Although the viral factors of host adaptation from domestic poultry to humans have been studied several times since the first cases of direct transmission of highly pathogenic avian influenza viruses from domestic poultry to humans were confirmed in 1997, the host-specific adaptation mechanisms from waterfowl to domestic poultry remain unknown. To study the mechanisms involved, a waterfowl-derived virus was passaged in a chicken fibroblast cell line. This passaged virus was found to have much higher growth titer than that of the original virus and several mutations were discovered in its genome. One of the most characteristics was an increase of the polymorphism of the internal genes. In addition, the general applicability of this property to the field isolates of influenza A viruses by database sequences analysis was confirmed, with the smallest amount of amino acid polymorphism in viral internal proteins observed in waterfowl-derived viruses, more in domestic poultry and the most in human-derived viruses. Although specific amino acid changes conserved in human-derived viruses were found, such amino acid changes were not observed in poultry-derived viruses.