Isolation of granulocytes and mononuclear cells from the blood of dogs, cats, horses and cattle

A simple discontinuous Percoll density-gradient technique was adapted for isolation of granulocytes and mononuclear cells from cats, dogs, horses and cattle. Separation was accomplished at low speeds using a standard tabletop centrifuge. Cell purity was 100% for both granulocytes and mononuclear cells and cell viability exceeded 95%. Percent recovery of leukocytes ranged from 69 to 83%.     

Effects of delayed serum separation and long-term storage on the measurement of thyroid hormones in equine blood samples

Studies were conducted to determine the effects of delaying the separation of serum from the clot and of long-term storage of serum samples on the measurement of thyroid hormones in blood from horses using a fluorescence polarization immunoassay. The measured concentrations of T3 and T4 were not affected by leaving serum on the clot for as long as 24 hours at room temperatures. Storage of serum for 19 to 22 months at -20 degrees C resulted in significant increases of measured T4, but not T3. These studies support previous work demonstrating that thyroid hormones are resistant to degradation, immunologically stable, and reasonably insensitive to potential problems of routine specimen handling when measured with an immunoassay.     

Conjunctival fungal flora in horses, cattle, dogs, and cats

Conjunctival swab specimens were obtained from both eyes of 43 horses, 25 cows, 50 dogs, and 25 cats without keratitis or other ophthalmologic problems. Fungi were isolated from 95% of the horses, 100% of the cows, 22% of the dogs, and 40% of the cats. Aspergillus spp were isolated from 56% of the horses, 12% of the cows, 8% of the cats, and none of the dogs. Penicillium spp and Cladosporium spp were isolated ubiquitously. Collectively, 28 species from 209 isolants were identified.     

Detection and quantification of cocaine metabolites in urine samples from horses administered cocaine

Cocaine is a naturally occurring alkaloid that is commonly abused by human-beings for its psychostimulatory effects. Occasionally, very small concentrations (i.e. <100 ng/mL) of the primary cocaine metabolite, benzoylecgonine (BZE) have been detected in urine collected from horses competing in athletic events. In this study urine samples, collected from four horses following the administration of 2.5 and 20 mg of cocaine sublingually and 50 mg of cocaine intravenously, were analyzed for the presence of cocaine and/or its metabolites by enzyme-linked immunosorbent assay (ELISA) and gas chromatography-mass spectrometry (GC-MS). The results of ELISA analysis of urine samples collected from all four horses suggested the presence of cocaine and/or its metabolites up to 10, 48, and 72 h after administration of 2.5, 20, and 50 mg of cocaine, respectively. The results of GC-MS analysis confirmed the presence of BZE above the limit of quantification (LOQ = 5 ng/mL) in urine samples collected from all four horses for up to 24 h after administration of 2.5 mg of cocaine and for up to 48 h after administration of 20 and 50 mg of cocaine. No obvious behavioral effects or overt alterations of heart rate or rhythm were noted in any of these horses after cocaine administration.     

Antimicrobial drug susceptibility of bacteria isolated from disease processes in cattle, horses, dogs and cats

Results are presented of antimicrobial disc diffusion susceptibility testing on commonly isolated bacterial pathogens made at the Ontario Veterinary College Diagnostic Bacteriology Laboratory in 1981 and 1982. Nearly 2 000 isolates from horses, cattle, dogs and cats were tested. Comparison of resistance in the same bacterial species isolated from different animal species showed significant differences between some of the same antibiotics.     

