Zika and dengue viruses infecting wild-caught mosquitoes in an environmental protection area in Brazil

Abstract

Species of the genus Flavivirus are widespread in Brazil and are a major public health concern. The city of Sao Paulo is in a highly urbanized area with some green spaces which are used for recreation and where potential vertebrate hosts and mosquito vectors of these arboviruses can be found, a scenario that can contribute to the transmission of flaviviruses to humans. This study therefore sought to investigate natural flavivirus infection in mosquitoes collected in the Capivari-Monos Environmental Protection Area (EPA) in the south of the city. Monthly mosquito collections were carried out from March 2016 to April 2017 with CO2-baited CDC light traps. Specimens were identified morphologically and grouped in pools. A total of 260 pools of non-engorged females were inoculated into the C6/36 cell lineage after analysis by indirect immunofluorescence assay (IFA). IFA-positive specimens were tested by qRT-PCR with genus-specific primers targeting a region of ~260 nucleotides in the flavivirus NS5 gene, and the PCR products were sequenced to confirm and identify the flavivirus species. Anopheles (Kerteszia) cruzii and Wyeomyia (Prosopolepis) confusa were the most frequent species collected. Zika virus (ZIKV) nucleotide sequences were detected in three mosquito species, An. cruzii, Limatus durhami and Wy. confusa, and dengue virus 2 (DENV-2) sequences in Culex. spp. and Culex. (Mel.) vaxus. To our knowledge, this is the first report of natural isolation of DENV-2 and ZIKV in sylvatic species of mosquitoes in the Capivari-Monos EPA. Our findings suggest that DENV-2 is present in Culex mosquitoes, and ZIKV in Anopheles, Wyeomyia and Limatus. The flavivirus species identified here are of medical importance; surveillance is therefore recommended in this EPA, where vertebrates and mosquitoes can act as flavivirus hosts and vectors.