Yellow fever virus (YFV) causing a deadly viral disease is transmitted by the bite of infected mosquitoes. In Brazil, YFV is restricted to a forest cycle maintained between non-human primates and forest-canopy mosquitoes, where humans can be tangentially infected. Since late 2016, a growing number of human cases have been reported in Southeastern Brazil at the gates of the most populated areas of South America, the Atlantic coast, with Rio de Janeiro state hosting nearly 16 million people. We showed that the anthropophilic mosquitoes Aedes aegypti and Aedes albopictus as well as the YFV-enzootic mosquitoes Haemagogus leucocelaenus and Sabethes albiprivus from the YFV-free region of the Atlantic coast were highly susceptible to American and African YFV strains. Therefore, the risk of reemergence of urban YFV epidemics in South America is major with a virus introduced either from a forest cycle or by a traveler returning from the YFV-endemic region of Africa.

The Pantanal is a tropical, seasonal wetland of approximately 140,000 km 2 that is fed by tributaries of the upper Paraguay River in the centre of South America, which covers mainly Brazilian but also Paraguayan and Bolivian territories and is classified as one of the largest freshwater wetland ecosystems in the world (Alho 2005). The Brazilian Pantanal, which represents 85% of the total Pantanal area, is located within the states of Mato Grosso do Sul and Mato Grosso in Central-West Brazil, which are recognised as South and North Pantanal, respectively (Alho et al. 1988). This region is ecologically classified into sub-regions according to vegetation, flooding and physiography (Silva & Abdon 1988). The Nhecolândia sub-region that is situated in South Pantanal is one of the largest, comprising approximately one fifth of the total area, and is characterised by hundreds of shallow lakes that display varying degrees of salinity and coalescence with the system during floods (Adámoli 1982).Recent studies have demonstrated that some crocodilian species may be infected by WNV (Steinman et al. 2003, Jacobson et al. 2005 and that Alligator mississippiensis may also have a viremic load that affords the infection of blood-feeding Culicidae vectors, which suggests that in areas with high population densities of these alligators, juvenile individuals could play an important role in WNV transmission (Klenk et al. 2004). Taking these data into account, the unsuccessful efforts to detect WNV circulation in the avian hosts in Brazil and the widespread, high prevalence of the Crocodilia species, Caiman crocodilus yacare, in the Brazilian Pantanal (Campos et al. 2005), our strategy was to investigate the WNV circulation in the potential vectors, dead-end hosts and natural secondary amplifying hosts through the collection and testing of mosquitoes and serum samples of horses and wild caimans from the Nhecolândia sub-region. No morbidity was observed during the sampling period and only apparently healthy horses and caimans were sampled in the present study. MATERiALS AND METHoDSStudy area -In February 2009, mosquitoes and blood samples from horses and wild caimans were collected in six different cattle ranches of the Nhecolândia sub-region (18º20' 19º40'S and 57º14' 55º00'W) during the rainy season. Considering the free circulation of mosquitoes, horses and wild caimans in large ranches, sample collections for the study were undertaken in a 700-square-kilometre (70,000 ha) area (Figure). The collections for this study were authorised by the Brazilian Institute of Environment and Natural Resources (license IBAMA 18363-1/2009).Samples collections -Adult mosquitoes were captured at sites that were randomly selected using CDC automatic light traps and manual aspirators while landing to blood-feed on horses and research team members as routinely reported. Living mosquito specimens were transported to a field laboratory where they were immobilised by chilling and the species were identified through direct observation of the morphological ...

The wetlands of the Brazilian Pantanal host large concentrations of diverse wildlife species and hematophagous arthropods, conditions that favor the circulation of zoonotic arboviruses. A recent study from the Nhecolândia sub-region of Pantanal reported serological evidence of various flaviviruses, including West Nile virus and Ilheus virus (ILHV). According to the age of seropositive horses, at least three flaviviruses, including ILHV, circulated in the Brazilian Pantanal between 2005 and 2009. To extend this study, we collected 3,234 adult mosquitoes of 16 species during 2009 and 2010 in the same sub-region. Mosquito pool homogenates were assayed for infectious virus on C6/36 and Vero cell monolayers and also tested for flaviviral RNA by a group-specific real-time RT-PCR. One pool containing 50 non-engorged female specimens of Aedes scapularis tested positive for ILHV by culture and for ILHV RNA by real-time RT-PCR, indicating a minimum infection rate of 2.5 per 1000. Full-length genomic sequence exhibited 95% identity to the only full genome sequence available for ILHV. The present data confirm the circulation of ILHV in the Brazilian Pantanal.