Dengue Hemorrhagic Fever in a Japanese Traveler with Pre-existing Japanese Encephalitis Virus Antibody

An adult Japanese man who had just returned from Thailand developed dengue hemorrhagic fever (DHF). A primary infection of dengue virus (DENV) was confirmed, specifically DENV serotype 2 (DENV-2), on the basis of the detection of the virus genome, a significant increase in the neutralizing antibody and the isolation of DENV-2. DHF is often observed following a secondary infection from another serotype of dengue virus, particularly in children, but this case was a primary infection of DENV. Japan is a non-endemic country for dengue disease. In fact, only Japanese encephalitis (JE) is known to be a member of the endemic flavivirus family. In this study, IgG antibody against Japanese encephalitis virus (JEV) was detected. JEV belongs to the family of dengue virus and prevails in Japan, particularly Kyushu. Among many risk factors for the occurrence of DHF, a plausible candidate could be a cross-reactive antibody-dependent enhancement (ADE) mechanism caused by JEV antibody. This indicates that most Japanese travelers who living in dengue non-endemic areas, particularly Kyushu, should be aware of the occurrence of DHF.     

Investigation of Dengue and Japanese Encephalitis Virus Transmission in Hanam,Viet Nam

This study investigated whether a large dengue epidemic that struck Hanoi in 2009 also affected a nearby semirural area. Seroconversion (dengue virus-reactive immunoglobulin G enzyme-linked immunosorbent assay) was high during 2009 compared with 2008, but neutralization assays showed that it was caused by both dengue virus and Japanese encephalitis virus infections. The findings highlight the importance of continued Japanese encephalitis virus vaccination and dengue surveillance.Dengue is an emerging health problem in northern Viet Nam, and in Hanoi, the capital, the largest recorded outbreak occurred in 2009, when 16,175 clinical cases were reported.¹ Clinical dengue incidence in northern Viet Nam is highest in urban Hanoi, and transmission commences around July, several months after the end of winter, and ceases in late December,¹ coinciding with changes in vector abundance.² Dengue cases have been detected sporadically in other northern provinces of Viet Nam,³ and 11% of dengue patients presenting to a Hanoi hospital during 2008 had come from other provinces.⁴ The extent to which this finding reflects local transmission or infections acquired during travel to endemic areas, such as Hanoi, is not clear. Serology is an important tool for understanding dengue virus (DENV) transmission, because a variable and sometimes large proportion of infections can be asymptomatic.⁵ Therefore, we used serology to investigate DENV infection and transmission in a semirural commune approximately 60 km from Hanoi.A cohort of 270 households was selected randomly from a semirural commune in Hanam province. Blood samples were collected in December of 2007, December of 2008, June of 2009, and April of 2010⁶ (i.e., outside the typical dengue season). The research was approved by the institutional review board of the National Institute of Hygiene and Epidemiology and the Oxford Tropical Research Ethics Committee, University of Oxford. All participants provided written informed consent. DENV-reactive immunoglobulin G (IgG) in sera was detected by indirect enzyme-linked immunosorbent assay (ELISA) using plates coated with purified DENV-1, -2, -3, and -4 virions (Panbio E-DEN01G; Alere, Brisbane, Australia). As per the manufacturer''s instructions, sera with absorbance values < 0.9 times the cutoff for the kit were classified as negative, sera with values > 1.1 times the cutoff were classified as positive, and sera with intermediate values were classified equivocal. ELISA seroconversion was defined as a change from negative to positive. Both Japanese encephalitis virus (JEV) and DENV circulate in northern Viet Nam, and there is substantial cross-reactivity between flavivirus antibodies.^7,8 Therefore, a subset of sera was assessed using the plaque reduction neutralization test (PRNT) with DENV-1, -2, -3, and -4 and JEV antigens performed as described previously.⁹In total, 606 participants provided blood samples in April of 2010; 29 of those participants had equivocal DENV IgG ELISA results and were excluded. Of the remaining 577 participants, 240 (41.6%) participants were male, and the median age was 31 years (interquartile range [IQR] = 13–45); 205 (35.5%) participants were DENV IgG-seropositive, and 372 (64.5%) participants were seronegative. Seroprevalence increased with age (Figure 1), and seropositive participants were significantly older than seronegative participants (median = 49 years, IQR = 32–59 years versus median = 25 years, IQR = 13–40 years, P < 0.001). Earlier samples were assessed to determine the proportions that seroconverted during 2008 or 2009 or were already seropositive at baseline in 2007; 3 participants did not have an earlier serum sample, and 13 participants had equivocal results, leaving 561 participants who could be assessed for seroconversion. In total, 143 of 561 (25.5%) participants were already seropositive at baseline, and 418 participants were seronegative, of whom 3 (0.7%, 95% confidence interval [95% CI] = 0.1–2.2%) participants seroconverted during 2008 compared with 43 of 415 (10.4%, 95% CI = 7.6–14.0%) participants seroconverted during 2009. Participants who seroconverted during 2009 were significantly younger (median = 32 years, IQR = 17–47 years) than participants who were already seropositive (median = 54 years, IQR = 42–63 years, P < 0.001). However, the proportion of children ages 10 years or less who seroconverted was very low (Figure 1).Open in a separate windowFigure 1.DENV IgG seroprevalence and seroconversion according to age. Results are shown as the proportion of participants that were DENV IgG-seropositive in either December of 2007 (gray bars) or April of 2010 (white bars). Also shown is the proportion of participants who seroconverted during 2009 (seronegative in December of 2008 and seropositive by April of 2010; black bars). The numbers shown below the chart are the denominator for each age group (N) and the denominator for seroconversion (N*; i.e., the number in each age group that was seronegative at baseline).Virus neutralization assays were performed on paired sera from participants who seroconverted and had sufficient sera remaining (31/46). The median age of this subset of participants was 31 years (IQR = 17–47 years), similar to the age of all seroconverters. DENV-1 and JEV but not DENV-2, -3, or -4 reactive antibodies were detected in post-infection sera from seroconverters by PRNT assay when titers were based on a 70% reduction in plaques (

