Clostridium difficile and Clostridium perfringens from wild carnivore species in Brazil

Despite some case reports, the importance of Clostridium perfringens and Clostridium difficile for wild carnivores remains unclear. Thus, the objective of this study was to identify C. perfringens and C. difficile strains in stool samples from wild carnivore species in Brazil. A total of 34 stool samples were collected and subjected to C. perfringens and C. difficile isolation. Suggestive colonies of C. perfringens were then analyzed for genes encoding the major C. perfringens toxins (alpha, beta, epsilon and iota) and the beta-2 toxin (cpb2), enterotoxin (cpe) and NetB (netb) genes. C. difficile strains were analyzed by multiplex-PCR for toxins A (tcdA) and B (tcdB) and a binary toxin gene (cdtB) and also submitted to a PCR ribotyping. Unthawed aliquots of samples positive for C. difficile isolation were subjected to the detection of A/B toxins by a cytotoxicity assay (CTA). C. perfringens was isolated from 26 samples (76.5%), all of which were genotyped as type A. The netb gene was not detected, whereas the cpb2 and cpe genes were found in nine and three C. perfringens strains, respectively. C. difficile was isolated from two (5.9%) samples. A non-toxigenic strain was recovered from a non-diarrheic maned wolf (Chrysocyon brachyurus). Conversely, a toxigenic strain was found in the sample of a diarrheic ocelot (Leopardus pardallis); an unthawed stool sample was also positive for A/B toxins by CTA, indicating a diagnosis of C. difficile-associated diarrhea in this animal. The present work suggests that wild carnivore species could carry C. difficile strains and that they could be susceptible to C. difficile infection.