Genotypic and phenotypic characterization of Clostridium perfringens and Clostridium difficile in diarrheic and healthy dogs

The objectives of this study were to examine the potential roles of Clostridium difficile and enterotoxigenic Clostridium perfringens in diarrhea in dogs by comparison of isolation, determination of toxin status via enzyme-linked immunosorbent assay (ELISA), and application of multiplex polymerase chain reaction (PCR). These techniques were used to evaluate fecal specimens in 132 healthy and diarrheic dogs. These dogs were prospectively evaluated by grouping them into the following 3 categories: hospitalized dogs with diarrhea (n = 32), hospitalized dogs without diarrhea (n = 42), and apparently healthy outpatient dogs without diarrhea (n = 58). All fecal specimens were cultured using selective media for C difficile, Salmonella spp., and Campylobacter spp. and selective media after heat shock for C perfringens. No significant difference was found in the isolation of C perfringens or C difficile among the 3 groups. A significant association was found between the presence of diarrhea and detection of C perfringens enterotoxin (CPE) or toxin A via ELISA for both C perfringens and C difficile, respectively. PCR performed on C difficile isolates for toxin A and toxin B genes revealed no significant differences among the 3 groups, but diarrheic dogs were significantly more likely to be positive for the enterotoxin gene of C perfringens. Based on the results of this study, the use of ELISA for detection of CPE in feces combined with the detection of enterotoxigenic fecal isolates obtained via heat shock provides the strongest evidence for the presence of C perfringens-associated diarrhea.     

Evaluation of a routine diagnostic fecal panel for dogs with diarrhea

OBJECTIVES: To assess the diagnostic yield of a routine fecal panel and determine whether Clostridium perfringens or C difficile toxin production is associated with acute hemorrhagic diarrheal syndrome (AHDS) in dogs. DESIGN: Case-control study. ANIMALS: 260 dogs with diarrhea and 177 dogs with normal feces. PROCEDURE: Medical records were reviewed for results of culture for C difficile, Campylobacterspp, and Salmonella spp; C perfringens fecal enterotoxin (CPE) assay via ELISA or reverse passive latex agglutination (RPLA) assay; fecal endospore enumeration; C difficile toxin A assay; and parasite evaluation. RESULTS: Prevalence of CPE in dogs with diarrhea was 22/154 (14.3%) via ELISA and 47/104 (45.2%) via RPLA assay, versus 9/74 (12%) via ELISA and 26/103 (25%) via RPLA assay in control dogs. Prevalence of C difficile was 47/260 (18%) in dogs with diarrhea and 41/74 (55%) in control dogs. Prevalence of C difficile toxin A was 26/254 (10.2%) in dogs with diarrhea and 0/74 in control dogs. Diagnosis of AHDS was made in 27 dogs; 8 had positive results for CPE, 7 had positive results for toxin A, and 1 had positive results for both toxins. Campylobacter spp were isolated from 13 of 260 (5%) dogs with diarrhea and 21 of 74 (28.4%) control dogs. Salmonella spp were isolated from 3 (1.2%) dogs with diarrhea. CONCLUSIONS AND CLINICAL RELEVANCE: Diagnostic value of a fecal panel in dogs with diarrhea appears to below.     

Bacteriologic examination of equine fecal flora as a diagnostic tool for equine intestinal clostridiosis

The fecal flora of 56 clinically healthy and 23 sick horses were examined bacteriologically for counts of Clostridium perfringens, molds, coliforms, alpha- and beta-hemolytic streptococci, and microbes belonging to genus Bacillus, as well as for the presence of Salmonella spp. Of the healthy horses, 85.7% had a C perfringens count less than 10(1) colony-forming units/g of feces. Of the healthy horses, lowest counts were found in race-horses. Of the sick horses, equine intestinal clostridiosis was diagnosed in 2 horses with large C perfringens counts (10(4) to 10(7) colony-forming units/g) and with acute diarrhea. The 7 isolates of C perfringens were identified as serotype A. Salmonella spp were not detected from any of the horses. The study indicated that diagnosing equine intestinal clostridiosis based on the determination of the fecal C perfringens count was suitable.     