First Isolation of Dengue Virus from the 2010 Epidemic in Nepal

Dengue is an emerging disease in Nepal and was first observed as an outbreak in nine lowland districts in 2006. In 2010, however, a large epidemic of dengue occurred with 4,529 suspected and 917 serologically-confirmed cases and five deaths reported in government hospitals in Nepal. The collection of demographic information was performed along with an entomological survey and clinical evaluation of the patients. A total of 280 serum samples were collected from suspected dengue patients. These samples were subjected to routine laboratory investigations and IgM-capture ELISA for dengue serological identification, and 160 acute serum samples were used for virus isolation, RT-PCR, sequencing and phylogenetic analysis. The results showed that affected patients were predominately adults, and that 10% of the cases were classified as dengue haemorrhagic fever/ dengue shock syndrome. The genetic characterization of dengue viruses isolated from patients in four major outbreak areas of Nepal suggests that the DENV-1 strain was responsible for the 2010 epidemic. Entomological studies identified Aedes aegypti in all epidemic areas. All viruses belonged to a monophyletic single clade which is phylogenetically close to Indian viruses. The dengue epidemic started in the lowlands and expanded to the highland areas. To our knowledge, this is the first dengue isolation and genetic characterization reported from Nepal.     

Contrasting the Practices of Virus Isolation and Characterization between the Early Period in History and Modern Times: The Case of Japanese Encephalitis Virus

Japanese encephalitis is a serious disease transmitted by mosquitoes. With its recent spread beyond the traditional territory of endemicity in Asia, the magnitude of global threat has increased sharply. While much of the current research are largely focused on changing epidemiology, molecular genetics of virus, and vaccination, little attention has been paid to the early history of virus isolation and phenotypic characterization of this virus. In this review, using this piece of history as an example, I review the transition of the concept and practice of virus isolation and characterization from the early period of history to modern times. The spectacular development of molecular techniques in modern times has brought many changes in practices as well as enormous amount of new knowledge. However, many aspects of virus characterization, in particular, transmission mechanism and host relationship, remain unsolved. As molecular techniques are not perfect in all respects, beneficial accommodation of molecular and biologic data is critically important in many branches of research. Accordingly, I emphasize exercising caution in applying only these modern techniques, point out unrecognized communication problems, and stress that JE research history is a rich source of interesting works still valuable even today and waiting to be discovered.     

