Enhanced Visualization of Influenza A Virus Entry Using Virus-View Atomic Force Microscopy

Abstract

Virus entry begins with attachment of virions to the cell surface, multivalent binding of viral proteins to receptors, signaling, and endocytosis. Using ViViD-AFM (Virus-View Dual confocal and Atomic Force Microscopy), we visualized the nanoscale morphology of influenza A virus (IAV) virions interacting with the cell membrane during virus entry. Following attachment to the cell surface, spherical IAV (90-100 nm in diameter) diffused in a sialic acid- and neuraminidase-dependent manner. Reduced diffusion signified the onset of clathrin coat assembly, followed by formation of actin-rich ruffles that promoted pit closure and IAV endocytosis. Cell surface ruffles sheared filamentous IAV (>1{micro}m in length) into shorter fragments that became internalized. ViViD-AFM is a powerful tool that provides nanoscale morphological insights of virus-cell membrane interplay in living cells.