H1N1 influenza viruses varying widely in hemagglutinin stability transmit efficiently from swine to swine and to ferrets

A pandemic-capable influenza virus requires a hemagglutinin (HA) surface glycoprotein that is immunologically unseen by most people and is capable of supporting replication and transmission in humans. HA stabilization has been linked to 2009 pH1N1 pandemic potential in humans and H5N1 airborne transmissibility in the ferret model. Swine have served as an intermediate host for zoonotic influenza viruses, yet the evolutionary pressure exerted by this host on HA stability was unknown. For over 70 contemporary swine H1 and H3 isolates, we measured HA activation pH to range from pH 5.1 to 5.9 for H1 viruses and pH 5.3 to 5.8 for H3 viruses. Thus, contemporary swine isolates vary widely in HA stability, having values favored by both avian (pH >5.5) and human and ferret (pH ≤5.5) species. Using an early 2009 pandemic H1N1 (pH1N1) virus backbone, we generated three viruses differing by one HA residue that only altered HA stability: WT (pH 5.5), HA1-Y17H (pH 6.0), and HA2-R106K (pH 5.3). All three replicated in pigs and transmitted from pig-to-pig and pig-to-ferret. WT and R106 viruses maintained HA genotype and phenotype after transmission. Y17H (pH 6.0) acquired HA mutations that stabilized the HA protein to pH 5.8 after transmission to pigs and 5.5 after transmission to ferrets. Overall, we found swine support a broad range of HA activation pH for contact transmission and many recent swine H1N1 and H3N2 isolates have stabilized (human-like) HA proteins. This constitutes a heightened pandemic risk and underscores the importance of ongoing surveillance and control efforts for swine viruses