Natural infection and vertical transmission of two flaviviruses (Yellow fever and Zika) in mosquitoes in primary forests in the Brazilian state of Rio de Janeiro (Diptera: Culicidae)

Abstract

BackgroundZika virus (ZIKV) was recently introduced in the American continent, probably transmitted by Aedes aegypti and possibly by Ae. albopictus and Culex quinquefasciatus in urban environments. ZIKV represents a known public health problem as it has been involved in newborn cases of congenital microcephaly in South America since 2005. The transmission of this virus in forested areas of other countries and its relative ubiquity in relation to its vectors and reservoirs raises suspicions of its adaptation to non-human modified environments (i.e., natural forests reserve) or on this continent, similar to those seen for Yellow fever virus (YFV). The objective of this work was to have an epidemiological monitoring tool mapping insects as well as circulating arboviruses in wild areas with low human interference. This study was based on the history of the insect flavivirus spreading cycle.\n\nMethods/Principal FindingsUsing a previously described sensitive PCR-based assay to assess the conserved NS5 region of the Flavivirus genus, both YFV partial genome and ZIKV were found in pools of Aedes albopictus, a sylvatic mosquito adapted to human-modified environments, and in Haemagogus leucocelaenus, a sylvatic mosquito.\n\nConclusionsThis is the first report of natural infection by ZIKV in mosquitoes in a sylvatic environment on the American continent. The wide distribution of these mosquitoes is probably important in the transmission of ZIKV. Vertical transmission indicates a higher efficiency for the maintenance and transmission of the virus in nature as well as the presence of the ZIKV in permanent character in the forest areas as it occurs with the YFV thus making more difficult the prevention of new cases of Zika in humans.\n\nAuthor SummaryArboviruses are diseases transmitted by arthropod vectors, hence the origin of the term ARthropod BOrne VIRUS, which is adopted since 1942. This work had as objective to survey the circulating insects as well as to detect the presence of viruses in them. Arboviruses circulate between insects and vertebrate hosts, having importance for promoting diseases in humans and animals. The diseases most known at the time, due to the recent cases reported by South America, are Dengue, Zika, Yellow Fever and Chikungunya. For this study, we used appropriate traps to collect the insects and their eggs in wild areas where there is little human interference. After collection, mosquitoes and / or eggs were identified and separated as to the source and species. The eggs were kept in laboratory conditions for the hatching of new insects. All the insects obtained were separated into pools to be macerated and thus extract the RNA from the viruses to be studied. Using molecular biology techniques, in our case the RT-PCR (Reverse Transcriptase Polymerase Chain Reaction), we amplified the RNA and in sequentially, we performed the sequencing reaction. With sequencing, it is possible to identify which virus material is present since each virus has a characteristic arrangement. For the identification of the sequences, we need to use some computational programs that guarantee us the correct result.