West Nile virus (WNV) is a flavivirus affecting mainly birds, and mammals, sometimes causing severe disease. Enzootic cycle is sustained between mosquitoes and wild birds, which are reservoir hosts and responsible for the virus dispersal during migration. WNV is endemic in Africa and bird migration is the most likely way for the translocation of the virus to Europe. WNV has been recognized for a long time in southern Europe, where it causes sporadic outbreaks, with few clinical cases, separated by long periods of epidemiological silence. However in North America, since its first appearance in 1999, the virus has spread relentlessly, causing a severe epidemic of important consequences for public health, livestock and wildlife. In Spain, WNV circulation has been revealed only very recently. Serological surveys in wild birds in southern wetlands have identified species exposed to the infection and involved in an enzootic cycle, highlighting the common coot as the species with the highest seroprevalence rates, and the only one in which seroconversion could be detected. On the other hand, WNV genome and specific antibodies have been repeatedly detected in endangered eagles in central Spain. In September 2007 in this region, WNV was isolated from two captive golden eagles showing symptoms of disease. Phylogenetically, WNV can be classified in 5 different lineages, but only lineages 1 (L1) and 2 (L2) have been associated with neurological disease, being both of them present in Europe. Our group (CISA-INIA) has developed a RRT-PCR for the simultaneous and differential detection of these two lineages in a single reaction. This method (rapid and highly sensitive and specific) may represent a helpful diagnostic tool in territories where the two lineages of WNV are supposed to co-circulate, for instance, Spain, where L1 has been isolated and an introduction of L2 may not be ruled out, given the bird migration routes between Europe and Africa. The present proposal aims at, on one hand, detect and characterize WNV strains circulating in Doñana and on the other hand, carry out a technologic transfer of the RRT-PCR for the specific detection of WNV L1 and L2 from CISA-INIA to EBD. We propose to collect and analyze a panel of common coot blood samples (that will be harvested during WNV transmission season in Doñana wetlands) with the aim of getting the detection (and if possible, the isolation) of WNV in these territories, where WNV circulation was probed.