PCA plots distinguish the frequencies of transmission bias loci (A) and vaccine evasion mutations (B). In (A) transmission bias mutations P-74, D-97, H-110, S-123, S-141, F-144, N-165 and M-226 (total = 8) have a high frequency in human isolates and closely clustering avian isolates but not in other avian isolates. In (B), vaccine-evasion mutations S-74, G-140, P-141, Y-144 and K-162 (total = 5) from Cattoli et al [12], which are responsible for resistance to the Mexican H5N2 vaccine strain commonly used in commercial poultry farms in Egypt are overrepresented in a different subcluster compared to the transmission bias mutations (A).