Human α-Defensin-6 Neutralizes Clostridioides difficile Toxins TcdA and TcdB by Direct Binding

Abstract

Rising incidences and mortalities have drawn attention to Clostridioides difficile infections (CDIs) in recent years. The main virulence factors of this bacterium are the exotoxins TcdA and TcdB, which glucosylate Rho-GTPases and thereby inhibit Rho/actin-mediated processes in cells. This results in cell rounding, gut barrier disruption and characteristic clinical symptoms. So far, treatment of CDIs is limited and mainly restricted to some antibiotics, often leading to a vicious circle of antibiotic-induced disease recurrence. Here, we demonstrate the protective effect of the human antimicrobial peptide α-defensin-6 against TcdA, TcdB and the combination of both toxins in vitro and in vivo and unravel the underlying molecular mechanism. The defensin prevented toxin-mediated glucosylation of Rho-GTPases in cells and protected human cells, model epithelial barriers as well as zebrafish embryos from toxic effects. In vitro analyses revealed direct binding to TcdB in an SPR approach and the rapid formation of TcdB/α-defensin-6 complexes, as analyzed with fluorescent TcdB by time-lapse microscopy. In conclusion, the results imply that α-defensin-6 rapidly sequesters the toxin into complexes, which prevents its cytotoxic activity. These findings extend the understanding of how human peptides neutralize bacterial protein toxins and might be a starting point for the development of novel therapeutic options against CDIs.

Trauma-toxicology: concepts, causes, complications

“…The growing mechanistic knowledge about the biology of C. difficile toxins has led to novel anti-toxin approaches, which might be useful in the future as supportive treatment options against C. difficile -associated diseases and/or post-traumatic complications. For instance, body-own antimicrobial peptides, such as certain defensins, were shown to inhibit TcdA/B and CDT either by direct interaction and formation of biologically inactive aggregates (Fischer et al 2020 ; Korbmacher et al 2020 ; Barthold et al 2022 ) or by inhibiting enzyme activities, as suggested for human α-defensin and TcdB (Giesemann et al 2008 ).…”