Prevalence of Clostridium sPP. in diarrhoeic and healthy dogs ...

Ann. Fac. Medic. Vet. di Parma (Vol. XXVII, 2007) pag. 143 - pag. 156different (P= 0.023, Fisher’s test). This is in disagreement with previous reports inwhich significant differences were not found in the isolation rates of C. difficilebetween the 2 groups [3]. Four of ten C. difficile-positive dogs expressed enteritisafter antibiotic therapy. Probably, antibiotic administration caused the overgrowthof C. difficile in intestine of the dogs, predisposing the animals to enteritis. Infact, a previous study has reported that the carriage rate of C. difficile and toxinsin animals including dogs that had recently received antibiotics was higher thanin the nonantibiotic-treated group, although these differences were not statisticallysignificant [3]. Another study has indicated that the presence of C. difficile in caninefeces was evident only after administration of antibiotic [9]. However, results ofseveral studies have supported the notion that antimicrobial treatment is not requiredfor C. difficile-associated disease (CDAD) development in dogs [3].Since we did not research the toxins A and B (neither in vivo directly in fecesnor in vitro from the C. difficile isolates), which seem the primary virulence factorsinvolved in the pathogenesis of CDAD, we cannot correlate the C. difficile isolationwith dog illness. However, we performed on the 10 C. difficile isolates a duplexPCR for the revelation of tcdA and tcdB genes, encoding for toxin A and toxin B,respectively [11]. The majority of C. difficile strains (6/10, 60.0%) were toxigenic(tcdA+/tcdB+) on the basis of the results of PCR assay (data not shown). The carriagerates of toxigenic isolates in diarrhoeic dogs (5/6, 83.3%) were higher than thosein non-diarrhoeic dogs (1/6, 16.7%). However, this difference was not statisticallysignificant (P= 0.08, Fisher’s test).It could be interesting to verify if tcdA/tcdB-positive strains by PCR willbe able to produce the two toxins also in vitro, because the presence of genes is notalways correlated with their expression.ConclusionsIt is particularly important to seek C. perfringens and C. difficile presencein canine fecal samples. However, C. perfringens-associated diarrhoea in the dogseems to be a complex disease, the pathogenesis of which is still poorly understood.Furthermore, despite various reports that have documented an association betweenthe detection of CPE and the presence of diarrhoea, it is still unclear whether or notC. perfringens is a primary cause of canine diarrhoea or a secondary determinant.Much work still needs to be done to clarify the exact role of C. perfringens incanine diarrhoea. Based on the results of several studies, the optimum diagnosticapproach for canine C. perfringens-associated diarrhoea is the use of a CPE ELISAin conjunction with PCR for detection of enterotoxigenic strains procured aftera heat shock or alcohol shock treatment [7]. To date, different PCRs (duplex andmultiplex) were evaluated for the revelation of genes encoding for C. perfringenstoxins (CPE, toxin α, toxin β, toxin β2, toxin ι and toxin ε) only for research purposes[2, 4]. It will be useful to evaluate these methods on our isolates for the presence ofC. perfringens toxin genes, especially of the CPE gene. The production in vivo and/orin vitro of CPE should be also investigated to correlate the C. perfringens isolationwith diarrhoea in dogs.153