Evaluation of methods to diagnose Clostridium perfringens-associated diarrhea in dogs

OBJECTIVE: To assess the prevalence of Clostridium perfringens enterotoxin in feces of dogs with and without diarrhea, and to compare the use of microbial cultures from fecal specimens and evaluation of stained fecal smears for endospores with the presence of enterotoxin as tools for diagnosing C perfringens-associated diarrhea. DESIGN: Prospective study. ANIMALS: 144 dogs representing hospitalized dogs with (n = 41) or without (50) diarrhea, and clinically normal dogs treated as outpatients (53). PROCEDURE: Fresh fecal specimens from all dogs were examined as Gram-stained fecal smears to determine numbers of Gram-positive spore-forming rods/100x objective field. Enterotoxin was assayed directly by use of a reverse passive latex agglutination assay. Fecal specimens were plated directly to prereduced egg yolk agar plates and incubated overnight at 37 C in an anaerobic chamber. At 24 hours, up to 3 lecithinase-positive colonies were subcultured to Brucella blood agar to evaluate for double zone hemolysis. Colonies with double zone hemolysis were tested for aerotolerance and Gram-stained. RESULTS: A significant difference was not detected among groups with respect to the presence of C perfringens as determined by culture, the presence of endospores, and the reaction patterns of fecal enterotoxin assays. An association was not found between number of endospores and the presence of fecal enterotoxin. CLINICAL IMPLICATIONS: The presence of C perfringens enterotoxin in feces of dogs, as detected by the latex agglutination assay used in this study, correlates poorly with the number of fecal endospores, regardless of the dog's clinical status.     

Prevalence of Diarrhea and Enteropathogens in Racing Sled Dogs

Background: Diarrhea is highly prevalent in racing sled dogs, although the underlying causes are poorly understood.
Hypothesis: Clostridium perfringens enterotoxin (CPE) and Clostridium difficile Toxin A and B are associated with diarrhea in racing sled dogs.
Animals: One hundred and thirty-five sled dogs.
Methods: Freshly voided feces were obtained from 55 dogs before racing and from 80 dogs after 400 miles of racing. Samples were visually scored for diarrhea, mucus, blood, and melena. CPE and C. difficile Toxin A and B were detected by ELISA. Samples were cultured for C. perfringens, C. difficile, Campylobacter, Salmonella , and Escherichia coli 0157; Giardia and Cryptosporidium spp. were detected via immunofluorescence.
Results: Diarrhea occurred in 36% of dogs during racing, and hematochezia, fecal mucus or melena, or all 3 occurred in 57.5% of dogs. Salmonella was isolated from 78.2% of dogs before racing, and from 71.3% of dogs during racing. C. perfringens and C. difficile were isolated from 100 and 58.2% of dogs before racing, and from 95 and 36.3% of dogs during racing. Dogs were more likely to test positive for CPE during than before racing (18.8 versus 5.5%, P = .021); however, no enteropathogens or their respective toxins were significantly associated with hematochezia or diarrhea.
Conclusions and Clinical Importance: Sled dogs participating in long distance racing have a high prevalence of diarrhea and hematochezia that is not associated with common enteropathogens. It is possible that diarrhea and hematochezia represent the effect of prolonged exercise on the gastrointestinal tract.     

