Japanese Encephalitis Virus Transmission Research Articles

SHORT REPORT: OBSERVATIONS ON THE MULTIPLE FEEDING BEHAVIOR OF CULEX TRITAENIORHYNCHUS (DIPTERA: CULICIDAE), THE VECTOR OF JAPANESE ENCEPHALITIS IN KERALA IN SOUTHERN INDIA

A study was undertaken in villages endemic for Japanese encephalitis (JE) in Kerala in southern India during the period 1998-2001 to determine the host-feeding pattern of Culex tritaeniorhynchus, the major vector of JE in southeast Asia. A total of 3,067 blood-engorged Cx. tritaeniorhynchus were tested and 2,553 (82.2%) of the samples could be identified. Culex tritaeniorhynchus had fed mainly (56.6%) on cattle. Pig feeding accounted 6.3% of the total samples. Some samples (n = 980, 38.3%) were of serologic mixed origin. Of 980 mixed blood-fed mosquitoes, 975 (99.5%) had imbibed blood from two distinct hosts and 5 (0.5%) imbibed blood from three distinct hosts. Mixed blood meals were mostly (96.7%) from cattle and goats. The epidemiologic implications of multiple feeding of Cx. tritaeniorhynchus on dampening (dead-end) hosts such as cattle and goats in the transmission of JE virus is discussed.

Mosquito host-feeding patterns and implications for Japanese encephalitis virus transmission in northern Australia and Papua New Guinea.

Japanese encephalitis (JE) virus spread to northern Australia during the 1990s, transmitted by Culex annulirostris Skuse and other mosquitoes (Diptera: Culicidae). To determine the relative importance of various hosts for potential vectors of JE virus, we investigated the host-feeding patterns of mosquitoes in northern Australia and Western Province of Papua New Guinea, with particular attention to pigs, Sus scrofa L. - the main amplifying host of JE virus in South-east Asia. Mosquitoes were collected by CDC light traps baited with dry ice and 1-octen-3-ol, run 16.00-08.00 hours, mostly set away from human habitations, if possible in places frequented by feral pigs. Bloodmeals of 2569 mosquitoes, representing 15 species, were identified by gel diffusion assay. All species had fed mostly on mammals: only <10% of bloodmeals were from birds. The predominant species was Cx. annulirostris (88%), with relatively few (4.4%) bloodmeals obtained from humans. From all 12 locations sampled, the mean proportion of Cx. annulirostris fed on pigs (9.1%) was considerably lower than fed on other animals (90.9%). Highest rates of pig-fed mosquitoes (>30%) were trapped where domestic pigs were kept close to human habitation. From seven of eight locations on the Australian mainland, the majority of Cx. annulirostris had obtained their bloodmeals from marsupials, probably the Agile wallaby Macropus agilis (Gould). Overall proportions of mosquito bloodmeals identified as marsupial were 60% from the Gulf Plains region of Australia, 78% from the Cape York Peninsula and 64% from the Daru area of Papua New Guinea. Thus, despite the abundance of feral pigs in northern Australia, our findings suggest that marsupials divert host-seeking Cx. annulirostris away from pigs. As marsupials are poor JE virus hosts, the prevalence of marsupials may impede the establishment of JE virus in Australia.

Vector competence of Australian mosquitoes (Diptera: Culicidae) for Japanese encephalitis virus.

