Analysis of Clostridium difficile strains causing multiple nosocomial outbreaks in Japan by genotypic typing methods

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The typing system sequencing the variable region of the slpA gene was applied to typing Clostridium difficile strains belonging to one PCR ribotype, type smz, which has been identified as frequently causing outbreaks in Japan. The PCR ribotype smz strains recovered from patients at different hospitals in Japan were examined. Among 10 type smz strains tested, three subtypes, smz-1,-2,and -3,were identified that differed from each other by one nucleotide. slpA sequence typing was also applied to direct typing on DNA extracted from stool specimens. Of 22 stool specimens examined, 17 were PCR positive for slpA ; eight were typed as slpA sequence type smz-1,and nine were type smz-2. C.difficile was cultured from 12 of the 17 stool specimens, and the sequence results of the recovered isolates were compared with those from DNA extracted from stool specimens. In all 12 stool specimens, the sequence results of DNA from recovered C.difficile isolates completely agreed with those of DNA extracted directly from stool specimens. The remaining five stool specimens were culture-negative for C.difficile. Sequence typing has the advantage of being easy to compare typing results among multiple laboratories via the Internet without exchanging reference strains to those of typing systems the results of which depend on banding-pattern analyses. slpA sequence typing appears to be a reproducible and reliable typing system for C.difficile as well as being useful for the typing of C.difficile when stool specimens contain only small numbers of C.difficile or are inappropriate for culturing.