The roles of Clostridium difficile and enterotoxigenic Clostridium perfringens in diarrhea in dogs

In this prospective study, feces of dogs with diarrhea were compared with feces of normal dogs for the presence of Clostridium difficile, C difficile toxins A and B, C perfringens, and C perfingens enterotoxin (CPE). C difficile toxins A, B, or both were present in feces of 18 of 87 (21%) dogs with diarrhea and 4 of 55 (7%) normal dogs (P = 0.03), whereas CPE was present in the feces of 24 of 87 (28%) dogs with diarrhea and 3 of 55 (5%) normal dogs (P = 0.01). C difficile was isolated from 2 of 87 (2%) dogs with diarrhea but was not isolated from the feces of 55 normal dogs, possibly because of poor survival of the organism in fecal samples. C perfringens was isolated from the feces of 23 of 24 (96%) CPE-positive dogs with diarrhea, 52 of 63 (83%) CPE-negative dogs with diarrhea, and 39 of 55 (71%) CPE-negative dogs with normal feces. No correlation was found between C perfringens spore number and the presence of CPE.     

Enteropathogenic bacteria in dogs and cats: diagnosis, epidemiology, treatment, and control

This report offers a consensus opinion on the diagnosis, epidemiology, treatment, and control of the primary enteropathogenic bacteria in dogs and cats, with an emphasis on Clostridium difficile, Clostridium perfringens, Campylobacter spp., Salmonella spp., and Escherichia coli associated with granulomatous colitis in Boxers. Veterinarians are challenged when attempting to diagnose animals with suspected bacterial-associated diarrhea because well-scrutinized practice guidelines that provide objective recommendations for implementing fecal testing are lacking. This problem is compounded by similar isolation rates for putative bacterial enteropathogens in animals with and without diarrhea, and by the lack of consensus among veterinary diagnostic laboratories as to which diagnostic assays should be utilized. Most bacterial enteropathogens are associated with self-limiting diarrhea, and injudicious administration of antimicrobials could be more harmful than beneficial. Salmonella and Campylobacter are well-documented zoonoses, but antimicrobial administration is not routinely advocated in uncomplicated cases and supportive therapy is recommended. Basic practices of isolation, use of appropriate protective equipment, and proper cleaning and disinfection are the mainstays of control. Handwashing with soap and water is preferred over use of alcohol-based hand sanitizers because spores of C. difficile and C. perfringens are alcohol-resistant, but susceptible to bleach (1:10 to 1:20 dilution of regular household bleach) and accelerated hydrogen peroxide. The implementation of practice guidelines in combination with the integration of validated molecular-based testing and conventional testing is pivotal if we are to optimize the identification and management of enteropathogenic bacteria in dogs and cats.     

Tylosin-responsive chronic diarrhea in dogs

Fourteen dogs had shown chronic or intermittent diarrhea for more than 1 year. Diarrhea had been successfully treated with tylosin for at least 6 months but recurred when treatment was withdrawn on at least 2 occasions. Tylosin-responsive diarrhea (TRD) affects typically middle-aged, large-breed dogs and clinical signs indicate that TRD affects both the small and large intestine. Treatment with tylosin eliminated diarrhea in all dogs within 3 days and in most dogs within 24 hours. Tylosin administration controlled diarrhea in all dogs, but after it was discontinued, diarrhea reappeared in 12 (85.7%) of 14 dogs within 30 days. Prednisone given for 3 days did not completely resolve diarrhea. Probiotic Lactobacillus rhamnosus GG did not prevent the relapse of diarrhea in any of 9 dogs so treated. The etiology of TRD, a likely form of antibiotic-responsive diarrhea (ARD) is unclear. The following reasons for chronic diarrhea were excluded or found to be unlikely: parasites, exocrine pancreatic insufficiency, inflammatory bowel disease, small intestinal bacterial overgrowth, enteropathogenic bacteria (Salmonella spp., Campylobacter spp., Yersinia spp., or Lawsoni intracellularis), and Clostridium perfringens enterotoxin and Clostridium difficile A toxin. A possible etiologic factor is a specific enteropathogenic organism that is a common resident in the canine gastrointestinal tract and is sensitive to tylosin but difficult to eradicate. Additional studies are required to identify the specific cause of TRD.     