Australian mosquitoes were evaluated for their ability to become infected with and transmit a Torres Strait strain of Japanese encephalitis virus. Mosquitoes, which were obtained from either laboratory colonies and collected using Centers for Disease Control and Prevention light traps baited with CO2 and octenol or reared from larvae, were infected by feeding on a blood/sucrose solution containing 10(4.5 +/- 0.1) porcine stable-equine kidney (PS-EK) tissue culture infectious dose50/mosquito of the TS3306 virus strain. After 14 d, infection and transmission rates of 100% and 81%, respectively, were obtained for a southeast Queensland strain of Culex annulirostris Skuse, and 93% and 61%, respectively, for a far north Queensland strain. After 13 or more days, infection and transmission rates of > 90% and > or = 50%, respectively, were obtained for southeast Queensland strains of Culex sitiens Wiedemann and Culex quinquefasciatus Say, and a far north Queensland strain of Culex gelidus Theobald. Although infection rates were > 55%, only 17% of Ochlerotatus vigilax (Skuse) and no Cx. quinquefasciatus, collected from far north Queensland, transmitted virus. North Queensland strains of Aedes aegypti L., Ochlerotatus kochi (Dönitz), and Verrallina funerea (Theobald) were relatively refractory to infection. Vertical transmission was not detected among 673 F1 progeny of Oc. vigilax. Results of the current vector competence study, coupled with high field isolation rates, host feeding patterns and widespread distribution, confirm the status of Cx. annulirostris as the major vector of Japanese encephalitis virus in northern Australia. The relative roles of other species in potential Japanese encephalitis virus transmission cycles in northern Australia are discussed.

Vertical transmission of Japanese encephalitis virus in Mansonia species, in an epidemic-prone area of southern India.

(2002). Vertical transmission of Japanese encephalitis virus in Mansonia species, in an epidemic-prone area of southern India. Annals of Tropical Medicine & Parasitology: Vol. 96, No. 4, pp. 419-420.

Presence of hemagglutination inhibition and neutralization antibodies to Japanese encephalitis virus in wild pigs on an offshore island in Singapore

A study was conducted to determine the presence of hemagglutination inhibition (HI) and neutralization antibodies against Japanese encephalitis virus (JEV) in wild pigs on an offshore island in Singapore. Blood samples were collected from 28 wild pigs on the island. All the sera tested with HI assay and plaque reduction neutralization tests were found to be positive for antibodies against JEV. Our results indicate that the wild pigs have been infected with JEV on the offshore island and there is JEV transmission.

Japanese encephalitis on Badu Island, Australia: the first isojation of Japanese encephalitis virus from Culex gelidus in the Australasian region and the role of mosquito host-feeding patterns in virus transmission cycles

During investigation of an outbreak of Japanese encephalitis (JE) in the Torres Strait, Australia, in 2000, mosquitoes were collected in Badu Island community and at a newly established communal piggery about 3 km from the community. A total of 94 285 mosquitoes, comprising 91 240 (96·8%) unengorged females, 1630 (1·7%) blood-engorged females and 1415 (1·5%) males, were processed for virus isolation. One isolate of JE virus was obtained from Culex gelidus, with a minimum infection rate of 12·4:1000. This is the first isolate of JE virus from Cx. gelidus in the Australasian region. No isolates were obtained from Cx. annulirostris, the primary implicated Australian JE vector. Analysis of mosquito host-feeding patterns, using gel diffusion, demonstrated that Cx. annulirostris and 5 other species fed predominately on mammals. Analysis of blood-fed mosquitoes collected within the community demonstrated that the proportion of Cx. annulirostris feeding on pigs in 2000 (2·3%) was significantly lower than that for the 1995–1997 period (31·3%). The removal of the pigs from Badu Island community has limited the contact between potential amplifying hosts and mosquitoes, thus potentially reducing the risk of transmission of JE virus to the human population.

Potential role of Armigeres subalbatus (Diptera: Culicidae) in the transmission of Japanese encephalitis virus in the absence of rice culture on Liu-chiu islet, Taiwan.

