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The emergence of West Nile virus (WNV) and Usutu virus (USUV) in Europe resulted in significant outbreaks leading to avifauna mortality and human infections. Both viruses have overlapping geographical, host and vector ranges, and are often co-circulating in Europe. In Germany, a nationwide bird surveillance network was established to monitor these zoonotic arthropod-borne viruses in migratory and resident birds. In this framework, co-infections with WNV and USUV were detected in six dead birds collected in 2018 and 2019. Genomic sequencing and phylogenetic analyses classified the detected WNV strains as lineage 2 and the USUV strains as lineages Africa 2 (n = 2), Africa 3 (n = 3) and Europe 2 (n = 1). Preliminary attempts to co-propagate both viruses in vitro failed. However, we successfully cultivated WNV from two animals. Further evidence for WNV-USUV co-infection was obtained by sampling live birds in four zoological gardens with confirmed WNV cases. Three snowy owls had high neutralizing antibody titres against both WNV and USUV, of which two were also positive for USUV-RNA. In conclusion, further reports of co-infections in animals as well as in humans are expected in the future, particularly in areas where both viruses are present in the vector population., (© 2021 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

West Nile virus (WNV), a zoonotic arbovirus, has recently established an autochthonous transmission cycle in Germany. In dead-end hosts like humans and horses the WNV infection may cause severe symptoms in the central nervous system. In nature, WNV is maintained in an enzootic transmission cycle between birds and ornithophilic mosquitoes. Bridge vector species, such as members of the Culex pipiens complex and Aedes spp., also widely distributed in Germany, might transmit WNV to other vertebrate host species. This study determined and compared the vector competence of field-collected northern-German Cx. pipiens biotype pipiens and laboratory-reared Ae. vexans Green River (GR) for WNV lineage 1 (strain: Magpie/Italy/203204) and WNV lineage 2 (strain: "Austria") under temperatures typical for northern Germany in spring/summer and autumn. For assessment of vector competence, 7- to 14-day-old female mosquitoes were offered a WNV containing blood meal via Hemotek membrane feeding system or cotton-stick feeding. After incubation at 18°C respectively 24°C for 14 days engorged female mosquitoes were salivated and dissected for determination of infection, dissemination and transmission rates by reverse transcriptase quantitative real-time PCR (RT-qPCR). Both Ae. vexans GR and Cx. pipiens biotype pipiens were infected with both tested WNV strains and tested 14 days post-inoculation. Disseminated infections were detected only in Ae. vexans GR incubated at 18°C and in Cx. pipiens pipiens incubated at 24°C after infection with WNV lineage 1. Transmission of WNV lineage 1 was detected in Cx. pipiens pipiens incubated at 24°C. These results indicate that Cx. pipiens pipiens from Northern Germany may be involved in the transmission of WNV, also to dead-end hosts like humans and horses., (© 2020 Blackwell Verlag GmbH.)