The antigenic landscape of N1 neuraminidase in human influenza A virus strains isolated between 2009 and 2020

Abstract

The immunogen composition of seasonal influenza vaccines is revised biannually to match the antigenic drift of the hemagglutinin of circulating influenza virus strains. Antibodies directed against the influenza neuraminidase also correlate with protection against influenza, yet the antigenic evolution of influenza neuraminidase remains underexplored. To evaluate the antigenic diversity of N1 neuraminidase, we generated a panel of immune sera in mice directed against 17 N1 neuraminidases derived from human H1N1 strains that were isolated between 2009 and 2020 and determined the neuraminidase inhibition (NAI) titers of these sera against a panel of 14 HxN1 viruses. The selected N1 neuraminidases cluster into two major phylogenetic groups while the NAI pattern revealed two antigenic groups corresponding to early and more recent H1N1 viruses. Sera raised against early N1 NAs inhibited NA activity from both early and recent H1N1 viruses equally well, whereas sera raised against more recent N1 NAs showed stronger NAI against recent H1N1 strains, indicating an asymmetric antigenic relationship between the two antigenic groups. Random forest modeling of antigenic distances indicated that the I321V, N386K/S and K432E substitutions played an important role in the N1 NA antigenic variation. Reverse-genetics experiments using mutant strains revealed that the N386K substitution in an early H1N1 virus increased susceptibility to inhibition by sera raised against recent N1 NAs, consistent with a role for the loss of the N-linked glycosylation motif and the acquisition of a lysine at this position near the lateral surface of the NA head. However, reciprocal substitutions in a recent N1 background did not restore the early antigenic phenotype, indicating a unidirectional effect. This finding suggests that anti-NA responses elicited against early pandemic H1N1 viruses remained effective against circulating N1 variants up to at least 2020.