Binary toxin locus analysis in Clostridium difficile.

The objective of this study was to compare full binary toxin loci (CDTloc) sequences from a collection of Clostridium difficile isolates in an effort to further understand the regulation of the binary toxin (CdtAB) and its putative regulator (CdtR). Sequences from different ribotypes and toxinotypes were analysed phylogenetically and for polymorphisms, non-sense mutations, promoter features and signal sequences. Expression of cdtA, which was also representative of cdtB expression, was measured by quantitative PCR (qPCR). Several consensus promoter features and various polymorphisms were identified including a non-sense mutation identified in a ribotype 078 cdtR gene that is predicted to result in a severely truncated protein. Despite this mutation, cdtA expression was still detected by qPCR. Dendrograms based on total sequences indicated that isolates belonging to the same ribotype shared the greatest similarity within the binary toxin locus. Although cdtR is thought to be involved in regulation of cdtA expression, a cdtR non-sense mutation did not inhibit expression of cdtA, suggesting that either the truncated protein is functional or another regulator of the binary toxin exists.