Infectious Agents Detected in the Feces of Diarrheic Foals: A Retrospective Study of 233 Cases (2003–2008)

Background: Diarrhea is common in foals but there are no studies investigating the relative prevalence of common infectious agents in a population of hospitalized diarrheic foals.
Objectives: To determine the frequency of detection of infectious agents in a population of hospitalized foals with diarrhea and to determine if detection of specific pathogens is associated with age, outcome, or clinicopathologic data.
Animals: Two hundred and thirty-three foals ≤ 10 months of age with diarrhea examined at a referral institution.
Methods: Retrospective case series. Each foal was examined for Salmonella spp., viruses, Clostridium difficile toxins, Clostridium perfringens culture, C. perfringens enterotoxin, Cryptosporidium spp., and metazoan parasites in feces collected at admission or at the onset of diarrhea.
Results: At least 1 infectious agent was detected in 122 foals (55%). Rotavirus was most frequently detected (20%) followed by C. perfringens (18%), Salmonella spp. (12%), and C. difficile (5%). Foals < 1 month of age were significantly more likely to be positive for C. perfringens (odds ratio [OR] = 15, 95% confidence interval [CI] = 3.5–66) or to have negative fecal diagnostic results (OR = 3.0, 95% CI = 1.7–5.2) than older foals. Foals > 1 month of age were significantly more likely to have Salmonella spp. (OR = 2.6, 95% CI = 1.2–6.0), rotavirus (OR = 13.3, 95% CI = 5.3–33), and parasites (OR = 23, 95% CI = 3.1–185) detected compared with younger foals. Overall 191 of the 223 foals (87%) survived. The type of infectious agent identified in the feces or bacteremia was not significantly associated with survival.
Conclusions and Clinical Importance: In the population studied, foals with diarrhea had a good prognosis regardless of which infectious agent was identified in the feces.     

Use of bacitracin in the prevention and treatment of experimentally-induced idiopathic colitis in horses.

Ten healthy ponies from a single herd were found by repeated fecal culture to be free of Salmonella species and Clostridium cadaveris. In a preliminary study, four ponies administered a single oral dose of 10 mg/kg lincomycin did not develop idiopathic colitis when the drug was administered alone. Four other ponies were administered 10 mg/kg lincomycin by stomach tube together with 0.45 L of colonic content from a horse with idiopathic colitis induced earlier by lincomycin alone. Two of the four ponies were treated with 25 g oral zinc bacitracin premix (110 g/kg active ingredient) 24 h later. Forty-two hours after inoculation the two untreated ponies had severe signs of idiopathic colitis and were euthanized. Postmortem findings were typical of idiopathic colitis. The two treated ponies had milder illness but the more severely affected was also euthanized; the other was retreated at 42 h with bacitracin pre-mix and again 12 h later. Its illness and diarrhea resolved over the next 24 h. Clostridium cadaveris was isolated in large numbers from the cecum of the euthanized ponies and their cecal content contained mouse lethal and guinea pig dermonecrotic, but not cytotoxic, activity. Enterotoxins of Clostridium perfringens and Clostridium difficile could not be demonstrated. No toxin could be demonstrated in culture supernatants of C. cadaveris or in supernatants of cecal contents treated with ethanol prior to culturing in anaerobically incubated broth. No Salmonella spp. were isolated. A further two ponies were administered 10 mg/kg lincomycin orally with 0.45 L colonic content from a horse with idiopathic colitis, as described.(ABSTRACT TRUNCATED AT 250 WORDS)     

Occurrence of anaerobic bacteria in diseases of the dog and cat.

