Clostridium difficile 027/BI/NAP1 Encodes a Hypertoxic and Antigenically Variable Form of TcdB

The Clostridium difficile exotoxin, TcdB, which is a major virulence factor, varies between strains of this pathogen. Herein, we show that TcdB from the epidemic BI/NAP1/027 strain of C. difficile is more lethal, causes more extensive brain hemorrhage, and is antigenically variable from TcdB produced by previously studied strains of this pathogen (TcdB(003)). In mouse intoxication assays, TcdB from a ribotype 027 strain (TcdB(027)) was at least four fold more lethal than TcdB(003). TcdB(027) caused a previously undescribed brain hemorrhage in mice and this correlated with a heightened sensitivity of brain microvascular endothelial cells to the toxin. TcdB(003) and TcdB(027) also differed in their antigenic profiles and did not share cross-neutralizing epitopes in a major immunogenic region of the protein. Solid phase humoral mapping of epitopes in the carboxy-terminal domains (CTD) of TcdB(027) and TcdB(003) identified 11 reactive epitopes that varied between the two forms of TcdB, and 13 epitopes that were shared or overlapping. Despite the epitope differences and absence of neutralizing epitopes in the CTD of TcdB(027), a toxoid form of this toxin primed a strong protective response. These findings indicate TcdB(027) is a more potent toxin than TcdB(003) as measured by lethality assays and pathology, moreover the sequence differences between the two forms of TcdB alter antigenic epitopes and reduce cross-neutralization by antibodies targeting the CTD.