Evidence of mosquito-transmitted flavivirus circulation in Piedmont, north-western Italy

Background: Flavivirus is a highly heterogeneous viral genus that includes important human pathogens and several viral strains with unknown zoonotic potential. Mosquito-borne flaviviruses have been isolated and characterized in Northern Italy: West Nile virus and Usutu virus were detected in mosquitoes and in different host species and recent studies provided evidence about the circulation of "insect Flavivirus" strains. Methods: In order to clarify the diffusion and the distribution of the mosquito-transmitted flaviviruses in Italy, we analyzed Culex and Ochlerotatus mosquitoes collected in 2009 and 2010 in an area divided evenly between hills and plains and where the landscape is dominated by mixed agricultural patches, rice fields, deciduous tree forests, and urban environments. Each mosquito pool was tested for the presence of Flavivirus strains and we characterized positive samples by genetic sequencing. Results: Positive mosquito pools revealed low infection prevalence, but suggested a continuous circulation of both Usutu virus and insect Flavivirus. Interestingly, phylogenetic analyses based on NS5 gene partial sequences showed a closer relationship among new Usutu virus strains from Piedmont and the reference sequences from the Eastern Europe, with respect to Italian samples characterized so far. Moreover, NS5 gene phylogeny suggested that mosquito flaviviruses found in Italy could belong to different lineages. Conclusions: Our results contribute to a wider point of view on the heterogeneity of viruses infecting mosquitoes suggesting a taxonomical revision of the Mosquito-borne Flavivirus group.