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The important veterinary pathogenClostridium perfringens type B is unique for producing the two most lethalC .perfringens toxins, i.e., epsilon-toxin and beta-toxin. Our recent study (K. Miyamoto, J. Li, S. Sayeed, S. Akimoto, and B. A. McClane, J. Bacteriol. 190:7178-7188, 2008) showed that most, if not all, type B isolates carry a 65-kb epsilon-toxin-encoding plasmid. However, this epsilon-toxin plasmid did not possess thecpb gene encoding beta-toxin, suggesting that type B isolates carry at least one additional virulence plasmid. Therefore, the current study used Southern blotting of pulsed-field gels to localize thecpb gene to ∼90-kb plasmids in most type B isolates, although a few isolates carried a ∼65-kbcpb plasmid distinct from theiretx plasmid. Overlapping PCR analysis then showed that the gene encoding the recently discovered TpeL toxin is located ∼3 kb downstream of the plasmid-bornecpb gene. As shown earlier for their epsilon-toxin-encoding plasmids, the beta-toxin-encoding plasmids of type B isolates were found to carry atcp locus, suggesting that they are conjugative. Additionally, IS1151 -like sequences were identified upstream of thecpb gene in type B isolates. These IS1151 -like sequences may mobilize thecpb gene based upon detection of possiblecpb- containing circular transposition intermediates. Most type B isolates also possessed a third virulence plasmid that carries genes encoding urease and lambda-toxin. Collectively, these findings suggest that type B isolates are among the most plasmid dependent of allC. perfringens isolates for virulence, as they usually carry three potential virulence plasmids.

Characterization of Virulence Plasmid Diversity among Clostridium perfringens Type B Isolates