A survey for anaerobic bacteria was conducted in 314 clinical specimens from dogs and cats. A total of 187 anaerobic isolates in pure and mixed culture were isolated from 111 of the specimens that contained anaerobic bacteria. Common isolated included Actinomyces (9.1%), Clostridium perfringens (19.3%), other Clostridium spp (11.2%), Peptostreptococcus anaerobius (7.5%), Bacteroides melaninogenicus (13.4%), other Bacteroides spp (17.6%), and Fusobacterium necrophorum (5.3%). Anaerobic bacteria were involved in serious lesions that often were life threatening to the animals. Antibiotic susceptibility data indicated that the lincomycin family, the penicillin family, chloramphenicol, and cephaloridine are preferred drugs for treatment of anaerobic infections. Data from the survey were used in formulation of a table to aid practitioners in clinical diagnosis of disease caused by anaerobes. Clostridium perfringens was isolated in large numbers from five of six dogs with a clinical diagnosis of canine hemorrhagic gastroenteritis and from one cat with hemorrhagic diarrhea. Experimental infections were induced in rats, using caine feces as inoculum. Induced lesions contained aerobic and anaerobic bacteria similar to those bacteria isolated in the clinical survey, indicating that feces may serve as a major source of these bacteria in clinical infections of the dog.     

Antimicrobial susceptibilities of canine Clostridium difficile and Clostridium perfringens isolates to commonly utilized antimicrobial drugs

Clostridium difficile and Clostridium perfringens are anaerobic, Gram-positive bacilli that are common causes of enteritis and enterotoxemias in both domestic animals and humans. Both organisms have been associated with acute and chronic large and small bowel diarrhea, and acute hemorrhagic diarrheal syndrome in the dog. The objective of this study was to determine the in vitro antimicrobial susceptibilities of canine C. difficile and C. perfringens isolates in an effort to optimize antimicrobial therapy for dogs with clostridial-associated diarrhea. The minimum inhibitory concentrations (MIC) of antibiotics recommended for treating C. difficile (metronidazole, vancomycin) and C. perfringens-associated diarrhea in the dog (ampicillin, erythromycin, metronidazole, tetracycline, tylosin) were determined for 70 canine fecal C. difficile isolates and 131 C. perfringens isolates. All C. difficile isolates tested had an MIC of or=256 microg/ml for both erythromycin and tylosin. A third C. perfringens isolate had an MIC of 32 microg/ml for metronidazole. Based on the results of this study, ampicillin, erythromycin, metronidazole, and tylosin appear to be effective antibiotics for the treatment of C. perfringens-associated diarrhea, although resistant strains do exist. However, because there is limited information regarding breakpoints for veterinary anaerobes, and because intestinal concentrations are not known, in vitro results should be interpreted with caution.     

Evaluation of five enzyme immunoassays compared with the cytotoxicity assay for diagnosis of Clostridium difficile-associated diarrhea in dogs.

Clostridium difficile-associated-diarrhea (CDAD) is a nosocomial infection in dogs. Diagnosis of this infection is dependent on clinical signs of disease supported by laboratory detection of C. difficile toxins A or B, or both, in fecal specimens via enzyme-linked immunosorbent assay (ELISA). Unfortunately, to the authors' knowledge, commercially available ELISAs have not been validated in dogs to date. We evaluated 5 ELISAs done on 143 canine fecal specimens (100 diarrheic and 43 nondiarrheic dogs) and on 29 C. difficile isolates. The results of each ELISA were compared with the cytotoxin B tissue culture assay (CTA). Clostridium difficile was isolated from 23% of the fecal specimens. Eighteen of the 143 fecal specimens were toxin positive (15 diarrheic and 3 nondiarrheic dogs). On the basis of multiplex polymerase chain reaction (PCR) analysis for toxin-A and -B genes, 72% of the isolates were toxigenic. The carriage rate of toxigenic isolates in diarrheic dogs was higher than that in the nondiarrheic dogs; however, these differences were not statistically significant. A good correlation was found between CTA, PCR, and culture results. The ELISAs done on fecal specimens collected from diarrheic dogs had low sensitivity (7-33%). In contrast, ELISA for toxin A or B, or both, performed on toxigenic isolates had high sensitivity (93%). These results suggest that commercially available human ELISAs are inadequate for the diagnosis of canine C. difficile-associated diarrhea when tested on fecal specimens. In contrast, the Premier ToxinA/B and Techlab ToxinA/B ELISAs may be useful for the diagnosis of canine CDAD when used on toxigenic isolates.     

