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Abstract.Intestinal colonization with toxigenic strains of Clostridium difficile was documented in 9 of 10 horses with acute onset diarrhea in a veterinary medical teaching hospital, whereas a similar isolate was detected in only 1 of 23 other horses without diarrhea in the hospital. One horse with diarrhea was infected simultaneously with both C. difficile and Salmonella krefeld. Clostridium difficile was detected by fecal culture on selective medium, confirmed with a latex particle agglutination test, and identified as toxigenic by polymerase chain reaction amplification of toxin A and toxin B gene sequences. Using an arbitrarily-primed polymerase chain reaction, 6 distinct C. difficile isolates were detected in the feces of the 9 affected horses at the time of the outbreak of diarrhea.Although species of the genus Clostridium, particMaterials and methods ularly C. perfringens, are known causes for enterocolitis in neonatal foals, the role of C. difficile as an enteroAnimals. In the spring of 1993, 10 adult horses developed pathogen in the adult horse is largely unknown. Intesdiarrhea in a 2-day period in the large animal clinic of the tinal colonization with C. difficile has been described University of California Veterinary Medical Teaching Hospital. The affected horses had been hospitalized for periods in the horse, 12 and there are reports of C. difficile in-of time ranging from 0 to 22 days, and 7 of the 10 horses fection in neonatal foals with associated necrotizing enterocolitis 7-9 and in an adult horse with typhlocolitis. 15 Despite the relative paucity of information regarding intestinal colonization of horses with C. difficile, the organism is a common nosocomial pathogen in human patients and causes pseudomembranous colitis associated with antibiotic therapy. 10,18 Several studies have confirmed the presence of C. difficile in animals in veterinary practices and teaching hospitals, although those reports were limited to examinations of small animals. 2,3,13,22 We recently dehad been in the hospital for 4 days or longer. All of the horses were receiving 1 or more antibiotics (ampicillin, 6 horses; gentamicin, 8 horses; ceftiofur, 1 horse; trimethoprim-sulfamethoxazole, 1 horse), and all but 2 of the horses were hospitalized for colic or diarrhea. Four of these horses had had celiotomies during their treatment for colic. Of the two horses without colic, one had uterine torsion and another was undergoing treatment for a subepiglottic mass. Affected horses were located in 4 separated areas (different buildings) of the hospital at the time of fecal sampling. There were 23 other horses without diarrhea in the hospital at this time.Laboratory methods. Specimens of the diarrheal feces were tected C. difficile in small animal patients in our teaching hospital, and although both toxigenic and nontoxigenic isolates were detected, the association between intestinal colonization and clinical signs was not clear. 20In the present study, we present evidence of colonization of adult horses with toxigenic strains...

The genes for Clostridium difficile toxins A and B (tcdA and tcdB) are part of a 19.6-kb pathogenicity locus (PaLoc) that includes the genes tcdD, tcdE, and tcdC. To determine whether the C. difficile PaLoc is a stable and conserved genetic unit in toxigenic strains, a multiplex polymerase chain reaction was used to analyze 50 toxigenic, 39 nontoxigenic, and 2 toxin-defective isolates. The respective amplicons were identified for tcdA-E in the toxigenic isolates; these were absent in the nontoxigenic isolates. C. difficile P-829 lacked at least a fragment of tcdD, tcdB, tcdE, and tcdC, but tcdA was present. C. difficile 8864 had deletions in the tcdA and tcdC genes. These data suggest that the PaLoc is highly stable in toxigenic C. difficile, nontoxigenic isolates lack the unit, and isolates with a defective PaLoc can still cause clinical disease. Further studies are needed to define the role of individual genes in the pathogenesis of C. difficile-associated diarrhea.

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Clostridium difficile is the etiological agent of antibiotic-associated colitis and the most common cause of hospital-acquired infectious diarrhea. Fluoroquinolones such as ciprofloxacin are associated with lower risks of C. difficile-associated diarrhea. In this study, we have analyzed 72 C. difficile isolates obtained from patients with different clinical courses of disease, such as toxic megacolon and relapses; the hospital environment; public places; and horses. They were investigated for their susceptibilities to moxifloxacin (MXF), metronidazole (MEO), and vancomycin (VAN). Mutants highly resistant to fluoroquinolones were selected in vitro by stepwise exposure to increasing concentrations of MXF. The resulting mutants were analyzed for the presence of mutations in the quinolone resistance-determining regions of DNA gyrase (gyrA), the production of toxins A and B, and the epidemiological relationship of these isolates. These factors were also investigated using PCR-based methods. All strains tested were susceptible to MEO and VAN. Twenty-six percent of the clinical isolates (19 of 72) were highly resistant to MXF (MIC > 16 g/ml). Fourteen of these 19 strains contained nucleotide changes resulting in amino acid substitutions at position 83 in the gyrA protein. Resistant strains selected in vitro did not contain mutations at that position. These findings indicate that resistance to MXF in a majority of cases may be due to amino acid substitution in the gyrA gene.