Different Cross-Reactivities of IgM Responses in Dengue,Zika and Tick-Borne Encephalitis Virus Infections

Flaviviruses circulate worldwide and cause a number of medically relevant human diseases, such as dengue, Zika, yellow fever, and tick-borne encephalitis (TBE). Serology plays an important role in the diagnosis of flavivirus infections, but can be impeded by antigenic cross-reactivities among flaviviruses. Therefore, serological diagnosis of a recent infection can be insufficiently specific, especially in areas where flaviviruses co-circulate and/or vaccination coverage against certain flaviviruses is high. In this study, we developed a new IgM assay format, which is well suited for the specific diagnosis of TBE, Zika and dengue virus infections. In the case of TBE and Zika, the IgM response proved to be highly specific for the infecting virus. In contrast, primary dengue virus infections induced substantial amounts of cross-reactive IgM antibodies, which is most likely explained by structural peculiarities of dengue virus particles. Despite the presence of cross-reactive IgM, the standardized nature and the quantitative read-out of the assay even allowed the serotype-specific diagnosis of recent dengue virus infections in most instances.     

An Outbreak of Dengue Virus Serotype 2 Cosmopolitan Genotype in Nepal, 2017

Dengue virus (DENV) is one of the most prevalent neglected tropical diseases, with half of the world’s population at risk of infection. In Nepal, DENV was first reported in 2004, and its prevalence is increasing every year. The present study aimed to obtain and characterize the full-length genome sequence of DENV from the 2017 outbreak. Hospital-based surveillance was conducted in two provinces of Nepal during the outbreak. Acute-phase serum samples were collected from 141 clinically suspected dengue patients after the rainy season. By serological and molecular techniques, 37 (26.9%) and 49 (34.8%), respectively, were confirmed as dengue patients. The cosmopolitan genotype of DENV-2 was isolated from 27 laboratory-confirmed dengue patients. Genomic analysis showed many amino acid substitutions distributed mainly among the E, NS3, and NS5 genes. Phylogenetic analyses of the whole genome sequence revealed two clades (Asian and Indian) among DENV-2 isolates from Nepal. The DENV isolates from hilly and Terai areas were similar to Asian and Indian strains, respectively. Further genomic study on different DENV serotypes is warranted to understand DENV epidemics in Nepal, where there are limited scientific resources and infrastructure.     

Age-Related Susceptibility to Japanese Encephalitis Virus in Domestic Ducklings and Chicks

Ardeid birds and pigs are known as major amplifying hosts for Japanese encephalitis virus, and ducklings and chickens have been considered to play at best a minor role in outbreaks because of their low or absent viremia. We hypothesized that viremia of sufficient magnitude would develop in young ducklings (Anas platyrhynchos) and chicks (Gallus gallus) for them to serve as reservoir hosts and thereby contribute to the transmission cycle. Infection was associated with reduced weight gain in both species, and ducklings infected at 10 days of age or less showed overt clinical signs of disease. The mean peak viremia in birds of both species decreased as the age at infection increased from 2 to 42 days, indicating the importance of age of infection on magnitude of viremia in birds from both species, and suggesting that young poultry may be amplifying hosts of importance in disease-endemic regions.     

