Clostridioides difficile infection (CDI) remains a leading cause of healthcare‑associated morbidity and mortality. While disease symptoms are primarily mediated by toxins A and B, a growing proportion of clinically severe and epidemic strains also express a third virulence factor, the binary toxin (CDT). CDT expression has been associated with increased disease severity, enhanced colonization, and worse clinical outcomes. Despite this, current vaccine strategies largely overlook CDT as a protective target.
Our researchers identified Receptor Binding Domain 2 (RBD2) of the CDT binding component (CDTb) as a critical and conserved structural domain required for toxin function. Sequence and structural analyses demonstrated that RBD2 is highly conserved across multiple hypervirulent and clinically relevant C. difficile ribotypes, supporting its broad applicability as a vaccine antigen.
Immunization studies showed that RBD2 induces strong systemic and mucosal antibody responses, unlike the adjacent RBD1 domain. Importantly, RBD2 vaccination provided complete protection in mice against lethal CDT challenge and significant protection in hamsters against spore‑mediated C. difficile infection, a clinically relevant disease model. Functional assays further confirmed that anti‑RBD2 sera neutralize CDT activity by inhibiting toxin‑mediated cell rounding, validating a clear mechanism of action.
Figure: RBD2 Immunization Protects Hamsters from C. difficile Infection
Golden Syrian hamsters were immunized intramuscularly with RBD2 or PBS control and subsequently challenged with C. difficile DSM101085 spores. RBD2‑immunized animals demonstrated significantly improved survival and reduced disease severity, including decreased weight loss, compared to controls. These results show that RBD2 vaccination confers robust protection in a clinically relevant infection