Evolution of the Hemagglutinin Protein of the New Pandemic H1N1 Influenza Virus: Maintaining Optimal Receptor Binding by Compensatory Substitutions

Abstract

e Pandemic influenza A H1N1 (pH1N1) virus emerged in 2009. In the subsequent 4 years, it acquired several genetic changes in its hemagglutinin (HA). Mutations may be expected while virus is adapting to the human host or upon evasion from adaptive immune responses. However, pH1N1 has not displayed any major antigenic changes so far. We examined the effect of the amino acid substitutions found to be most frequently occurring in the pH1N1 HA protein before 1 April 2012 on the receptor-binding properties of the virus by using recombinant soluble HA trimers. Two changes (S186P and S188T) were shown to increase the receptor-binding avidity of HA, whereas two others (A137T and A200T) decreased binding avidity. Construction of an HA protein tree revealed the worldwide emergence of several HA variants during the past few influenza seasons. Strikingly, two major variants harbor combinations of substitutions (S186P/A137T and S188T/A200T, respectively) with opposite individual effects on binding. Stepwise reconstruction of the HA proteins of these variants demonstrated that the mutations that increase receptorbinding avidity are compensated for by the acquisition of subsequent mutations. The combination of these substitutions restored the receptor-binding properties (avidity and specificity) of these HA variants to those of the parental virus. The results strongly suggest that the HA of pH1N1 was already optimally adapted to the human host upon its emergence in April 2009. Moreover, these results are in agreement with a recent model for antigenic drift, in which influenza A virus mutants with high and low receptor-binding avidity alternate.T he spread of new animal influenza A viruses (IAVs) in the human population requires specific adaptations in several viral proteins. Clearly, the hemagglutinin (HA) receptor-binding and fusion protein needs to adapt to the new sialic acid (SIA) receptor repertoire present on epithelial cells in the human upper airway (1-3). However, optimal adaptation does not simply translate into the HA protein acquiring high-avidity binding to human receptors. Strong binding by HA of decoy receptors, for example, receptors present on soluble proteins in the mucus, is probably detrimental for virus replication. Also, efficient release of newly assembled viral particles requires a delicate balance between binding of HA to host cell receptors and neuraminidase (NA)-mediated cleavage of this interaction (4-6).The emergence of the new pandemic H1N1 (pH1N1) virus (7, 8) in 2009 has resulted in the accumulation of sequences of large numbers of pH1N1 virus isolates over a number of consecutive influenza seasons. Thus, for the first time in history, the evolution of a newly emerged IAV spreading in the human population can be followed in great detail. Apart from the possible selection of adaptive changes in HA that result in optimized receptor-binding properties, HA is also under selective pressure by the host immune response. IAV can escape from antibody recognition by the accumulation of amino acid substitut...

Expansion of Genotypic Diversity and Establishment of 2009 H1N1 Pandemic-Origin Internal Genes in Pigs in China

“…This is consistent with our earlier work (2,16,17), reports from other regions (38,39), and experimental coinfections of pigs with EA and TR viruses (40). At its emergence, the HA of pdm/09 virus was suggested to have been optimally adapted to humans, maintaining its receptor binding avidity via compensatory substitutions (41). Phenotypic differences and a heightened variability of the virus in infected swine suggested insufficient adaptation to pigs (5).…”

Amino acid substitution D222N from fatal influenza infection affects receptor-binding properties of the influenza A(H1N1)pdm09 virus

“…the two mutants with S185N already displayed recoverable virus, whereas rEng/195-wt virus did not, and at 24 h, the higher titers for the two mutants were significant (p o0.0001). The slow replication of the rEng/195-wt virus is probably due to differences in adaptation to the HTBE system; our data are in agreement with recent results that showed a delayed viral growth for the rEng/195-wt used as prototype virus from the first pandemic wave compared to a recombinant virus from the third wave (Baillie et al, 2012;de Vries et al, 2013;Elderfield et al, 2014). Similar to the wt, the rEng/195-D222N showed null viral growth at 12 h.p.i.…”

“…We also addressed the relevance of the mutation S185N, located at the antigenic site Sb (Xu et al, 2010). A different amino acid change, S185T, is reported to be conserved in all A (H1N1)pdm09 viruses in current circulation, and its role in the enhancement of receptor-binding avidity of the early A(H1N1) pdm09 virus has been addressed (de Vries et al, 2013;ECDC 2013;Elderfield et al, 2014;Klimov et al, 2012).…”

Identification of Amino Acid Substitutions Supporting Antigenic Change of Influenza A(H1N1)pdm09 Viruses

“…Additionally, finegrain differences in receptor specificity beyond the coarse distinction for ␣2,3-SA or ␣2,6-SA preference that allow more efficient replication in the human host and that may change the virus antigenically are currently incompletely understood (13,30). Because of the limited insight on the molecular basis for previous antigenic change of A(H1N1) viruses, we selected substitutions to introduce into the A/Netherlands/602/09 representative virus HA gene based on the following three approaches.…”