RTX-family toxin EhxA drives morphological remodeling and thrombogenesis in RBCs during enterohemorrhagic <i>Escherichia coli</i> infection.

Abstract

Enterohemorrhagic <i>Escherichia coli</i> (EHEC) causes thrombotic microangiopathy, yet the red blood cell (RBC)-centered mechanism has remained unclear. We identify the RTX-family hemolysin EhxA as the driver of RBC-mediated thrombogenesis. Deletion of <i>ehxA</i> abolishes Ca²⁺ influx, phosphatidylserine (PS) exposure, progression from discocyte to echinocyte to spherocyte, thrombin generation, RBC-endothelium adhesion, and RBC aggregation. Genetic complementation restores these readouts to wild type, and purified EhxA in bacteria-free assays recapitulates them while localizing to intact RBC membranes. By contrast, Δ<i>stx2</i> mutants do not elicit these RBC phenotypes, distinguishing this pathway from Shiga toxin-dependent effects. Multiple regression quantifies the link between PS exposure, morphology, and procoagulant outputs. In rats, infection with wild type increased RBC remodeling and venous thrombosis, whereas infection with Δ<i>ehxA</i> did not. Together, the data define an EhxA-Ca²⁺-PS pathway that drives RBC structural remodeling and procoagulant activation during EHEC infection and nominate RTX toxins as targets for preventing toxin-induced coagulopathies.