Shiga Toxin 1a Blunts Shiga Toxin 2a-pathogenic Effects in Blood

Once released into human blood, Shiga toxins (Stx) interact with platelets and leukocytes, stimulating them to form aggregates and to release pathogenic extracellular vesicles (EV) containing Stx. These EV are considered the trigger driving the transition from bloody diarrhea to the life-threatening hemolytic uremic syndrome (HUS) during human infections by Stx-producing Escherichia coli (STEC). In children, HUS is characterized by hemolytic anemia, thrombocytopenia, and acute renal failure. The risk of any STEC-infected patient of developing HUS varies significantly depending on the Stx type produced by the bacteria, i.e., it is negligible for Shiga toxin 1 (Stx1), relevant for Shiga toxin 2 (Stx2), and considerably reduced when both toxins are present.To mimic what happens in the bloodstream of patients, human blood was challenged with Stx2a, Stx1a, or both toxins, and the formation of leukocyte/platelet aggregates was evaluated by direct-flow cytometric analysis. Pathogenic blood cell-derived EV were then isolated, their number and size determined by nanoparticle tracking analysis, and their proteins characterized by capillary Western blotting.We found that the presence of Stx1a during Stx2a challenge significantly reduced the formation of pathogenic EV, particularly the large (>300 nm) EV population causing HUS development. Notably, the amount of Stx2a significantly decreased in Stx1a + Stx2a-triggered EV with respect to Stx2a-induced EV.Our findings suggest that in STEC-infected children the presence of Stx1 in association with Stx2 reduces the risk of developing HUS by lowering the release of Stx2-containing blood cell-derived EV which are considered the main culprits for HUS onset.