Mosquitoes known to be involved in the transmission of Japanese encephalitis virus (JE) on Taiwan typically develop in rice fields. However, recent serological evidence indicated that JE virus was being transmitted on Liu-Chiu, a rice-free islet. To identify the mosquito vector in this unusual epidemiological situation, 4 mosquito species commonly found in Liu-Chiu were evaluated for their vector competence for a strain of JE (CH1392) virus isolated from central Taiwan. Armigeres subalbatus (Coquillett) was the most susceptible species, indicating its status as a potential vector. In addition, an isolate of JE (T1P1) virus from Ar. subalbatus collected on Liu-Chiu readily infected the salivary glands of orally infected Ar. subalbatus originating from Liu-Chiu. The infection rate reached 79% (11/14) after a 20-d period of extrinsic incubation at 28 degrees C. We conclude that JE likely was transmitted between vertebrate hosts by Ar. subalbatus in this rice-free islet and that this species should be considered as a potential vector in similar ecological conditions.

Outbreak of Japanese encephalitis on the island of Saipan, 1990.

During October 1990, an outbreak of encephalitis occurred on Saipan. Although no virus was isolated, patients seroconverted to Japanese encephalitis (JE) virus, indicating the first known occurrence of JE on US territory since 1947. Ten cases occurred among a population of 40,000. The prevalence of antibody to JE virus among 234 lifelong Saipan residents surveyed after the outbreak was 4.2%. Age, household crowding, and lack of air conditioning were risk factors for infection. The seroprevalence in pigs, which are important amplifying hosts of JE virus, was 96% (n = 52). None of 288 stored serum specimens from lifelong Saipan residents sampled in 1984 were seropositive. These data suggest that JE virus was recently introduced onto Saipan and that peridomestic factors affected the risk of human infection. Transmission of JE virus probably ended with exhaustion of the supply of susceptible amplifying hosts. Surveillance for human cases and seroconversions in pigs during 1991 revealed no evidence of ongoing JE virus transmission.

Japanese encephalitis virus infection in mosquitoes reared from field-collected immatures and in wild-caught males.

Immature stages of mosquitoes collected in JE endemic areas of Karnataka, India between 1985 and 1987 were reared to adults and processed for the detection and isolation of Japanese encephalitis (JE) virus in an attempt to find naturally occurring vertical transmission of the virus. Males collected during 1985-1986 were also processed. A total of 15,785 adults reared from immatures and divided into 445 pools and 1,756 wild-collected males divided into 128 pools were processed using mosquito inoculation and immunofluorescence techniques. JE virus antigen was detected in 9 pools, 4 of which yielded JE virus. These were 2 pools of males and 1 pool of female Culex tritaeniorhynchus and 1 pool of male C. pseudovishnui, suggesting vertical transmission of JE virus in the mosquitoes.

Experimental Vertical Transmission of Japanese Encephalitis Virus by Culex Tritaeniorhynchus and other Mosquitoes

Vertical transmission of Japanese encephalitis virus to the F1 adult stage was demonstrated in Culex tritaeniorhynchus, Cx. annulus, Cx. quinquefasciatus, and Armigeres subalbatus. Transmission to the F1 larval stage was demonstrated in Cx. pipiens, Aedes vexans, Ae. alcasidi, and A. flavus. In Cx. tritaeniorhynchus, vertical transmission rates (the percentage of parent females transmitting to progeny) varied (12-100%). Filial infection rates (the percentage of progeny infected) for a given mosquito virus combination were markedly affected by the interval of time between parental infection and oviposition, suggesting that vertical infection was not transovarial in nature but occurred at oviposition. Filial infection rates for Cx. tritaeniorhynchus also varied widely by family and, as measured in F1 larvae, rates in excess of 20% were observed in a family. Filial infection rates in Cx. tritaeniorhynchus F1 adults were about 4 times lower than those in larvae. Japanese encephalitis virus was sexually transmitted from male to female Cx. tritaeniorhynchus.

A Longitudinal Study of the Prevalence of Japanese Encephalitis Virus in Adult and Larval Culex Tritaeniorhynchus Mosquitoes in Northern Taiwan

Adult females and larvae of Culex tritaeniorhynchus were collected year-round for 3 1/2 years at a site near Taipei, Taiwan. One hundred sixty-four isolates of Japanese encephalitis (JE) virus were obtained from approximately 142,000 adult females and 1 isolate of the virus was obtained from approximately 382,000 larvae. Virus was recovered from adult females every year, except the first, beginning in May. The single larval isolate was from specimens collected in June. The vertical transmission of JE virus in Cx. tritaeniorhynchus as a possible inter-epidemic viral survival mechanism is examined.

