Avian influenza virus (AIV) poses a major threat to the poultry sector, especially in endemic areas such as Egypt. This research examined the contemporary circulation dynamics and molecular attributes of AIV subtypes within poultry farms in Qalyubia Governorate. Tissue samples obtained from duck, turkey, and chicken farms were subjected to viral isolation in specific pathogen-free embryonated chicken eggs, followed by hemagglutination (HA) assays. Viral isolates were confirmed by real-time PCR (RT-qPCR) targeting the matrix (M) gene and subsequent subtyping. The hemagglutinin (HA) gene of each subtype was exposed to partial sequencing, followed by phylogenetic analysis. Subtyping confirmed the circulation of three distinct viral strains: H5N1 in turkey, H5N8 in duck, and H9N2 in chicken. Phylogenetic investigation exposed that H5N1and H5N8 isolates belonged to the Eurasian lineage of clade 2.3.4.4b. The H9N2 isolate was assigned to the G-like lineage's genotype III. This finding is consistent with recent surveillance data since clade 2.3.4.4b emerged in Egypt in 2020. The existence of polybasic amino acids at the cleavage site confirms the high pathogenicity of the H5N1 and H5N8 isolates. In H5N8 and H5N1 isolates, the presence of N182, G221, Q222, and G224 in Receptor Binding Sites (RBS) indicates an avian-like alpha 2,3-sialic acid receptor binding preference. The continued circulation and genetic evolution within the antigenically distinct H5 clade 2.3.4.4b and H9N2 genotype III suggest a potential mismatch with current commercial H5 and H9N2 vaccines previously used in Egypt. we urge continual epidemiological and genetic surveillance of AIVs with continuous updated vaccination programs