开江县306例流行性乙型脑炎流行病学调查分析

目的 探讨乙型脑炎发病规律和临床特征,为乙脑防制提供科学依据.方法 对开江县1985～2005年临床诊治的乙脑患者306例进行流行病学个案调查及分析,取2002～2005年乙脑病人血清,采用反向被动血凝抑制试验(RPHI)进行乙脑IgM抗体检测.结果 306例乙脑患者,分布于开江县20个乡镇168个村和居委会,农村发病是城镇的37.25倍,病死率为3.27%(10/306),发病高峰集中在8月,占63.73%(195/306),2～6岁儿童病例占78.43%(240/306),男女发病之比为1.41∶1,以散居儿童发病最多,占57.19%(175/306).临床上体温≥39℃占88.56%(271/306),患者以中、重型为多,占71.90%(220/306),随着年龄增大重型和极重型有增多的趋势.接种过乙脑疫苗占10.46%(32/306),家庭中有防蚊措施的占52.81%(47/89).临床病例血清学确诊率为63.33%(19/30). 结论 乙脑的防制策略,以农村2～6岁散居儿童为重点,采取防蚊灭蚊及全程足量接种乙脑疫苗,是控制乙脑发生和流行的有效措施.     

Case Report: West Nile Virus Encephalitis: The First Human Case Recorded in Brazil

A Brazilian ranch worker with encephalitis and flaccid paralysis was evaluated in the regional Acute Encephalitis Syndromic Surveillance Program. This was the first Brazilian patient who met the Centers for Disease Control and Prevention (CDC) confirmation criteria for West Nile virus disease. Owing to the overlapping of neurological manifestations attributable to several viral infections of the central nervous system, this report exemplifies the importance of human acute encephalitis surveillance. The syndromic approach to human encephalitis cases may enable early detection of the introduction of unusual virus or endemic occurrence of potentially alarming diseases within a region.     

Case Report: Dengue Virus Infection Triggering Thrombotic Thrombocytopenic Purpura in Pregnancy

We report a case of thrombotic thrombocytopenic purpura (TTP) that immediately followed symptomatic dengue virus infection in a pregnant lady. The patient developed dengue fever at 16 weeks of gestation, resulting in spontaneous abortion. Subsequently, fever reappeared with persistent thrombocytopenia and jaundice. Investigations revealed microangiopathic hemolysis; there was no evidence of disseminated intravascular coagulation. The TTP episode resolved after six cycles of therapeutic plasma exchange with fresh-frozen plasma. An ADAMTS13 (a disintegrin and metalloprotease with thrombospondin type 1 motif 13 repeats) activity assay, done during convalescence, showed normal activity. The patient had an uneventful second pregnancy and has remained free of TTP recurrence for more than 2 years now. We review the pathophysiological basis of TTP in dengue infection, and suggest that jaundice with disproportionate elevation of serum aspartate aminotransferase level in a patient with dengue should arouse the suspicion of TTP.     

Japanese Encephalitis Virus: The Emergence of Genotype IV in Australia and Its Potential Endemicity

A fatal case of Japanese encephalitis (JE) occurred in northern Australia in early 2021. Sequence studies showed that the virus belonged to genotype IV (GIV), a genotype previously believed to be restricted to the Indonesian archipelago. This was the first locally acquired case of Japanese encephalitis virus (JEV) GIV to occur outside Indonesia, and the second confirmed fatal human case caused by a GIV virus. A closely related GIV JEV strain subsequently caused a widespread outbreak in eastern Australia in 2022 that was first detected by fetal death and abnormalities in commercial piggeries. Forty-two human cases also occurred with seven fatalities. This has been the first major outbreak of JEV in mainland Australia, and geographically the largest virgin soil outbreak recorded for JEV. This outbreak provides an opportunity to discuss and document the factors involved in the virus’ spread and its ecology in a novel ecological milieu in which other flaviviruses, including members of the JE serological complex, also occur. The probable vertebrate hosts and mosquito vectors are discussed with respect to virus spread and its possible endemicity in Australia, and the need to develop a One Health approach to develop improved surveillance methods to rapidly detect future outbreak activity across a large geographical area containing a sparse human population. Understanding the spread of JEV in a novel ecological environment is relevant to the possible threat that JEV may pose in the future to other receptive geographic areas, such as the west coast of the United States, southern Europe or Africa.     