Encephalitis in Nepal: The Visitation of The Goddess of The Forest

Examination of hospital records from BMH Dharan and the large Nepalese hospital at Biratnagar has revealed a worrying trend in the incidence of acute encephalitis in the area of the British Military Cantonment at Dharan. The weight of available evidence strongly favours Japanese Encephalitis Virus (JEV) as the aetiological agent. Serological studies from healthy Nepalese from the Dharan area and animals from the Cantonment farm indicate that intense transmission of JEV occurs within the Cantonment area. Japanese Encephalitis (JE) must now be regarded as a serious threat to British Military personnel and dependants living in or visiting Nepal. Protective measures, particularly active immunisation are discussed.

Transplacental Japanese encephalitis virus (JEV) infection in mice during consecutive pregnancies.

Transplacental transmission of Japanese encephalitis virus (JEV) has been demonstrated in consecutive pregnancies of mice. Pregnant mice inoculated intraperitoneally with JEV transmit the virus to the foetus. When such female mice were mated again after 6 months, the virus could be isolated from the foetuses of the ensuing pregnancy. The incidence of abortion was increased significantly though the neonatal deaths were considerably less than during the first pregnancy. Intra-uterine infection occurred in spite of the presence of HAI antibodies against JEV in the preconceptional sera of the mice. The findings of the present study indicate the value of such a system for further investigations of the pathogenesis of JEV infection during pregnancy in humans.

Congenital infection of mice with Japanese encephalitis virus.

Transplacental transmission of Japanese encephalitis virus (JEV) when given intraperitoneally was demonstrated in pregnant mice as shown by isolation of the virus from placenta and fetal tissues. Furthermore, JEV could be isolated from the brain, liver, and spleen of newborn mice. The effect of JEV at different periods of gestation in pregnant mice was demonstrated for the first time, and the consequences of maternal infection on fetuses and neonates were studied. JEV infection during the 1st week of gestation caused a significantly higher number of fetal and neonatal deaths (66%) than during the 3rd week of gestation (13.8%). The number of abortions, stillbirths, and neonatal deaths was higher in infected mothers than in controls. No congenital abnormalities were found in any of the newborn mice. Sera obtained from 5-week-old health mice delivered by mothers infected during the 3rd week of gestation contained JEV hemagglutination inhibiting and immunoglobulin M antibodies. The results of these preliminary experiments show the usefulness of mice as a model for further elucidation of JEV infection during pregnancy and its effects on the fetus.

Transovarial transmission of Japanese encephalitis virus by Culex tritaeniorhynchus mosquitoes.

Transovarial transmission of Japanese encephalitis virus was demonstrated in three different strains of Culex tritaeniorhynchus mosquitoes by examination of F1 larvae.

Japanese Encephalitis Virus in Mosquito Salivary Glands

Culex tritaeniorhynchus and C. pipiens mosquitoes were infected with Japanese encephalitis virus either by intrathoracic injection or by membrane feeding. The virus maturation sites and the process of virus particle concentration in salivary gland cells were studied by electron microscopy. Occurrence of mature virions was primarily associated with intracytoplasmic viral matrices which were extraordinarily large and had a perinuclear location in C. pipiens mosquitoes. The other sign of virus replication was the proliferation of small spherical vesicles throughout the cytoplasm. It appeared that mature virions were entrapped in intracellular vacuoles and later released into the apical cavity of salivary gland cells through the fusion of these vacuoles with the apical plasma membrane. This process seemed to be associated with primary resynthesis of saliva in mosquitoes following blood feeding activity. Another type of shedding involved virus particles either singly or in mass being released directly through the apical plasma membrane. All of these events occurred only in cells of the lateral lobes of the salivary glands, which fact was confirmed by immunofluorescent staining of infected glands. The median lobe of mosquito salivary glands may have a minor or no role in the transmission of Japanese encephalitis virus.

