Comparison of toxinotyping and PCR ribotyping of Clostridium difficile strains and description of novel toxinotypes

Toxinotyping and PCR ribotyping are two methods that have been used to type Clostridium difficile isolates. Toxinotyping is based on PCR-RFLP analysis of a 19 kb region encompassing the C. difficile pathogenicity locus. PCR r ibotyping is based on comparison of patterns of PCR products of the 165-23S rRNA intergenic spacer region. Representative strains (101) from a C. diff icile PCR ribotype library and 22 strains from previously described toxinot ypes were analysed to compare ribotyping with toxinotyping. Within this pan el of strains all 11 toxinotypes (0-X) described previously and an addition al 5 novel toxinotypes (XI-XV) were observed. PCR ribotyping and toxinotypi ng correlated well and usually all strains within a given ribotype had simi lar changes in toxin genes. The new toxinotype XI comprises strains that di d not express toxins TcdA or TcdB at detectable levels, but contained part of the tcdA gene. Strains of toxinotype XII exhibit changes only in the 5' end of the tcdB gene. Toxinotype XIV is composed of strains that have a lar ge insertion at the beginning of the tcdA gene. A total of 25 of the 89 tes ted PCR ribotypes of C. difficile contained variant strains. It was estimat ed that they represent 7.7% of the total number of strains in the Anaerobe Reference Unit collection.