Positive selection at the receptor-binding site of haemagglutinin H5 in viral sequences derived from human tissues

Highly pathogenic H5N1 avian influenza virus has spread through at least 45 countries in three continents. Despite the ability to infect and cause severe disease in humans, the virus cannot transmit efficiently from human to human. The lack of efficient transmission indicates the incompletion of the adaptation of the avian virus to the new host species. The required mutations for the complete adaptation and the emergence of a potential pandemic virus are likely to originate and be selected within infected human tissues. Differential receptor preference plays an important role in the species-tropism of avian influenza. We have analysed quasispecies of sequences covering the receptor-binding domain of the haemagglutinin gene of H5N1 viruses derived from fatal human cases. We employed a likelihood ratio test to identify positive-selection sites within the quasispecies. Nine of seventeen positive-selection sites identified in our analyses were found to be located within or flanking the receptor-binding domain. Some of these mutations are known to alter receptor-binding specificity. This suggests that our approach could be used to screen for mutations with significant functional impact. Our data provide new candidate mutations for the viral adaptation to a human host, and a new approach to search for new genetic markers of potential pandemic viruses.