Potential Role of Birds in Japanese Encephalitis Virus Zoonotic Transmission and Genotype Shift

Japanese encephalitis (JE) is a vaccine-preventable disease caused by the Japanese encephalitis virus (JEV), which is primarily prevalent in Asia. JEV is a Flavivirus, classified into a single serotype with five genetically distinct genotypes (I, II, III, IV, and V). JEV genotype III (GIII) had been the most dominant strain and caused numerous outbreaks in the JEV endemic countries until 1990. However, recent data shows the emergence of JEV genotype I (GI) as a dominant genotype and it is gradually displacing GIII. The exact mechanism of this genotype displacement is still unclear. The virus can replicate in mosquito vectors and vertebrate hosts to maintain its zoonotic life cycle; pigs and aquatic wading birds act as an amplifying/reservoir hosts, and the humans and equines are dead-end hosts. The important role of pigs as an amplifying host for the JEV is well known. However, the influence of other domestic animals, especially birds, that live in high abundance and close proximity to the human is not well studied. Here, we strive to briefly highlight the role of birds in the JEV zoonotic transmission, discovery of birds as a natural reservoirs and amplifying host for JEV, species of birds susceptible to the JEV infection, and the proposed effect of JEV on the poultry industry in the future, a perspective that has been neglected for a long time. We also discuss the recent in vitro and in vivo studies that show that the newly emerged GI viruses replicated more efficiently in bird-derived cells and ducklings/chicks than GIII, and an important role of birds in the JEV genotype shift from GIII to GI.     

North American Birds as Potential Amplifying Hosts of Japanese Encephalitis Virus

Japanese encephalitis virus (JEV) is an emerging arbovirus, and inter-continental spread is an impending threat. The virus is maintained in a transmission cycle between mosquito vectors and vertebrate hosts, including birds. We detected variation in interspecies responses among North American birds to infection with strains of two different JEV genotypes (I and III). Several native North American passerine species and ring-billed gulls had the highest average peak viremia titers after inoculation with a Vietnamese (genotype I) JEV strain. Oral JEV shedding was minimal and cloacal shedding was rarely detected. The majority of birds, both viremic (72 of 74; 97.3%) and non-viremic (31 of 37; 83.8%), seroconverted by 14 days post-inoculation and West Nile virus-immune individuals had cross-protection against JEV viremia. Reservoir competence and serologic data for a variety of avian taxa are important for development of JEV surveillance and control strategies and will aid in understanding transmission ecology in the event of JEV expansion to North America.     

1976～2007年云南省流行性乙型脑炎流行病学特征分析

目的分析云南省流行性乙型脑炎（简称乙脑）流行病学特点,提供防制参考。方法收集全省乙脑疫情资料,用Excel建立数据库,用SPSS13．0统计软件进行统计分析。结果1976—2007年云南省共报告乙脑发病29693例,死亡3326例,年均发病率为2．46／10万,年均死亡率为0．29／10万,病死率为11．2％。发病率和病死率在20世纪70,80年代较高,90年代以来逐步下降,2007年发病率略有上升;发病地区主要集中在德宏、大理、保山、文山、红河、曲靖、昭通和西双版纳州（市）;全年均有发病,主要流行季节为7～10月,其中8月为高峰;报告病例中80．33％为≤15岁儿童。结论乙脑在云南省广泛分布,但主要流行地区为滇中、滇西和滇南地区;防制工作成效显著,但仍应加强高发地区儿童的预防接种和蚊虫控制工作。     

Molecular Mechanism and Role of Japanese Encephalitis Virus Infection in Central Nervous System-Mediated Diseases

The Japanese encephalitis virus (JEV) is the most common cause of neurodegenerative disease in Southeast Asia and the Western Pacific region; approximately 1.15 billion people are at risk, and thousands suffer from permanent neurological disorders across Asian countries, with 10–15 thousand people dying each year. JEV crosses the blood-brain barrier (BBB) and forms a complex with receptors on the surface of neurons. GRP78, Src, TLR7, caveolin-1, and dopamine receptor D2 are involved in JEV binding and entry into the neurons, and these receptors also play a role in carcinogenic activity in cells. JEV binds to GRP78, a member of the HSP70 overexpressed on malignant cells to enter neurons, indicating a higher chance of JEV infection in cancer patients. However, JEV enters human brain microvascular endothelial cells via an endocytic pathway mediated by caveolae and the ezrin protein and also targets dopamine-rich areas for infection of the midbrain via altering dopamine levels. In addition, JEV complexed with CLEC5A receptor of macrophage cells is involved in the breakdown of the BBB and central nervous system (CNS) inflammation. CLEC5A-mediated infection is also responsible for the influx of cytokines into the CNS. In this review, we discuss the neuronal and macrophage surface receptors involved in neuronal death.     