Effects of storage time and temperature on pH,specific gravity,and crystal formation in urine samples from dogs and cats

OBJECTIVE: To determine effects of storage temperature and time on pH and specific gravity of and number and size of crystals in urine samples from dogs and cats. DESIGN: Randomized complete block design. ANIMALS: 31 dogs and 8 cats. PROCEDURE: Aliquots of each urine sample were analyzed within 60 minutes of collection or after storage at room or refrigeration temperatures (20 vs 6 degrees C [68 vs 43 degrees F]) for 6 or 24 hours. RESULTS: Crystals formed in samples from 11 of 39 (28%) animals. Calcium oxalate (CaOx) crystals formed in vitro in samples from 1 cat and 8 dogs. Magnesium ammonium phosphate (MAP) crystals formed in vitro in samples from 2 dogs. Compared with aliquots stored at room temperature, refrigeration increased the number and size of crystals that formed in vitro; however, the increase in number and size of MAP crystals in stored urine samples was not significant. Increased storage time and decreased storage temperature were associated with a significant increase in number of CaOx crystals formed. Greater numbers of crystals formed in urine aliquots stored for 24 hours than in aliquots stored for 6 hours. Storage time and temperature did not have a significant effect on pH or specific gravity. CONCLUSIONS AND CLINICAL RELEVANCE: Urine samples should be analyzed within 60 minutes of collection to minimize temperature- and time-dependent effects on in vitro crystal formation. Presence of crystals observed in stored samples should be validated by reevaluation of fresh urine.     

Detection and quantification of Anaplasma marginale DNA in blood samples of cattle by real-time PCR

A TaqMan-based real-time PCR assay was developed for the diagnosis of Anaplasma marginale infection of cattle. The established assay was proven to be highly specific, since no cross-reactions were observed with other Anaplasma species of ruminants, including the closely related Anaplasma centrale, or other haemoparasites of ruminants (Anaplasma bovis, Anaplasma ovis, Anaplasma phagocytophilum, Babesia bovis, Babesia bigemina, Theileria annulata and Theileria buffeli). The detection limit was equal to that of nested (n)PCR (10(1) copies of standard DNA and 3 x 10(1) infected erythrocytes ml(-1) of blood). The assay was also reproducible, as shown by satisfactory low intra-assay and inter-assay coefficients of variation. Fifty-four blood samples of ruminants (cattle, n = 51; sheep, n = 2; goats, n = 1), that had been tested previously by reverse line blot (RLB) hybridisation, were subjected to an nPCR assay and the newly established real-time PCR assay. By using real-time PCR, A. marginale DNA was detected in 39/51 bovine samples, with DNA titres ranging from 3.60 x 10(3) to 5.70 x 10(8) copies ml(-1) of blood, whereas sheep and goat samples tested negative. The concordance with nPCR was 100%, whereas a unique sample that had tested negative by RLB gave positive results by nPCR and real-time PCR. The established assay could overcome the limitations of existing diagnostic methods, allowing for simultaneous detection and quantification of the A. marginale DNA in bovine blood, that is essential to support the clinical diagnosis, to assess the carrier status of the animals and to evaluate the efficacy of vaccines and antirickettsial drugs.     

Quantitative buffy coat analysis of blood collected from dogs, cats, and horses

Using quantitative buffy coat analysis (QBCA), rapid and accurate measurements can be made of the erythrocyte PCV, total WBC count, and platelet count, and the leukocyte population can be differentiated into total granulocytes (including quantitation of eosinophils), and lymphocytes and monocytes. The QBCA is performed by placing a blood sample (50 to 111 microliters) into a high-precision-bore microhematocrit tube that contains a freely moving, closely fitting, cylindrical plastic float. After centrifugation for 5 minutes, the buffy coat components separate by density. The plastic cylinder floats in the buffy coat, thereby expanding the lengths of the buffy coat layers. The layers are measured in a manner that is similar to that used for measuring PCV. Results of QBCA of blood samples from dogs, cats, and horses indicated that the hematologic values obtained correlated with results obtained by use of conventional methods. The accuracy and ease of use of QBCA and the availability of results while the animal is still being examined make QBCA a useful tool for hematologic evaluation of animals.     

