Clostridium difficile is the most common cause of antibiotic-associated diarrhoea worldwide. Over the past 10 years, the incidence and severity of disease have increased in North America and Europe due to the emergence of a hypervirulent clone designated PCR ribotype 027. In this study, we sought to identify phenotypic differences among a collection of 26 presumed PCR ribotype 027 strains from the US and the UK isolated between 1988 and 2008 and also re-evaluated the PCR ribotype. We demonstrated that some of the strains typed as BI by restriction endonuclease analysis, and presumed to be PCR ribotype 027, were in fact other PCR ribotypes such as 176, 198 and 244 due to slight variation in banding pattern compared to the 027 strains. The reassigned 176, 198 and 244 ribotype strains were isolated in the US between 2001 and 2004 and appeared to have evolved recently from the 027 lineage. In addition, the UK strains were more motile and more resistant to most of the antibiotics compared to the US counterparts. We conclude that there should be a heightened awareness of newly identified PCR ribotypes such as 176, 198 and 244, and that they may be as problematic as the notorious 027 strains.