Prediction,dynamics, and visualization of antigenic phenotypes of seasonal influenza viruses

Human seasonal influenza viruses evolve rapidly, enabling the virus population to evade immunity and reinfect previously infected individuals. Antigenic properties are largely determined by the surface glycoprotein hemagglutinin (HA), and amino acid substitutions at exposed epitope sites in HA mediate loss of recognition by antibodies. Here, we show that antigenic differences measured through serological assay data are well described by a sum of antigenic changes along the path connecting viruses in a phylogenetic tree. This mapping onto the tree allows prediction of antigenicity from HA sequence data alone. The mapping can further be used to make predictions about the makeup of the future A(H3N2) seasonal influenza virus population, and we compare predictions between models with serological and sequence data. To make timely model output readily available, we developed a web browser-based application that visualizes antigenic data on a continuously updated phylogeny.Seasonal influenza viruses evade immunity in the human population through frequent amino acid substitutions in their hemagglutinin (HA) and neuraminidase (NA) surface glycoproteins (1). To maintain efficacy, vaccines against seasonal influenza viruses need to be updated frequently to match the antigenic properties of the circulating viruses. To facilitate informed vaccine strain selection, the genotypes and antigenic properties of circulating viruses are continuously monitored by the World Health Organization (WHO) Global Influenza Surveillance and Response System (GISRS), with a substantial portion of the virological characterizations being performed by the WHO influenza Collaborating Centers (WHO CCs) (2).Antigenic properties of influenza viruses are measured in hemagglutination inhibition (HI) assays (3) that record the minimal antiserum concentration (titer) necessary to prevent crosslinking of red blood cells by a standardized amount of virus based on hemagglutinating units. An antiserum is typically obtained from a single ferret infected with a particular reference virus. For a panel of test viruses, the HI titer is determined by a series of twofold dilutions of each antiserum. An antiserum is typically potent against the homologous virus (the reference virus used to produce the antiserum), but higher concentrations (and hence lower titers) are frequently required to prevent hemagglutination by other (heterologous) test viruses. HI titers typically decrease with increasing genetic distance between reference and test viruses (1).Given multiple antisera raised against different reference viruses and a panel of test viruses, WHO CCs routinely measure the HI titers T_aβ of all combinations of test viruses a and sera β, resulting in a matrix of HI titers (see Fig. 1A). The HI titer of a test virus a using antiserum β raised against the reference virus b is typically standardized as H_aβ = log₂(T_bβ) ? log₂(T_aβ), i.e., the difference in the number of twofold dilutions between homologous and heterologous titer. Standardized log₂ titers from many HI assays can be visualized in two dimensions via multidimensional scaling—an approach termed “antigenic cartography” (4). Although standard cartography does not use sequence information, sequences have been used as priors for positions in a Bayesian version of multidimensional scaling (5). To infer contributions of individual amino acid substitutions to antigenic evolution, Harvey et al. and Sun et al. (6, 7) have used models that predict HI titer differences by comparing sequences of reference and test viruses.Open in a separate windowFig. 1.Antigenic data and models for HI titers. (A) A typical table reporting HI titer data. Each number in the table is the maximum dilution at which the antiserum (column) inhibited hemagglutination of red blood cells by a virus (row). The red numbers on the diagonal indicate homologous titers. A typical HI assay consists of all reciprocal measurements of the available antisera and reference viruses, and a number of test viruses that are measured against all antisera, but for which no homologous antiserum exists. To make measurements using different antisera comparable, we define standardized log-transformed titers H_aβ relative to the homologous titer. (B) Each HI titer between antiserum α and virus b can be associated with a path on the tree connecting the reference and test viruses a and b, respectively, indicated as a thick line. The tree model seeks to explain the antigenic differences as additive contributions of branches. (C) In the substitution model, the sum over branches on the tree is replaced by a sum of contributions of amino acid substitutions.Here, we show that antigenic properties of seasonal influenza viruses are accurately described by a model based on the phylogenetic tree structure of their HA sequences. We use the model to show that HI titers have a largely symmetric and tree-like structure that can be used to define an antigenic distance between viruses. We show that large-effect substitutions account for about half of the total antigenic change and that the effect of specific substitutions is dependent on the genetic background in which they occur. We further investigate the ability of HI measurements to predict dominant clades in the next influenza season. To visualize antigenic properties on the phylogenetic tree, we have integrated the models of antigenic distances and the raw HI titer data into nextflu.org—an interactive real-time tracking tool for influenza virus evolution (8).This comprehensive summary of HA sequences from past and current influenza viruses linked to their antigenic properties has the potential to inform vaccine strain selections and facilitate efforts to predict successful influenza lineages (9–13).