Wild passerines as potential carriers and sources of avian influenza viruses in Ukraine

Wild waterfowl and shorebirds are the primary reservoir of influenza A viruses in nature. The role of wild birds from other taxonomic groups remains insufficiently studied or is a subject of debate. This applies in particular to Passeriformes, the most diverse avian order, accounting for approximately 60% of the global bird population, where the role in circulation of influenza A viruses is underexplored. We used serological, virological, and PCR-based methods to survey avian influenza viruses in Passeriformes birds (65 species, 20 families) in Ukraine over a 20-year period, 2004-2025. Antibodies to influenza viruses were detected in serum and egg yolk of seven passerine species, with average seroprevalence 1.24% in sera and 8.94% in yolk samples. Seroprevalence varied across species, ranging from 1.96 to 27.2%. Virological screening resulted in the isolation of two viruses from Fieldfares (<i>Turdus pilaris</i>) of the subtypes H1N1 and H7N1. The overall infection rate based on virus isolation was 0.15%, while local infection rate in Fieldfares reached 11.1%. According to PCR results, 41 positive samples were detected, representing 3.61% of all tested birds (ranging from 1.42-9.1%), and by location ranged from 6.25-9.1%. Sequencing and phylogenetic analyses of H1N1 (Fieldfare), H7N1 (Fieldfare), H3N8 (Great Tit <i>Parus major</i>) influenza viruses confirmed them as Eurasian lineage low pathogenic avian influenza viruses and with close relatedness to viruses of the same subtypes circulating among wild waterfowl.