Receptor Specificity of Influenza A H3N2 Viruses Isolated in Mammalian Cells and Embryonated Chicken Eggs

Abstract

Isolation of human subtype H3N2 influenza viruses in embryonated chicken eggs yields viruses with amino acid substitutions in the hemagglutinin (HA) that often affect binding to sialic acid receptors. We used a glycan array approach to analyze the repertoire of sialylated glycans recognized by viruses from the same clinical specimen isolated in eggs or cell cultures. The binding profiles of whole virions to 85 sialoglycans on the microarray allowed the categorization of cell isolates into two groups. Group 1 cell isolates displayed binding to a restricted set of ␣2-6 and ␣2-3 sialoglycans, whereas group 2 cell isolates revealed receptor specificity broader than that of their egg counterparts. Egg isolates from group 1 showed binding specificities similar to those of cell isolates, whereas group 2 egg isolates showed a significantly reduced binding to ␣2-6-and ␣2-3-type receptors but retained substantial binding to specific O-and N-linked ␣2-3 glycans, including ␣2-3GalNAc and fucosylated ␣2-3 glycans (including sialyl Lewis x), both of which may be important receptors for H3N2 virus replication in eggs. These results revealed an unexpected diversity in receptor binding specificities among recent H3N2 viruses, with distinct patterns of amino acid substitution in the HA occurring upon isolation and/or propagation in eggs. These findings also suggest that clinical specimens containing viruses with group 1-like receptor binding profiles would be less prone to undergoing receptor binding or antigenic changes upon isolation in eggs. Screening cell isolates for appropriate receptor binding properties might help focus efforts to isolate the most suitable viruses in eggs for production of antigenically well-matched influenza vaccines.

Identification of Influenza A/PR/8/34 Donor Viruses Imparting High Hemagglutinin Yields to Candidate Vaccine Viruses in Eggs

“…A striking linkage between HA antigen yield and the source of HA-NA surface protein-coding genes was observed regardless of the PR8 gene source, reaching up to 65-fold differences when comparing 0.15 mg HA/100 eggs (by IDMS) from H5N1-Hubei to 9.5 mg HA/100 eggs from H7N2-NY, confirming previous reports [12,13,14,15,16,17,35]. The variability among HA antigen yields re-emphasized that virus surface genes are key determinants of viral growth in embryonated eggs, in agreement with other lines of evidence, such as positive selection in the HA and NA of candidate vaccine viruses selected for increased yields in eggs [36,37,38,39]. In addition, closely related HA-NA surface genes such as the two A(H3N2) and A(H5N1) viruses can impart substantially different yields.…”

Glycosylation changes in the globular head of H3N2 influenza hemagglutinin modulate receptor binding without affecting virus virulence

“…Recent glycan microarray analysis of the human H3N2 IAVs binding indicated a change in HA receptor binding affinity preferences from recognition of a broad range of SA-containing receptors by earlier viruses, to recognition of mainly long linear glycan structures by more contemporary isolates1314. Consistent with this, recent H3N2 IAVs have a reduced capacity to agglutinate a range of red blood cell types15161718 and to grow in cell cultures or embryonated chicken eggs1920. Data from several studies suggested a link between declining morbidity and mortality associated with H3N2 infection in humans and reduced binding to SA-containing receptors182122.…”

Recent H3N2 Viruses Have Evolved Specificity for Extended, Branched Human-type Receptors, Conferring Potential for Increased Avidity

“…eggs, MDCK cells). Moreover, recent H3 isolates showed increasingly reduced avidity and inconsistent specificity for human-type receptors in receptor binding assays (Gulati et al, 2013; Lin et al, 2012; Medeiros et al, 2001; Nobusawa et al, 2000; Stevens et al, 2010; Yang et al, 2015). These observations have suggested that human H3N2 viruses have not maintained consistent receptor specificity during evolution in the human population.…”