Blood serum level of primary bile acids in cattle, horses, swine and dogs

The levels of the two primary bile acids, cholic acid (CA) and chenodeoxycholic acid (CDCA), were determined by radioimmunoassay in cattle, horse, pig and dog serum. The mean serum cholic acid (SCA) and deoxycholic acid (SCDCA) levels of cows varied with their reproductive status, being 7.8 (+/- 3.3) and 1.5 (+/- 1.0) mumol/l in dry cows, 17.8 (+/- 6.9) and 2.3 (+/- 1.0) mumol/l in freshly calved dams, and 15.8 (+/- 5.7) and 2.3 (+/- 0.8) mumol/l, respectively, in lactating cows. The SCA level found in the immediate prepartal period and also on the day of calving corresponded to those found during the dry period, then, they tended to rise 2 days after calving and attained the peak characteristic for freshly calved dams on day 3 or 4 post partum. Feed consumption had no influence on the serum levels of primary bile acids, and circadian variations of SCA and SCDCA were also negligible. Suckling calves had much lower SCA levels (2.3 (+/- 1.0) mumol/l before feeding than cows. This initial concentration rose to 10.3 (+/- 2.9) mumol/l 1 h after feeding and returned to 5.0 (+/- 2.1) mumol/l 3 h later. Like cows, horses showed no appreciate difference between pre- and post-feeding levels of SCA (2.2 (+/- 1.2) mumol/l) and SCDCA (1.1 (+/- 0.3) mumol/l). Unlike bovines, pigs and dogs showed a considerable increase in the serum levels of the primary bile acids after feeding.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of syringe type and storage temperature on results of blood gas analysis in arterial blood of horses

BACKGROUND: Results of arterial blood gas analysis can be biased by pre-analytical factors, such as time to analysis, syringe type, and temperature during storage. However, the acceptable delay between time of collection and analysis for equine arterial blood gas remains unknown. HYPOTHESIS: Dedicated plastic syringes provide better stability of arterial blood gases than multipurpose plastic syringes. ANIMALS: Eight mares, 1 stallion, and 1 gelding, ages 3 to 10 years old. METHODS: Arterial blood samples were collected in a glass syringe, a plastic syringe designated for blood gas collection, and a multipurpose tuberculin plastic syringe. Blood samples were stored at ambient temperature or in iced water. For each sample, partial pressure of oxygen in arterial blood (PaO2), partial pressure of carbon dioxide in arterial blood (PaCO2), and pH were measured within a few minutes of collection and at 5, 20, 30, 60, 90, and 120 minutes after collection. RESULTS: Collection into glass syringes stored in iced water provided adequate PaO2 results for up to 117 +/- 35 minutes, whereas blood collected in either of the plastic syringes resulted in a variation >10 mm Hg after 10 +/- 3 to 17 +/- 2 minutes, depending on the storage conditions. Plastic syringes kept at ambient temperature offered more stability for PaCO2 analysis because they could be stored up to 83 +/- 16 minutes without significant variations. Values of pH did not show variations more than 0.02 for the first hour, irrespectively of storage condition. CONCLUSIONS AND CLINICAL IMPORTANCE: Glass syringes placed on ice are preferable for analysis of PaO2. Blood collected in plastic syringes should be analyzed within 10 minutes, irrespective of the storage temperature, to ensure the accuracy of PaO2 values.     

Thiopurine methyltransferase in red blood cells of dogs, cats, and horses

Our objective was to determine if thiopurine methyltransferase (TPMT), the enzyme important in the metabolism of azathioprine in human beings, is detectable in red blood cell lysates (RBCL) of healthy dogs, cats, and horses. Values for TPMT activity were determined from blood collected from 20 healthy dogs, cats, and horses. The TPMT activity in each animal's RBCL was determined using a radioenzymatic end point involving TPMT-facilitated metabolism of 6-mercaptopurine to 6-methylmercaptopurine (6-MMP). One unit of TPMT activity represents the formation of 1 nmol of 6-MMP per milliliter of packed red blood cells per hour of incubation at 37 degrees C. TPMT activity in RBCL was detectable in all species, with mean RBC values +/- standard deviation of 17.9 +/- 3.79 U/mL in dogs; 2.76 +/- 0.70 U/mL in cats; and 2.185 +/- 0.36 U/mL in horses. Values for TPMT in the 3 species were significantly (P < .05) different from one another. TPMT values for dogs were significantly higher than the other species, and TPMT values for cats were significantly higher than those for horses. We conclude that RBCL TPMT values are measurable in dogs. cats, and horses and that dogs have higher values than cats or horses. These findings are consistent with the lower tolerance for azathioprine in cats as compared with dogs. It remains to be determined whether RBCL TPMT values in these species correlate with TPMT activity in the liver, where most of the metabolization of azathioprine is believed to occur.     

