

Molecular Detection and Genetic Characterization of Japanese Encephalitis Virus in Animals from 11 Provinces in China.

Japanese encephalitis virus (JEV), which uses a mosquito primary vector and swine as a reservoir host, poses a significant risk to human and animal health. JEV can be detected in cattle, goats and dogs. A molecular epidemiological survey of JEV was conducted in 3105 mammals from five species, swine, fox, racoon dog, yak and goat, and 17,300 mosquitoes from 11 Chinese provinces. JEV was detected in pigs from Heilongjiang (12/328, 3.66%), Jilin (17/642, 2.65%), Shandong (14/832, 1.68%), Guangxi (8/278, 2.88%) and Inner Mongolia (9/952, 0.94%); in goats (1/51, 1.96%) from Tibet; and mosquitoes (6/131, 4.58%) from Yunnan. A total of 13 JEV envelope (E) gene sequences were amplified in pigs from Heilongjiang (5/13), Jilin (2/13) and Guangxi (6/13). Swine had the highest JEV infection rate of any animal species, and the highest infection rates were found in Heilongjiang. Phylogenetic analysis indicated that the predominant strain in Northern China was genotype I. Mutations were found at residues 76, 95, 123, 138, 244, 474 and 475 of E protein but all sequences had predicted glycosylation sites at 'N154. Three strains lacked the threonine 76 phosphorylation site from non-specific (unsp) and protein kinase G (PKG) site predictions; one lacked the threonine 186 phosphorylation site from protein kinase II (CKII) prediction; and one lacked the tyrosine 90 phosphorylation site from epidermal growth factor receptor (EGFR) prediction. The aim of the current study was to contribute to JEV prevention and control through the characterization of its molecular epidemiology and prediction of functional changes due to E-protein mutations.

Japanese Encephalitis Enzootic and Epidemic Risks across Australia.

Japanese encephalitis virus (JEV) is an arboviral, encephalitogenic, zoonotic flavivirus characterized by its complex epidemiology whose transmission cycle involves reservoir and amplifying hosts, competent vector species and optimal environmental conditions. Although typically endemic in Asia and parts of the Pacific Islands, unprecedented outbreaks in both humans and domestic pigs in southeastern Australia emphasize the virus' expanding geographical range. To estimate areas at highest risk of JEV transmission in Australia, ecological niche models of vectors and waterbirds, a sample of piggery coordinates and feral pig population density models were combined using mathematical and geospatial mapping techniques. These results highlight that both coastal and inland regions across the continent are estimated to have varying risks of enzootic and/or epidemic JEV transmission. We recommend increased surveillance of waterbirds, feral pigs and mosquito populations in areas where domestic pigs and human populations are present.

Increasing seroprevalence of Japanese encephalitis virus in even-toed hoofed mammals (Artiodactyla species) associated with an upsurge of human cases in South Korea.

OBJECTIVES: The annual incidence of Japanese encephalitis (JE) has increased markedly in South Korea since 2010. We hypothesized that this increase was associated with higher frequencies of JE virus in animals. METHODS: We analyzed 5201 serum samples collected from even-toed hoofed mammals (Artiodactyla species) across South Korea from 2008 to 2012 using a stratified two-stage probability approach. RESULTS: The highest annual incidence of human JE cases and deaths occurred in 2010. Cases increased from six (no deaths) in 2008-2009 to 26 cases (seven deaths) in 2010. The JE virus seroprevalence in deer and elk fawns increased from 2.4% in 2008 to 24.1% in 2009, and in wild boars, it increased from 19.3% to 55.0% in the same period, which preceded the surge of human cases. Furthermore, the seroprevalence in calves increased from 15.3% in 2008 to 35.8% in 2010, and that in lambs and goat kids, increased from 8.5% in 2009 to 26.2% in 2010, which coincided with the surge in humans. CONCLUSION: Our findings show that the increased incidence of human JE in South Korea was temporally associated with an increasing seroprevalence in the Artiodactyla species. Surveillance of sentinel animals may be useful to predict the emergence of JE in humans.

Japanese Encephalitis Virus Genotypes 1 and 3 Isolation in Kochi, Japan.

Japanese encephalitis virus (JEV) is a mosquito-borne virus belonging to the JEV serocomplex within the genus Flavivirus, family Flaviviridae. It has 5 genotypes, G1-G5, based on the envelope (E) protein nucleotide sequence. JEV G3 circulated in Japan until the early 1990s when it was replaced by G1. JEV G3 was isolated from swine serum samples (sw/Kochi/1/2004) in the Kochi Prefecture, western Japan, in 2004. In addition, the 2018 isolates from pigs and cows (sw/Kochi/492/2018 and bo/Kochi/211/2018) in the same prefecture were identified as G3. The nucleotide sequencing results of the sw/Kochi/492/2018 and bo/Kochi/211/2018 polyprotein region differed from those of the sw/Kochi/1/2004 strain described in our previous report. Seven JEV isolates were identified as G1 in the same geographical area as that in this study. This result indicates that both JEV G1 and G3 are present in the Kochi area.