Occurrence of Japanese Encephalitis Virus Mosquito Vectors in Relation to Urban Pig Holdings

Japanese encephalitis virus (JEV) is transmitted to humans from pigs or birds by mosquitoes. In this study, the association between urban pig keeping and mosquito vectors was analyzed. A total of 7, 419 mosquitoes were collected overnight in urban households with and without pigs in Can Tho City, Vietnam. The most prevalent vectors were Culex tritaeniorhynchus (36%), Cx. gelidus (24%), and Cx. quinquefasciatus (15%), which were present in all parts of the city. Pigs were associated with increased numbers of Cx. tritaeniorhynchus. Traps close to pigs had higher numbers of Cx. tritaeniorhynchus and Cx. gelidus than traps close to humans. Increased number of persons in the household was associated with increased numbers of Cx. quinquefasciatus. We demonstrate that JEV vector species are present at urban households with and without pigs, and show that keeping pigs in an urban area increase the number of mosquitoes competent as vectors for JEV.     

从云南省宾川县蝙蝠体内分离到乙型脑炎病毒

１９９１年９月，从捕自云南省宾川县的１３２只双色帝蝙蝠和棕果蝙蝠体内分离到４株病毒。经电镜观察、血凝抑制试验和单克隆抗体免疫荧光试验证实为乙型脑炎病毒。结果表明蝙蝠在乙型脑炎病毒保存和传播中起重要作用。     

Surveillance of Japanese Encephalitis Virus Infection in Mosquitoes in Vietnam from 2006 to 2008

Japanese encephalitis virus (JEV) infection in mosquitoes was monitored in Vietnam from 2006 to 2008. A total of 15,225 mosquitoes, identified as 26 species in five genera were collected and 12,621 were grouped into 447 pools for examination of JEV infection by assays for cytopathic effects in C6/36 cells and by RT-PCR to detect flavivirus RNA. Three JEV strains were isolated from Culex tritaeniorhynchus Giles collected in northern and southern Vietnam and two JEV strains were isolated from Culex vishnui Theobald collected in the highlands of Vietnam. Genetic and phylogenetic analyses, based on complete E gene nucleotide sequences, revealed that the five JEV strains were classified into the genotype I group and six amino acid differences were found in these five strains. These results indicated that multiple JEV genotype I populations are circulating countrywide in Vietnam, transmitted by bites of their Cx. tritaeniorhynchus and Cx. vishnui.     

流行性乙型脑炎患者血清IgG抗体对乙脑病毒蛋白的识别

目的：了解流行性乙型脑炎病毒各蛋白成分在刺激机体免疫应答方面所起的作用，给亚单位疫苗研制提供理论依据，进行该项研究。方法：建立乙脑病毒免疫转印方法，应用该法检查乙脑病人血清IgG抗体对乙脑病毒蛋白的反应情况，分析正常人群中筛选出的乙脑病毒隐性感染者血清IgG对乙脑病毒蛋白的反应性，并比较二者的差异。结果：经检测，绝大部分病人血清IgG可识别E蛋白，部分病人尚可识别NS5，NS3，NS1蛋白，而隐性感染者血清IgG对98kd（NS5）乙脑病毒蛋白的反应性显著低于乙脑患者。结论：研究结果显示，E蛋白是乙脑病人血清IgG识别的主要蛋白，在免疫保护方面起着主要作用。隐性感染者对NS5蛋白的反应机率明显低于显性感染者，提示NS5蛋白在乙脑病毒的致病过程中起重要作用。