Abstract Thrombocytopenia is a common complication of influenza virus infection, and its severity predicts the clinical outcome of critically ill patients. The underlying cause(s) remain incompletely understood. In this study, in patients with an influenza A/H1N1 virus infection, viral load and platelet count correlated inversely during the acute infection phase. We confirmed this finding in a ferret model of influenza virus infection. In these animals, platelet count decreased with the degree of virus pathogenicity varying from 0% in animals infected with the influenza A/H3N2 virus, to 22% in those with the pandemic influenza A/H1N1 virus, up to 62% in animals with a highly pathogenic A/H5N1 virus infection. This thrombocytopenia is associated with virus-containing platelets that circulate in the blood. Uptake of influenza virus particles by platelets requires binding to sialoglycans and results in the removal of sialic acids by the virus neuraminidase, a trigger for hepatic clearance of platelets. We propose the clearance of influenza virus by platelets as a paradigm. These insights clarify the pathophysiology of influenza virus infection and show how severe respiratory infections, including COVID-19, may propagate thrombocytopenia and/or thromboembolic complications.

Understanding how emerging influenza viruses recognize host cells is critical in evaluating their zoonotic potential, pathogenicity, and transmissibility between humans. The surface of the influenza virus is covered with hemagglutinin (HA) proteins that can form multiple interactions with sialic acid-terminated glycans on the host cell surface. This multivalent binding affects the selectivity of the virus in ways that cannot be predicted from the individual receptor–ligand interactions alone. Here, we show that the intrinsic structural and energetic differences between the interactions of avian- or human-type receptors with influenza HA translate from individual site affinity and orientation through receptor length and density on the surface into virus avidity and specificity. We introduce a method to measure virus avidity using receptor density gradients. We found that influenza viruses attached stably to a surface at receptor densities that correspond to a minimum number of approximately 8 HA–glycan interactions, but more interactions were required if the receptors were short and human-type. Thus, the avidity and specificity of influenza viruses for a host cell depend not on the sialic acid linkage alone but on a combination of linkage and the length and density of receptors on the cell surface. Our findings suggest that threshold receptor densities play a key role in virus tropism, which is a predicting factor for both their virulence and zoonotic potential.

The influenza A virus (IAV) interacts with the glycocalyx of host cells through its surface proteins hemagglutinin (HA) and neuraminidase (NA). Quantitative biophysical measurements of these interactions may help to understand these interactions at the molecular level with the long-term aim to predict influenza infectivity and answer other biological questions. We developed a method, called multivalent affinity profiling (MAP), to measure virus binding profiles on receptor density gradients to determine the threshold receptor density, which is a quantitative measure of virus avidity toward a receptor. Here, we show that imaging of IAVs on receptor density gradients allows the direct visualization and efficient assessment of their superselective binding. We show how the multivalent binding of IAVs can be quantitatively assessed using MAP if the receptor density gradients are prepared around the threshold receptor density without crowding at the higher densities. The threshold receptor density increases strongly with increasing flow rate, showing that the superselective binding of IAV is influenced by shear force. This method of visualization and quantitative assessment of superselective binding allows not only comparative studies of IAV−receptor interactions, but also more fundamental studies of how superselectivity arises and is influenced by experimental conditions.

Influenza A viruses (IAVs) infect humans and a variety of other animal species. Infections with some subtypes of IAV were also reported in domestic cats and dogs. Besides animal health implications, close contact between companion animals and humans also poses a potential risk of zoonotic IAV infections. In this study, serum samples from different cat and dog cohorts were analyzed for IAV antibodies against 7 IAV subtypes, using three distinctive IAV-specific assays differing in IAV subtype-specific discriminatory power and sensitivity. Enzyme-linked immunosorbent assays against the complete hemagglutinin (HA) ectodomain or the HA1 domain were used, as well as a novel nanoparticle-based, virus-free hemagglutination inhibition (HI) assay. Using these three assays, we found cat and dog sera from different cohorts to be positive for antibodies against one or more IAV subtypes/strains. Cat and dog serum samples collected after the 2009 pandemic H1N1 outbreak exhibit much higher seropositivity against H1 compared with samples from before 2009. Cat sera furthermore displayed higher reactivity for avian IAVs than dog sera. Our findings show the added value of using complementary serological assays, which are based on reactivity with different numbers of HA epitopes, to study IAV antibody responses and for improved serosurveillance of IAV infections. We conclude that infection of cats and dogs with both human and avian IAVs of different subtypes is prevalent. These observations highlight the role of cats and dogs in IAV ecology and indicate the potential of these companion animals to give rise to novel (reassorted) viruses with increased zoonotic potential.