Mouse Anti-influenza A H1N1 Neuraminidase monoclonal antibody, clone DE7

Characterising dynamics of Influenza A Viruses (IAV) within-host evolution is an active field of research which may lead to a better understanding of viral pathogenesis. Using a pregnant mouse model, a study has recently suggested that immune modulation during pregnancy could promote the emergence of IAV quasispecies with increased virulence. Herein, we assess the clinical relevance of these findings in humans. We studied IAV intra-host diversity (ihD) in pregnant (n = 36) and non-pregnant (n = 23) women hospitalized in Lyon for IAV infection (01/2015-05/2018). Whole IAV genomes present in nasopharyngeal samples were sequenced in duplicate to analyze reproducible intra-host single nucleotide variants (ihSNV). Counts, relative frequencies and locations of ihSNV were used as indicators of ihD. The median ihSNV/kb counts per segment were between 0 and 1.3. There was >81% ihSNV at relative frequencies between 1-5% for H1N1 and >51% for H3N2 IAV. No significant difference was noted between pregnant and non-pregnant women when considering all or only non-synonymous ihSNV. Seven convergent non-synonymous ihSNV were found; none were significantly associated with pregnancy. These results suggest that modulation of the immune system during pregnancy in humans does not impact IAV ihD, in contrast to mice.

Influenza A virus (IAV) increases the presentation of class I human leukocyte antigen (HLA) proteins that limit antiviral responses mediated by natural killer (NK) cells, but molecular mechanisms for these processes have not yet been fully elucidated. We observed that infection with A/Fort Monmouth/1/1947(H1N1) IAV significantly increased the presentation of HLA-B, -C, and -E on lung epithelial cells. Virus entry was not sufficient to induce HLA upregulation because UV-inactivated virus had no effect. Aberrant internally deleted viral RNAs (vRNAs) known as mini viral RNAs (mvRNAs) and defective interfering RNAs (DI RNAs) expressed from an IAV minireplicon were sufficient for inducing HLA upregulation. These defective RNAs bind to retinoic acid-inducible gene I (RIG-I) and initiate mitochondrial antiviral signaling (MAVS) protein-dependent antiviral interferon (IFN) responses. Indeed, MAVS was required for HLA upregulation in response to IAV infection or ectopic mvRNA/DI RNA expression. The effect was partially due to paracrine signaling, as we observed that IAV infection or mvRNA/DI RNA-expression stimulated production of IFN-beta and IFN-lambda 1 and conditioned media from these cells elicited a modest increase in HLA surface levels in naive epithelial cells. HLA upregulation in response to aberrant viral RNAs could be prevented by the Janus kinase (JAK) inhibitor ruxolitinib. While HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral nonstructural 1 (NS1) protein; we determined that NS1 limits cell-intrinsic and paracrine mechanisms of HLA upregulation. Taken together, our findings indicate that aberrant IAV RNAs stimulate HLA presentation, which may aid viral evasion of innate immunity. IMPORTANCE Human leukocyte antigens (HLAs) are cell surface proteins that regulate innate and adaptive immune responses to viral infection by engaging with receptors on immune cells. Many viruses have evolved ways to evade host immune responses by modulating HLA expression and/or processing. Here, we provide evidence that aberrant RNA products of influenza virus genome replication can trigger retinoic acid-inducible gene I (RIG-I)/mitochondrial antiviral signaling (MAVS)-dependent remodeling of the cell surface, increasing surface presentation of HLA proteins known to inhibit the activation of an immune cell known as a natural killer (NK) cell. While this HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral nonstructural 1 (NS1) protein, which limits RIG-I activation and interferon production by the infected cell.