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.     

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 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.     

A survey of intestinal parasites in dogs from Saskatoon, Saskatchewan

Fresh fecal samples from 124 apparently healthy dogs and 333 random source canine fecal samples from dog walking areas were analyzed by centrifuged flotation in Sheather's solution to determine the prevalence of gastrointestinal parasites in dogs from Saskatoon, Saskatchewan. Fecal flotation was positive in 4.4% of samples. Roundworm species were found in 1.5% of samples, hookworm species were found in 0.4% of samples, whipworm species were found in 0.7%, Strongyloides spp. were found in 0.6%, Giardia spp., Cystoisospora spp., and Alaria spp. were found in 0.4% of samples.     

Experimentally induced coronavirus infections in calves: viral replication in the respiratory and intestinal tracts

Eleven 3- to 50-day-old colostrum-deprived gnotobiotic calves and seven 25- to 63-day-old colostrum-deprived conventional calves were allotted into 3 groups. Each group was inoculated with a fecal isolate of bovine coronavirus via different routes: orally/intranasally OR/IN, No. 1 through 8, group 1 calves; OR, No. 9 through 13, group 2 calves; IN, No. 14 through 18, group 3 calves. Nasal swab specimens and fecal specimens were collected daily and were examined for coronavirus antigen by use of direct immunofluorescent staining (nasal epithelial cells) or by use of immune electron microscopy (fecal specimens). All but 4 calves (No. 11, 13, 17, and 18) were euthanatized on postinoculation days (PID) 3 to 7. Calves 11 and 17 became severely dehydrated and died at PID 5. Calves 13 and 18 were evaluated for nasal and fecal shedding of coronavirus through PID 14. Distribution of coronavirus antigen in the respiratory and intestinal tracts of the 14 euthanatized calves was evaluated by use of direct immunofluorescent staining. All calves developed profuse diarrhea by PID 2 to 4; however, calves did not develop clinical signs of respiratory tract disease before euthanasia or death. Inoculated calves shed coronavirus in their feces as detected by use of immune electron microscopy. Infected nasal epithelial cells were detected in all but 2 orally inoculated calves (No. 9 and 10). Route of inoculation influenced the sequence of initial detection of coronavirus antigen from fecal specimens or nasal swab specimens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Recent epidemiological status of canine viral enteric infections and Giardia infection in Japan.

Epidemiology of canine enteric infections was studied. Rectal swabs collected from 95 dogs presented at animal hospitals during a period from January to June of 2000 were examined for enteric pathogens, including viruses and Giardia lamblia (G. lamblia). Most frequently detected in both diarrheal and normal feces were canine coronavirus (55.4%) and G. lamblia (48.2%). Canine parvovirus type 2 (CPV-2) was specifically associated with diarrheal cases and CPV-2b was the predominant antigenic type. Although canine rotavirus, canine adenovirus, and canine distemper virus were also detected in a small number of diarrheal cases, no evidence for calicivirus infection was obtained.     

Coronavirus infection of spotted hyenas in the Serengeti ecosystem

Sera from 38 free-ranging spotted hyenas (Crocuta crocuta) in the Serengeti ecosystem, Tanzania, were screened for exposure to coronavirus of antigenic group 1. An immunofluorescence assay indicated high levels of exposure to coronavirus among Serengeti hyenas: 95% when considering sera with titer levels of > or = 1:10 and 74% when considering sera with titer levels of > or = 1:40. Cubs had generally lower mean titer levels than adults. Exposure among Serengeti hyenas to coronavirus was also confirmed by a serum neutralisation assay and an ELISA. Application of RT-PCR to 27 fecal samples revealed viral RNA in three samples (11%). All three positive fecal samples were from the 15 juvenile animals (<24 months of age) sampled, and none from the 12 adults sampled. No viral RNA was detected in tissue samples (lymph node, intestine, lung) from 11 individuals. Sequencing of two amplified products from the S protein gene of a positive sample revealed the presence of coronavirus specific RNA with a sequence homology to canine coronavirus of 76 and 78% and to feline coronavirus type II of 80 and 84%, respectively. Estimation of the phylogenetic relationship among coronavirus isolates indicated considerable divergence of the hyena variant from those in European, American and Japanese domestic cats and dogs. From long-term observations of several hundred known individuals, the only clinical sign in hyenas consistent with those described for coronavirus infections in dogs and cats was diarrhea. There was no evidence that coronavirus infection in hyenas caused clinical signs similar to feline infectious peritonitis in domestic cats or was a direct cause of mortality in hyenas. To our knowledge, this is the first report of coronavirus infection in Hyaenidae.     

The use of enzyme-linked immunosorbent assay systems for serology and antigen detection in parvovirus, coronavirus and rotavirus infections in dogs in The Netherlands.

Complex trapping blocking (CTB) enzyme-linked immunosorbent assays (ELISAs) and indirect ELISAs for the detection of antibodies to canine parvovirus (CPV), canine coronavirus (CCV) and rotavirus in sera of dogs were established. Double antibody sandwich ELISAs for the detection of CPV-, CCV- and rotavirus antigens in fecal samples were also developed. Both the serological and antigen-detection ELISAs were used to screen samples from dogs in The Netherlands, with or without a history of acute diarrhea. It was shown that the results of the respective serological ELISAs correlated well and that CPV was the major cause of virus-induced acute diarrhea in dogs in The Netherlands.     

