Recent investigations of Clostridium difficile cell wall components have revealed the presence of an S-layer encoded by the slpA gene. The aim of this study was to determine whether slpA genotyping can be used as an alternative to serotyping. The variable regions of slpA were amplified by PCR from serogroup reference strains and various clinical isolates chosen randomly. Amplified products were analyzed after restriction enzyme digestion and DNA sequencing. The sequences of the variable region of the SlpA protein were found to be strictly identical within a given serogroup but divergent between serogroups. These preliminary results suggest that PCR-restriction fragment length polymorphism, in conjunction with DNA sequencing of the slpA variable region, could constitute an alternative typing method for determining C. difficile serotypes.