Epizootiologic investigations of a diarrheic syndrome in fattening pigs

A diarrheic syndrome linked to Clostridium perfringens was observed in fattening pigs. A good correlation was observed between the onset and the severity of diarrhea and the fecal passage of C perfringens enterotoxin. Intestinal fluids from affected pigs had activity comparable with that detected in a purified C perfringens enterotoxin. Seven pigs excreting enterotoxin were shown to develop serum antibodies to C perfringens enterotoxin.     

Cryptosporidiosis associated with bacterial enteritis in a goat kid

A 7-day-old male Nubian-Alpine crossbred goat was examined because of listlessness, anorexia, and diarrhea. The presumptive diagnosis was severe enteritis. Large numbers of Clostridium perfringens and a non-pathogenic heavily encapsulated Escherichia coli were isolated from the feces. Cryptosporidium parvum was identified on the qualitative fecal examination. The kid improved after treatment with fluids and antibiotics.     

Enteric Clostridium perfringens infection associated with parvoviral enteritis in dogs: 74 cases (1987-1990).

Parvovirus infection was confirmed by fluorescent antibody staining of or viral isolation from specimens of small intestine in 181 (17%) of 1,110 dogs necropsied between July 1, 1987 and Dec 31, 1990. Clostridium perfringens was isolated from 74 (69%) of 108 dogs with parvovirus infection from which specimens of jejunum also had been obtained for culture of anaerobic bacteria. Gram-positive bacilli in association with focal to diffuse necrosis of the superficial portions of the villi were observed in histologic sections of specimens of small intestine from 56 (98%) of 57 dogs from which parvovirus and C perfringens had been identified. These findings indicate that C perfringens frequently proliferates in dogs with parvovirus infection.     

Recurrent diarrhea associated with enterotoxigenic Clostridium perfringens in 2 dogs

Two dogs were diagnosed with enterotoxigenic Clostridium perfringens-associated diarrhea. Diarrhea was responsive to antimicrobial therapy, but recurred after treatment was ceased. Clostridium perfringens enterotoxin was present in feces during diarrheic episodes but not when feces were normal. Both dogs responded to a prolonged course of oral cephalexin and dietary modification.     

Bacteriological evaluation of commercial canine and feline raw diets

Twenty-five commercial raw diets for dogs and cats were evaluated bacteriologically. Coliforms were present in all diets, ranging from 3.5 x 10(3) to 9.4 x 10(6) CFU/g (mean 8.9 x 10(5); standard deviation 1.9 x 10(6)). Escherichia coli was identified in 15/25 (64%) diets; however, E. coli O157 was not detected. Salmonella spp. were detected in 5/25 (20%) diets; 1 each of beef-, lamb-, quail-, chicken-, and ostrich-based diets. Sporeforming bacteria were identified from 4/25 (16%) samples on direct culture and 25/25 (100%) samples using enrichment culture. Clostridium perfringens was identified in 5/25 (20%) samples. A toxigenic strain of C. difficile was isolated from one turkey-based food. Staphylococcus aureus was isolated from 1/25 (4%) diets. Campylobacter spp. were not isolated from any of the diets.     

Prevalence of Clostridium perfringens enterotoxin and Clostridium difficile toxin A in feces of horses with diarrhea and colic.