ELISA法检测犬腹泻粪样中的犬冠状病毒

用ＦＥ细胞增殖犬冠状病毒（ＣＣＶ）参考株，分别免疫家兔和ＢＡＬＢ／ｃ小鼠制备ＣＣＶ多抗和单抗，建立了夹心ＥＬＩＳＡ及Ｄｏｔ－ＥＬＩＳＡ诊断方法。在检测的８４例犬腹泻粪样中，多抗、单抗夹心法显示ＣＣＶ阳性１６例，Ｄｏｔ－ＥＬＩＳＡ阳性１３例，后１３例包括在前１６例中。从８４例腹泻犬粪样中随机取３８例作ＣＣＶ、犬细小病毒（ＣＰＶ）双项检测，ＣＣＶ阳性１６例，ＣＰＶ阳性６例，ＣＣＶ、ＣＰＶ混合感染４例。结果显示，在南京地区流行的犬腹泻中，ＣＣＶ感染比例有超过ＣＰＶ的趋势。     

Comparison of direct immunofluorescence, immunoassays, and fecal flotation for detection of Cryptosporidium spp. and Giardia spp. in naturally exposed cats in 4 Northern California animal shelters

BACKGROUND: Giardia spp. and Cryptosporidium spp. are common intestinal protozoan parasites in domestic cats. Few studies have critically evaluated the performance characteristics of commercially available immunoassays for detection of these organisms in the cat. HYPOTHESIS: Human-based immunoassays are suboptimal for the detection of Giardia spp. and Cryptosporidium spp. in cats. ANIMALS: Three-hundred-and-forty-four cats with diarrheic and nondiarrheic fecal specimens at 4 northern California animal shelters. METHODS: A fecal specimen was collected from each cat in a case-controlled fashion. Fecal specimens were tested for Giardia spp. and Cryptosporidium spp. by using centrifugation flotation and 5 commercially available immunoassays (SNAP Giardia, ProSpecT Giardia Microplate Assay, ProSpecT Cryptosporidium Microplate Assay, ImmunoCard STAT! Cryptosporidium/ Giardia Rapid Assay, and Xpect Giardia/Cryptosporidium). Results were compared with a reference standard, the MeriFluor direct immunofluorescence assay. RESULTS: Overall prevalences of Giardia spp. and Cryptosporidium spp. were 9.8 and 4.7%, respectively. The ProSpecT Microplate Assay had the highest sensitivities and specificities for Giardia spp. (91.2 and 99.4%) and Cryptosporidum spp. (71.4 and 96.7%), respectively. The SNAP Giardia antigen assay was easier to use and equally sensitive (85.3%) and specific (100%) to fecal flotation. CONCLUSIONS AND CLINICAL IMPORTANCE: Caution should be exercised when using human-based immunoassays for the diagnosis of Giardia and Cryptosporidium spp. in cats. Fecal flotation remains a useful method for detection of Giardia spp., can be used to detect other parasites, and has a sensitivity of 97.8% for detection of Giardia spp. when combined with the SNAP Giardia immunoassay.     

Potential pathogens in feces from unweaned llamas and alpacas with diarrhea

OBJECTIVE: To identify potential pathogens in feces from llama and alpaca crias with diarrhea. DESIGN: Prospective observational study. ANIMALS: 45 unweaned crias with diarrhea. PROCEDURE: Fecal samples were evaluated for Eimeria spp, Giardia spp, Cryptosporidium spp, enteric viruses, and Salmonella spp. A questionnaire yielded information concerning herd management and presence of other affected camelids. RESULTS: 28 crias were < or = 31 days old, 11 were 32 to 62 days old, and 6 were 63 to 210 days old. Potential pathogens were isolated from feces from 32 of the 45 crias. A total of 39 potential pathogens were obtained, including coronavirus (n = 19 crias; 42%), Giardia spp (8; 18%), Eimeria spp (6; 13%), Cryptosporidium spp (4; 9%), rotavirus (1; 2%), and nematode ova (1; 2%). Salmonella spp were not isolated. Most crias from which potential pathogens were isolated were identified during outbreaks of diarrhea involving other camelids, although only coronavirus was isolated from crias identified during outbreaks involving adult camelids. Coronavirus was detected throughout the year, whereas protozoa were most commonly isolated during the fall and winter. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that a variety of potential pathogens may be isolated from young crias with diarrhea. Many crias shed coronavirus, which may also have been affecting older camelids. Protozoa were isolated most often during wetter months, suggesting that crias born during these months may have greater exposure to protozoal pathogens.     

Association of diarrhea in cattle with torovirus infections on farms.

