Tracking the Pathways of West Nile Virus: Phylogenetic and Phylogeographic Analysis of a 2024 Isolate from Portugal

Birds are natural hosts for numerous zoonotic viral pathogens, including West Nile virus, which is transmitted by mosquitoes. During migration, birds can act as vectors for the geographic spread of viruses. WNV is endemic in Portugal, causing annual outbreaks, particularly in horses. Here, we report the first detection of an avian WNV strain isolated from a wild bird (<i>Astur gentilis</i>) collected in Portugal in mid-September 2024. Phylogenetic and phylogeographic analyses were conducted to trace the virus's origin and potential transmission routes, integrating the obtained full-length genomic sequence with a dataset of WNV strains from Africa and Europe (1951-2024). Phylogenetic analysis of 92 WNV sequences spanning lineages 1-5 positioned the 2024 isolate within lineage 1a. Results obtained using phylodynamics-based analysis showed that this isolate likely originated in Africa and reached Portugal via Spain's Cádiz coast, confirming previously described WNV dispersal patterns between Africa and Europe. The data suggest a migratory route from West Africa to Europe, extending through countries such as Senegal, Mauritania, Morocco, Portugal, Spain, Italy, and France, indicating a reciprocal flow of the virus back into Africa. These transmission routes match the migratory paths of Afro-Palearctic bird species, emphasizing the role of migratory birds in the long-distance spread of WNV.