Serological Evidence for Japanese Encephalitis and West Nile Virus Infections in Domestic Birds in Cambodia.

Mosquito-borne flaviviruses with an enzootic transmission cycle like Japanese encephalitis virus (JEV) and West Nile virus (WNV) are a major public health concern. The circulation of JEV in Southeast Asia is well-documented, and the important role of pigs as amplification hosts for the virus is long known. The influence of other domestic animals especially poultry that lives in high abundance and close proximity to humans is not intensively analyzed. Another understudied field in Asia is the presence of the closely related WNV. Such analyses are difficult to perform due to the intense antigenic cross-reactivity between these viruses and the lack of suitable standardized serological assays. The main objective of this study was to assess the prevalence of JEV and WNV flaviviruses in domestic birds, detailed in chickens and ducks, in three different Cambodian provinces. We determined the flavivirus seroprevalence using an hemagglutination inhibition assay (HIA). Additionally, we investigated in positive samples the presence of JEV and WNV neutralizing antibodies (nAb) using foci reduction neutralization test (FRNT). We found 29% (180/620) of the investigated birds positive for flavivirus antibodies with an age-depended increase of the seroprevalence (OR = 1.04) and a higher prevalence in ducks compared to chicken (OR = 3.01). Within the flavivirus-positive birds, we found 43% (28/65) with nAb against JEV. We also observed the expected cross-reactivity between JEV and WNV, by identifying 18.5% double-positive birds that had higher titers of nAb than single-positive birds. Additionally, seven domestic birds (10.7%) showed only nAb against WNV and no nAb against JEV. Our study provides evidence for an intense JEV circulation in domestic birds in Cambodia, and the first serological evidence for WNV presence in Southeast Asia since decades. These findings mark the need for a re-definition of areas at risk for JEV and WNV transmission, and the need for further and intensified surveillance of mosquito-transmitted diseases in domestic animals.

First dengue virus seroprevalence study on Madeira Island after the 2012 outbreak indicates unreported dengue circulation.

BACKGROUND: In 2012, the first dengue virus outbreak was reported on the Portuguese island of Madeira with 1080 confirmed cases. Dengue virus of serotype 1 (DENV-1), probably imported from Venezuela, caused this outbreak with autochthonous transmission by invasive Aedes aegypti mosquitoes. RESULTS: We investigated the seroprevalence among the population on Madeira Island four years after the outbreak. Study participants (n = 358), representative of the island population regarding their age and gender, were enrolled in 2012 in a cross-sectional study. Dengue antibodies were detected with an in-house enzyme-linked immunosorbent assay (ELISA) using the dimer of domain III (ED3) of the DENV-1 envelope protein as well as commercial Panbio indirect and capture IgG ELISAs. Positive ELISA results were validated with a neutralization test. The overall seroprevalence was found to be 7.8% (28/358) with the in-house ELISA, whereas the commercial DENV indirect ELISA detected IgG antibodies in 8.9% of the individuals (32/358). The results of the foci reduction neutralization test confirmed DENV-1 imported from South America as the causative agent of the 2012 epidemic. Additionally, we found a higher seroprevalence in study participants with an age above 60 years old and probable secondary DENV infected individuals indicating unreported dengue circulation before or after 2012 on Madeira Island. CONCLUSIONS: This study revealed that the number of infections might have been much higher than estimated from only confirmed cases in 2012/2013. These mainly DENV-1 immune individuals are not protected from a secondary DENV infection and the majority of the population of Madeira Island is still naïve for DENV. Surveillance of mosquitoes and arboviruses should be continued on Madeira Island as well as in other European areas where invasive vector mosquitoes are present.

Broad and long-lasting immune protection against various Chikungunya genotypes demonstrated by participants in a cross-sectional study in a Cambodian rural community.

Chikungunya virus (CHIKV) is an alphavirus circulating worldwide. Its presence in Asia has been reported since the 1950s, constituting the Asian genotype. Since 2005, strains from the Eastern, Central, and Southern African (ECSA) genotype have caused several outbreaks across Asia. Viruses from the ECSA genotype were also detected in Cambodia in late 2011 and led to an outbreak in a rural community in 2012. A former investigation from 2012 found a higher risk of infection in people younger than 40 years, suggesting a pre-existing herd immunity in the older Cambodian population due to infection with an Asian genotype. In 2016, we collected serum from equivalent numbers of individuals born before 1975 and born after 1980 that were also part of the 2012 study. We analyzed the 154 serum samples from 2016 for neutralization against the Cambodian ECSA isolate and three strains belonging to the Asian genotype. This experiment revealed that 22.5% (18/80) of the younger study participants had no CHIKV antibodies, whereas 5.4% (4/74) of the older population remained naive. Study participants infected during the ECSA outbreak had twofold neutralizing titers against the ECSA and the most ancient Asian genotype virus (Thailand 1958) compared to the other two Asian genotype viruses. The neutralization data also support the older population's exposure to an Asian genotype virus during the 1960s. The observed cross-reactivity confirms that the investigated CHIKV strains belong to a single serotype despite the emergence of novel ECSA genotype viruses and supports the importance of the development of a Chikungunya vaccine.