An epidemiologic survey was performed to determine the incidence of torovirus infections in 2 disease entities of cattle: diarrhea of replacement calves up to 2 months old, and winter dysentery of adult cattle. Samples were obtained from 187 diarrheal and 115 healthy calves from 15 farms, as well as 149 diarrheal and 67 healthy cows from 27 farms with or without winter dysentery. Enzyme-linked immunosorbent assays for detection of torovirus, rotavirus, and coronavirus antigen in feces, and of torovirus and coronavirus antibodies in serum were used to monitor infections in these groups. Torovirus was detected in 9 of the 15 farms in the study, and in 6% of calves with diarrhea, which was significantly higher than in healthy calves (2%). Seroconversion to torovirus was found significantly more often after winter dysentery episodes than on farms without a disease history; coronavirus seroconversion was less common.     

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.     

Salmon Poisoning Disease in Dogs: 29 Cases

Background: Salmon poisoning disease (SPD) is a trematode‐borne disease of dogs caused by Neorickettsia helminthoeca. Objectives: To determine risk factors and spatial epidemiology of SPD in dogs from northern California; to describe the clinicopathologic, microbiologic, and imaging findings of SPD in these dogs; and to evaluate treatments and outcomes for SPD. Animals: Twenty‐nine dogs with SPD based on the finding of trematode ova in the feces, or organisms consistent with N. helminthoeca in specimens submitted for microscopic examination. Methods: Information regarding signalment, fish exposure, clinical signs, diagnostic evaluation, treatments, and outcomes was obtained for each dog. Archived lymph node aspirates and histopathology specimens were subjected to polymerase chain reaction (PCR) testing for Neorickettsia spp. Results: Labrador Retrievers and intact male dogs were overrepresented. Exposure locations were often distant from the dogs' residence. Some dogs had neurologic signs, including twitching and seizures. Dogs lacking peripheral lymphadenomegaly had abdominal lymphadenomegaly on ultrasound examination. A combination of centrifugation fecal flotation and sedimentation had greatest sensitivity for finding fluke ova. N. helminthoeca DNA was amplified by PCR from 4/10 dogs. Penicillins, cephalosporins, and chloramphenicol did not appear to be effective treatments. Mortality rate was 4/29 (14%). Conclusions and Clinical Importance: SPD should be suspected in dogs with inappetence, gastrointestinal, or neurologic signs, with or without fever or peripheral lymphadenomegaly in the appropriate geographical setting. Diagnosis is facilitated by a combination of fecal sedimentation and centrifugal flotation, abdominal ultrasonography, and PCR‐based assays on lymphoid tissue. The treatment of choice is tetracycline antimicrobials.     

Viral antibody studies of laboratory dogs with diarrheal disease

Viral antibody studies were done on laboratory dogs in an epizootic of gastrointestinal disease. Increased hemagglutination-inhibition antibody titers to a parvovirus (PV) antigenically related to feline panleukopenia virus were found in convalescent serum specimens of 78% (20/26) of the affected dogs and in 83% (5/6) of apparently healthy dogs. With one exception, all dogs tested had significant levels of hemagglutination-inhibition antibody to this PV. Similar increased antibody titers were found to feline panleukopenia virus. Also, neutralizing antibody responses were detected to the canine coronavirus in 24% (6/25) and canine herpesvirus in 45% (10/22) of the affected dogs. However, antibody titers did not increase to canine distemper virus, infectious canine hepatitis virus, canine parainfluenza virus, or minute virus of canines. Subsequent serotesting of the colony provided evidence that additional PV infections occurred in pups from each of 8 litters born 3 to 8 months after the epizootic. These findings indicated the continued presence of the PV for more than 1 year in the infected colony. Of 19 laboratory personnel who worked with the affected dogs, none, including 4 with a concurrent diarrheal disease, developed or had antibodies to the PV or canine coronavirus.     

Assessment of three automated assays for C-reactive protein determination in dogs

OBJECTIVE: To determine the characteristics of an automated canine C-reactive protein (CRP) assay and evaluate 2 human CRP assays for use in dogs. Animals-56 client-owned dogs with pyometra and 11 healthy control dogs. PROCEDURES: Samples from 11 dogs with high (> 100 mg/L) or low (< 10 mg/L) CRP concentrations (determined by use of a canine ELISA) were evaluated by use of the automated canine CRP assay. Intra- and interassay imprecision was determined (by use of those 2 plasma pools), and assay inaccuracy was assessed by use of logistic regression analysis of results obtained via ELISA and the automated canine CRP assay. Two automated human CRP assays were used to measure plasma CRP concentration in 10 dogs. RESULTS: By use of the ELISA, mean +/- SD plasma CRP concentration was 96.1 +/- 38.5 mg/L and 10.1 +/- 23.2 mg/L in dogs with pyometra and control dogs, respectively. The automated canine assay had intra-assay coefficients of variation (CVs) of 7.8% and 7.9%, respectively, and interassay CVs of 11.1% and 13.1%, respectively. Results from the automated assay were highly correlated with results obtained via ELISA. The human assay results did not exceed 0.4 mg/L in any dog. CONCLUSIONS AND CLINICAL RELEVANCE: The automated canine CRP assay had less interassay imprecision, compared with the ELISA. The 2 human CRP assays were not suitable for analysis of canine plasma samples. The automated canine CRP assay was more precise than the ELISA for serial evaluations of plasma CRP concentration in dogs.