The Molecular Determinants of Antibody Recognition and Antigenic Drift in the H3 Hemagglutinin of Swine Influenza A Virus

Abstract

Influenza A virus (IAV) of the H3 subtype is an important respiratory pathogen that affects both humans and swine. Vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA) is the primary method used to control disease. However, due to antigenic drift, vaccine strains must be periodically updated. Six of the 7 positions previously identified in human seasonal H3 (positions 145, 155, 156, 158, 159, 189, and 193) were also indicated in swine H3 antigenic evolution. To experimentally test the effect on virus antigenicity of these 7 positions, substitutions were introduced into the HA of an isogenic swine lineage virus. We tested the antigenic effect of these introduced substitutions by using hemagglutination inhibition (HI) data with monovalent swine antisera and antigenic cartography to evaluate the antigenic phenotype of the mutant viruses. Combinations of substitutions within the antigenic motif caused significant changes in antigenicity. One virus mutant that varied at only two positions relative to the wild type had a >4-fold reduction in HI titers compared to homologous antisera. Potential changes in pathogenesis and transmission of the double mutant were evaluated in pigs. Although the double mutant had virus shedding titers and transmissibility comparable to those of the wild type, it caused a significantly lower percentage of lung lesions. Elucidating the antigenic effects of specific amino acid substitutions at these sites in swine H3 IAV has important implications for understanding IAV evolution within pigs as well as for improved vaccine development and control strategies in swine. IMPORTANCEA key component of influenza virus evolution is antigenic drift mediated by the accumulation of amino acid substitutions in the hemagglutinin (HA) protein, resulting in escape from prior immunity generated by natural infection or vaccination. Understanding which amino acid positions of the HA contribute to the ability of the virus to avoid prior immunity is important for understanding antigenic evolution and informs vaccine efficacy predictions based on the genetic sequence data from currently circulating strains. Following our previous work characterizing antigenic phenotypes of contemporary wild-type swine H3 influenza viruses, we experimentally validated that substitutions at 6 amino acid positions in the HA protein have major effects on antigenicity. An improved understanding of the antigenic diversity of swine influenza will facilitate a rational approach for selecting more effective vaccine components to control the circulation of influenza in pigs and reduce the potential for zoonotic viruses to emerge. I nfluenza A virus (IAV) of the H3 subtype is an important pathogen that infects both humans and swine. The main strategy used to prevent or reduce morbidity of IAV in humans is the implementation of vaccine programs (1). Likewise, swine producers employ commercially available and farm-specific autogenous vaccines to prevent IAV clinical disease in swine (2, 3). ...

Amino Acids in Hemagglutinin Antigenic Site B Determine Antigenic and Receptor Binding Differences between A(H3N2)v and Ancestral Seasonal H3N2 Influenza Viruses

“…The amino acid residues identified in the present study as critical for the antigenic difference between A(H3N2)v and seasonal H3N2 viruses are all located near the RBS in HA, which is in agreement with the observations in other studies that examined H3N2 antigenic evolution (19,23,24). Our study indicated that the amino acid mutations which switched the antigenic phenotype of MN/10 and BJ/92 influenced receptor binding.…”

“…In addition, we demonstrated that the amino acid substitutions that confer the antigenic phenotype impact both receptor specificity and antibody binding. Our findings provide additional evidence that residues near the HA RBS, especially those in antigenic site B, play significant role in the antigenic evolution of H3N2 viruses (23,24). Seasonal H3N2 viruses experience antigenic drift, which necessitates frequent updates of H3N2 candidate viruses for seasonal influenza vaccines (24,29).…”

“…antigenic phenotype of A(H3N2)v viruses. Residues 155, 156, 158, 189, and 193 in antigenic site B of MN/10 HA were examined because these residues are commonly associated with antigenic drift of human seasonal and swine influenza viruses (23,24,28,29). Reassortant MN/10 viruses bearing each of the 5 single mutations from MN/10 to BJ/92, Y155H, N156K, N158E, K189R, or K193S, were inhibited by MN/10 antiserum at slightly lower HI titers (160, 226, or 320) than wt MN/10.…”

“…Although amino acids close to the receptor binding site (RBS) in hemagglutinin (HA) are associated with antigenic evolution of human seasonal H3N2 and swine H3N2 viruses (23,24), the specific residues that explain why swine-origin A(H3N2)v viruses are antigenically distinct from the human strains have not been identified. This information is important for our understanding of the emergence of novel antigenic influenza virus strains that pose a threat to public health.…”

Detection and characterization of an H4N6 avian-lineage influenza A virus in pigs in the Midwestern United States

“…The diversity of contemporary strains co-circulating in the United States has been relatively well characterized in large part due to the US Department of Agriculture (USDA) surveillance efforts operated through the National Animal Health Laboratory Network and the National Veterinary Services Laboratories. The increased reporting and accessibility to publically available sequence data resulted in a more comprehensive genetic and antigenic understanding of circulating strains (Abente et al, 2016; Anderson et al, 2015; Anderson et al, 2013; Lewis et al, 2014). Furthermore, it facilitated the early detection of novel HA introductions into the swine population, such as a 2010–11 human H3 that has sustained onward transmission in the pig population (Rajao et al, 2015).…”