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Housekeeping genes encoding metabolic enzymes may provide alternative markers to 16S ribosomal DNA (rDNA) for genotypic and phylogenetic characterization of bacterial species. We have developed a PCR-restriction fragment length polymorphism (PCR-RFLP) assay, targeting the triosephosphate isomerase ( tpi) gene, which allows the differentiation of twelve pathogenic Clostridium species. Degenerate primers constructed from alignments of tpi sequences of various Gram-positive bacteria allowed the amplification of a 501 bp target region in the twelve Clostridium type strains. A phylogenetic tree constructed from the nucleotidic sequences of these tpi amplicons was well correlated with that inferred from analysis of 16S rDNA gene sequences. The analysis of tpi sequences revealed restriction sites of enzyme Alu I that could be species-specific. Indeed, Alu I digestion of amplicons from the twelve type strains provided distinct restriction patterns. A total of 127 strains (three to sixteen strains for each species) was further analyzed by PCR-RFLP of the tpi gene, and confirmed that each species could be characterized by one to three restriction types (RTs). The differences between RTs within species could be explained by point mutations in Alu I restriction sites of the tpi sequences. PCR-restriction analysis of the tpi gene offers an accurate tool for species identification within the genus Clostridium, and provides an alternative marker to 16S rDNA for phylogenetic analyses.

Clostridium sordellii is an anaerobic gram-positive spore-forming bacillus that is microaerotolerant; it can be isolated from environmental sources and, occasionally, from normal human or animal feces. In rare cases, this histotoxic Clostridium species has been reported as the cause of such human infections as wound or surgical infections, pleuropulmonary infections, postpartum maternal infections, spontaneous endometritis, and necrotizing omphalitis in neonates. The fatal outcome observed in association with some of these clinical features was most often due to shock that was presumably induced by toxin. Infection is an important determinant of morbidity and mortality following orthotopic liver transplantation. We report, to the best of our knowledge, the first case of septic shock and bacteremia due to C. sordellii that occurred following liver biopsy in a liver transplant recipient who had an episode of acute rejection.