The genus Flavivirus, family Flaviviridae, includes a number of important arthropod-transmitted human pathogens such as dengue viruses, West Nile virus, Japanese encephalitis virus and yellow fever virus. In addition, the genus includes flaviviruses without a known vertebrate reservoir, which have been detected only in insects, particularly in mosquitoes, such as cell fusing agent virus, Kamiti River virus, Culex flavivirus, Aedes flavivirus, Quang Binh virus, Nakiwogo virus and Calbertado virus. Reports of the detection of these viruses with no recognized pathogenic role in humans are increasing in mosquitoes collected around the world, particularly in those sampled in entomological surveys targeting pathogenic flaviviruses. The presence of six potential flaviviruses, detected from independent European arbovirus surveys undertaken in the Czech Republic, Italy,Portugal, Spain and the UK between 2007 and 2010, is reported in this work. Whilst the Aedes flaviviruses, detected in Italy from Aedes albopictus mosquitoes, had already been isolated in Japan, the remaining five viruses have not been reported previously: one was detected in Italy, Portugal and Spain from Aedes mosquitoes (particularly from Aedes caspius), one in Portugal and Spain from Culex theileri mosquitoes, one in the Czech Republic and Italy from Aedes vexans, one in the Czech Republic from Aedes vexans and the last in the UK from Aedes cinereus. Phylogenetic analysis confirmed the close relationship of these putative viruses to other insect-only flaviviruses.

A total of 56 fleas were collected from mice, rats, and one hedgehog in national parks of mainland Portugal and the Madeira Island. All fleas were tested for the presence of bacteria of the genera Rickettsia and Bartonella using PCR assays. In fleas from mainland Portugal, we detected Rickettsia felis in one Archaeopsylla erinacei maura flea and in one Ctenophtalmus sp. In five Leptopsylla segnis fleas taken from rats in the Madeira Island, we identified Rickettsia typhi. In addition, in four fleas from the genera Ornithophaga and Stenoponia collect from mice and a rat in mainland Portugal, we detected the presence of two new Bartonella genotypes closely related to Bartonella elizabethae. Our findings emphasize the potential risk of flea-transmitted infections in mainland Portugal and the Madeira archipelago, and extend our knowledge of the potential flea vectors of human pathogens.

Mosquito surveillance in Europe is essential for early detection of invasive species with public health importance and prevention and control of emerging pathogens. In Portugal, a vector surveillance national program—REVIVE (REde de VIgilância de VEctores)—has been operating since 2008 under the custody of Portuguese Ministry of Health. The REVIVE is responsible for the nationwide surveillance of hematophagous arthropods. Surveillance for West Nile virus (WNV) and other flaviviruses in adult mosquitoes is continuously performed. Adult mosquitoes—collected mainly with Centre for Disease Control light traps baited with CO₂—and larvae were systematically collected from a wide range of habitats in 20 subregions (NUTS III). Around 500,000 mosquitoes were trapped in more than 3,000 trap nights and 3,500 positive larvae surveys, in which 24 species were recorded. The viral activity detected in mosquito populations in these years has been limited to insect specific flaviviruses (ISFs) non-pathogenic to humans. Rather than emergency response, REVIVE allows timely detection of changes in abundance and species diversity providing valuable knowledge to health authorities, which may take control measures of vector populations reducing its impact on public health. This work aims to present the REVIVE operation and to expose data regarding mosquito species composition and detected ISFs.