Survey of Japanese Encephalitis Virus in amplifying host and vector in four mountain districts of Nepal

Japanese Encephalitis Virus (JEV) is a mosquito-borne zoonotic virus that can cause severe neurologic disease and is the leading cause of viral encephalitis in Asia. Recently, laboratory-confirmed human cases of JEV have been reported from the mountain districts in Nepal, which were previously thought not to harbor JEV due to the higher elevation creating unfavorable breeding habitats for mosquitoes. Further investigation, however, found no history of travel to endemic areas in the majority of these human cases. This study consisted of a cross-sectional survey conducted in July and August 2010 and focused on four mountain districts known to have human cases of JEV. The objectives were to 1) estimate the prevalence of pigs with anti-JEV antibodies, which are an amplifying host for JEV, 2) to identify risk factors associated with JEV seropositivity, 3) to assess the presence of potential vectors of JEV and 4) to test potential JEV vectors for JEV infection. A total of 454 pig serum samples were collected and tested by competitive-ELISA. A 35-question questionnaire was administered to 109 pig owners and data was analyzed by multivariate logistic regression. In addition, mosquitoes were collected using 1-octen-3-ol baited CDC light traps. Mosquito samples were tested by Multiplex and RT-PCR for identification of mosquito species and detection of JEV, respectively. Seventy-six pigs (16.7%, 95% CI: 13.2, 20.2) tested positive for anti-JEV antibodies. No seropositive pig had been vaccinated against JEV nor came from known JEV endemic districts where previous exposure may have occurred. Prevalence of pigs with anti-JEV antibodies in each district was 16.7% (17/102), 4% (4/100), 6.6% (10/151) and 44.6% (45/101) in Sindhupalchowk, Dolakha, Solukhumbu and Kavrepalanchowk, respectively. Risk factors associated with increased risk of JEV seropositivity were summer abortion , wells as water source , urban location , and reported presence of mosquitoes . Increasing elevation was associated with decreased risk for JEV seropositivity. A total of 143 mosquitoes were collected from 12 different locations. Sequence analysis of PCR products from the pools of mosquitoes identified the presence of Culex vishnui, Culex quinquefasciatus and Aedes albopictus all known or suspected to be vectors of JEV. Other mosquito species with unknown vector potential were also identified including Culex annulus, Anopheles vagus, and Armigeres obturans. JEV was not detected in any of the mosquito pools; however, the small sample size, non-ideal trapping methods and time of season may have precluded identification of the virus. The identification of anti-JEV antibodies in pigs in the mountain districts, in conjunction with the presence of potential JEV vectors and historical reports of human cases of JEV in people without travel history to endemic areas, suggest that JEV is likely circulating in the mountain districts of Nepal. Infection with JEV should be considered a risk to residents and travelers in this area