Analyses of phylogenetics, natural selection, and protein structure of clade 2.3.4.4b H5N1 Influenza A reveal that recent viral lineages have evolved promiscuity in host range and improved replication in mammals in North America

Influenza A virus has been circulating in birds from Eurasia for more than 146 years, but human infection has been sporadic. H5N1 (clade 2.3.4.4b) has recently infected hundreds of species of wild and domestic birds and mammals in North America. Infections include 70 people with two fatalities. We have developed an analytical bioinformatics, genomics, and structural workflow to understand better how H5N1 is circulating in North America and adapting to new host species.Our time-series analysis reveals that the circulation of H5N1 (clade 2.3.4.4b) in North America follows a distinct annual pattern, with cases in the United States consistently peaking each December. Separate from this seasonal cycle, our analysis also documents an increase in the total number of cases reported since 2021. We also show that H5N1 (clade 2.3.4.4b) spreads in North America as two distinct subclades of interest for human and animal health. These viral lineages have achieved a vast host range by efficiently binding the viral surface protein Hemagglutinin to both mammalian and avian cell surface receptors. This novel promiscuity of host range is concomitant with the additional strengthening of the Polymerase basic 2 viral proteins binding for mammalian and avian immune proteins. Once bound, the immune proteins will have diminished ability to fight the virus, thus allowing for more efficient replication of H5N1 in mammalian and avian cells than seen in the recent past. Finally, structural docking analyses predict that while most current antivirals remain effective, a fatal human isolate showed significantly reduced binding to multiple drugs from different classes. In conclusion, the H5N1 virus is causing an animal pandemic through promiscuity of host rage and strengthening ability to evade the innate immune systems of both mammalian and avian cells.