Glycan Analysis and Influenza A Virus Infection of Primary Swine Respiratory Epithelial Cells THE IMPORTANCE OF NeuAcα2-6 GLYCANS

To better understand influenza virus infection of pigs, we examined primary swine respiratory epithelial cells (SRECs, the primary target cells of influenza viruses in vivo), as a model system. Glycomic profiling of SRECs by mass spectrometry revealed a diverse range of glycans terminating in sialic acid or Gal alpha Gal. In terms of sialylation, alpha 2-6 linkage was more abundant than alpha 2-3, and NeuAc was more abundant than NeuGc. Virus binding and infection experiments were conducted to determine functionally important glycans for influenza virus infection, with a focus on recently emerged swine viruses. Infection of SRECs with swine and human viruses resulted in different infectivity levels. Glycan microarray analysis with a high infectivity "triple reassortant" virus ((A/Swine/MN/593/99 (H3N2)) that spread widely throughout the North American swine population and a lower infectivity human virus isolated from a single pig (A/Swine/ONT/00130/97 (H3N2)) showed that both viruses bound exclusively to glycans containing NeuAc alpha 2-6, with strong binding to sialylated polylactosamine and sialylatedN-glycans. Treatment with mannosamine precursors of sialic acid (to alter NeuAc/NeuGc abundances) and linkage-specific sialidases prior to infection indicated that the influenza viruses tested preferentially utilize NeuAc alpha 2-6-sialylated glycans to infect SRECs. Our data indicate that NeuAc alpha 2-6-terminated polylactosamine and sialylated N- glycans are important determinants for influenza viruses to infect SRECs. As NeuAc alpha 2-6 polylactosamine glycans play major roles in human virus infection, the importance of these receptor components in virus infection of swine cells has implications for transmission of viruses between humans and pigs and for pigs as possible adaptation hosts of novel human influenza viruses.