OBJECTIVE: To determine prevalence of clostridial enterotoxins in feces of horses with diarrhea and colic, and to determine whether an association exists between detection of clostridial enterotoxins in feces and development of diarrhea as a complication of colic. DESIGN: Prospective case series and case-control study. ANIMALS: 174 horses with diarrhea, colic, or problems not related to the gastrointestinal tract. PROCEDURE: Horses were assigned to 1 of 4 groups: colic with diarrhea (group 1; n = 30); colic without diarrhea (group 2; 30); diarrhea without colic (group 3; 57); and control (group 4; 57). Feces were evaluated by use of ELISA to detect Clostridium perfringens enterotoxin (CPE) and C difficile toxin A (TOXA). Frequency of detection of CPE or TOXA in groups 1 and 3 was compared with that in groups 2 and 4, respectively. RESULTS: Prevalence of enteric clostridiosis in horses in group 3 was 25%. Clostridium perfringens enterotoxin was detected in 9 of 57 (16%), TOXA in 8 of 57 (14%), and both toxins in 3 of 57 (5%) fecal samples collected from these horses. Neither toxin was detected in feces of the age-matched horses in group 4. Clostridial enterotoxins were detected in feces of 7 of 60 (12%) horses with colic (groups 1 and 2), however, a significant association was not found between detection of enterotoxins in feces and development of diarrhea as a complication of colic. CONCLUSIONS AND CLINICAL RELEVANCE: Clostridia are important etiologic agents of diarrhea in horses. Additionally, changes in intestinal flora of horses with colic may allow for proliferation of clostridia and elaboration of enterotoxins regardless of whether diarrhea develops.     

Dietary effects on bifidobacteria and Clostridium perfringens in the canine intestinal tract

Dietary effects on the intestinal microflora have gained increasing interest because of the evidence that a balanced micro ecology in the gut is important for health and well being. The aim of the present study was to evaluate the effect of different diets on faecal counts of bifidobacteria and Clostridium perfringens in dogs. Two extruded, dry diets, one supplemented with 3% chicory (1.5% inulin), a non-digestible oligosaccharide (NDO) and the other with 3% glucose (GLU) were compared with a protein rich diet (PR+) based on low quality animal derived protein sources (NDO 265, GLU 259, PR+ 726 g crude protein/kg dry matter; greaves meal and bovine lung as protein sources in PR+). Nine adult beagles were subjected to a consecutive cross-over trial. All dogs started with diet PR+, after which groups of four dogs (group A) received GLU and the other five dogs (group B) received NDO. After an intermediate wash-out period with diet PR+ for 3 weeks the A dogs were switched to diet NDO and B dogs to GLU. In the final period all dogs were fed with diet PR+. Faecal samples were collected during each period for dry matter and pH measurements. Faecal bifidobacteria and Cl. perfringens were quantified in fresh samples at the end of each feeding period and additionally on the first days after feed change from the dry diets to diet PR+. Diets NDO and GLU increased faecal dry matter and reduced faecal pH from 6.9 to 7.4 with the high protein diet to 5.9-6.5. The dry diets induced a firmer faecal consistency and a lower faecal pH, with no significant difference between NDO or GLU. Clostridium perfringens was found in all faecal specimens after feeding PR+ with counts of log 8.2-8.8 colony forming units (cfu)/g faeces. Both dry diets reduced the counts of Cl. perfringens significantly (log 3.3-4.0 cfu/g faeces). Switching from the dry diets to the high protein diet induced an increase of Cl. perfringens within 1 day, independent of the previous diet. In dogs fed PR+, bifidobacteria were detected in only four faecal samples and exclusively in the initial feeding period. During the remainder of the experiment the counts fell below the detection limit (log 6 cfu/g faeces). The faecal concentrations of bifidobacteria increased with both dry diets. Slightly higher concentrations (log 9.6-9.7 cfu/g faeces) were obtained from dogs fed the dry diet containing NDO compared with the diet containing glucose (log 9.3-9.4 cfu/g faeces). The increase was small which may be related to the level of total fermentable carbohydrates in both diets which alone increase remarkably the total counts of bifidobacteria. In conclusion, distinct dietary effects on the faecal counts of Cl. perfringens and bifidobacteria with a clear antagonistic pattern were observed. The main factor was the protein source and level in the diet. In this case, NDO favoured the concentrations of bifidobacteria to a limited degree. Further studies are needed to evaluate time effects, metabolic consequences and the potential implication for health promotion in pets.