Effects of plasma sample storage on blood ammonia, bilirubin, and urea nitrogen concentrations: cats and horses

Ten horses, a pony, and 13 cats were used to evaluate base-line blood ammonia, bilirubin, and urea nitrogen concentrations and to determine The effects of prolonged cold storage (-20 degrees C) before assay. Base-line plasma ammonia concentrations in cats (0.992 +/- 0.083 [SE] micrograms/ml) did not change significantly after 48 hours of storage (0.871 +/- 0.073 micrograms/ml); however, they were increased 4.2- and 13-fold after 168 and 216 hours of storage, respectively. In contrast to base-line plasma-ammonia values in cats, those of horses were significantly (0.265 +/- 0.044 micrograms/ml) lower, and significantly increased from base-line values after 48 hours of storage (0.861 +/- 0.094 micrograms/ml) and continued to increase 25.6-fold at 168 hours and 18.4-fold at 216 hours. Plasma urea nitrogen concentrations in cats (25.8 +/- 1.06 mg/dl) and horses (11.2 +/- 0.749 mg/dl) did not change significantly during 168 hours of storage. Total plasma bilirubin values from both cats (0.19 +/- 0.049 mg/dl) and horses (0.75 +/- 0.064 mg/dl) also did not change significantly during storage. These results indicate that feline plasma samples for ammonia determinations may be stored at -20 degrees C for up to 48 hours, whereas equine plasma ammonia values tend to increase during that time. The reason for the increase remains unexplained. Both feline and equine plasma urea nitrogen and total bilirubin are stable for at least 168 hours of storage at -20 degrees C.     

Epidemiologic analysis of oral and pharyngeal cancer in dogs, cats, horses, and cattle.

Four hundred sixty-nine oral-pharyngeal malignancies diagnosed in dogs, cats, horses, and cattle and submitted to the Viterinary Medical Data Program between March 1, 1964, and Dec 31, 1974, were analyzed. Of these cases, 84% were in dogs. The most frequent oral-pharyngeal cancer in dogs was melanoma; in cats and horses, it was squamous cell carcinoma. In dogs, the risk of developing melanoma increased more with age than did the risk of developing squamous cell carcinoma and fibrosarcoma. Male dogs had significantly greater risk of developing fibrosarcomas and melanomas than did female dogs. The German Shorthaired Pointer, Weimaraner, Golden Retriever, Boxer, and Cocker Spaniel breeds had significantly higher risk and Dachshunds and Beagles had significantly lower risk, as compared with all breeds combined. There was no significant difference between observed and expected numbers of tonsillar carcinomas diagnosed at veterinary colleges located in small urban areas (less than 50,000 persons) as compared with large urban populations (greater than 500,000).     

Relationship of serum total calcium to serum albumin in dogs, cats, horses and cattle

A retrospective study was performed in order to assess the relationship between serum calcium and serum albumin concentrations in domestic animals. Results of 9041 canine, 1564 feline, 2917 equine, and 613 bovine serum samples from hospitalized patients were examined by regression analysis. Subpopulations of cases with concurrent elevations in creatinine or that were less than six months of age were evaluated separately. Statistically significant linear relationships between calcium and albumin concentrations were established for each species (p <0.05). The coefficients of determination (r²) were 0.169 for dogs, 0.294 for cats, 0.222 for horses, and 0.032 for cattle. The correlation coefficients (r) computed were: dogs = 0.411, cats = 0.543, horses = 0.471, cattle = 0.182. Neither increases in creatinine concentration nor juvenile age appreciably influenced the relationship between calcium and albumin concentrations. Interspecies variation was marked, and a strong correlation between calcium and albumin concentrations was not established in any species.