Transovarial Transmission of Japanese Encephalitis Virus by Mosquitoes

Female Aedes albopictus and Aedes togoi mosquitoes infected with Japanese encephalitis virus either by intrathoracic inoculation or by ingestion of a virus-sucrose-erythrocyte mixture transmitted the virus to a small percentage of their F1 progeny. Adult F1 female Aedes albopictus thus infected transmitted the virus in turn to newly hatched chickens by feeding on them.

滋賀県下における日本脳炎の疫学的研究(1965～1973年)

In the ecology of Japanese encephalitis virus (JEV), the host animal for the virus during the non-epidemic season still remains uncertain.In this paper, the author sought to determine the animal species which play an important role in the transmission of JEV just before an epidemic.Sera were collected from wild herons, rats, snakes, lizards, and domestic fowls. Antibody-free dogs and rabbits, placed at two points as sentinels, were bled biweekly for two years. A prevalence of hemagglutination inhibition (HI) antibody was shown in dogs, rabbits, domestic fowls, herons and snakes, while not in rats and lizards. Seasonal transitions of HI titer chased in the sera of sentinel animals. The serum of each sentinel animal showed ascending HI titer late in July to early in August. All these results and other literature suggest that hibernating animals may play an important role as viral host during the winter season. Migratory birds may transmit JEV from endemic areas, such as South East Asia, to Japan and these two groups of animals may transmit JEV and cause epidemics in Japan.

The effects of environmental and physiological conditions of Culex tritaeniorhynchus on the pattern of transmission of Japanese encephalitis virus.

A log-probit regression was established between the infection rate of a colony of Culex tritaeniorhynchus and concentration of Japanese encephalitis virus in the mosquitoes' blood meals. The 50% infective dose of the virus for mosquitoes was 101.7 mouse ICLD50 per blood meal. A simple technique for detecting the oral secretion of virus by infected mosquitoes is described. The quantity of virus expelled by an infected the mosquito under prolonged incubation at 28°C was determined to be more than 104.0 mouse ICLD50. The interval between virus infection and initiation of virus secretion in mosquitoes was not influenced by mosquito age, concentration of sugar given to adults, history of blood engorgement in advance of infective blood meal, or history of blood engorgement following infective blood meal. A slight difference in this interval was observed among mosquitoes with different larval nutrition and those fed blood meals with different virus concentrations. However, the interval was markedly influenced by environmental temperature. Virus secretion was delayed and suppressed when infected mosquitoes were kept at 20°C. The suppression is suspected to be a consequence of the relatively slow physiological activity of mosquito tissues at low temperature rather than of a delay in the velocity of virus multiplication.

Japanese Encephalitis Virus in Northern Taiwan, 1969–1973 *

Sentinel pigs were bled and mosiquitoes were collected for isolation of Japanese encephalitis virus (JEV) between 1969 and 1973 in a rural area of northern Taiwan which reported a high annual incidence of human cases. The study site included a farmyard, schools, a bat cave, rice paddies and a heronry. Although Culex annulus was collected in every month of the year, isolations were made only in midsummer and only from Culex annulus and (once) from Culex tritaeniorhynchus. These isolates were usually collected from pig-related sources. Human cases in the vicinity of the study site were reported 3 to 6 weeks following the initial isolations from sentinel pigs. Isolations were made only for 4 to 8 weeks each summer. The isolation of JEV was not related to the availability of susceptible pigs or the spring or summer peaks in prevalence of Culex annulus. Despite periodic draining of rice paddies and the application of insecticides, sufficient numbers of Culex annulus were available to support transmission of JEV in each year of the study.