In vitro cytopathic effects of a cysteine protease of Tritrichomonas foetus on cultured bovine uterine epithelial cells

OBJECTIVE: To evaluate the cytopathic effects of Tritrichomonas foetus and a purified cysteine protease (ie, CP30) of T foetus on cultured bovine uterine epithelial cells (BUECs) in vitro. SAMPLE POPULATION: 10 reproductive tracts were obtained from late-term bovine fetuses at a commercial abattoir. PROCEDURE: An in vitro culture system of BUECs was developed to study the cytopathic effects of T foetus and purified CP30 of T foetus on host cells. Cytotoxicity of T foetus or CP30 on exposed BUECs was determined. Fluorescence microscopy and flow cytometry analyses were used to detect apoptosis. A fluorometric assay was used to detect BUEC caspase 3 activation. The CP inhibitor E-64 and a caspase inhibitor were used to inhibit apoptosis. RESULTS: Cytopathic effects were observed in BUECs treated with parasites or CP30 and were concentration and time dependent. The BUECs underwent apoptosis in the presence of parasites or CP30. The specific CP inhibitor E-64 abolished the induction of apoptosis in BUECs by CP30. The caspase inhibitor reduced the amount of apoptosis in BUECs. CONCLUSIONS AND CLINICAL RELEVANCE: T foetus and its CP30 induce apoptosis in cultured BUECs in vitro. Induction of apoptosis by CP30 is correlated with protease activity. Endometrial cell death as a result of a T foetus infection is likely to be more important in mediating infertility than a direct effect on the conceptus. Provoking an apoptotic reaction in the host may mitigate an inflammatory reaction or immune response and therefore favor survival of the parasite in a chronic infection.     

Relative sensitivity of polymerase chain reaction assays used for detection of feline herpesvirus type 1 DNA in clinical samples and commercial vaccines

OBJECTIVE: To determine relative detection rates and detection limits for 6 published polymerase chain reaction (PCR) assays used for detection of feline herpesvirus type 1 (FHV-1) DNA. SAMPLE POPULATION: 5 vaccines licensed for use in preventing FHV-1-associated disease; 15 conjunctival biopsy specimens collected from cats with keratitis, conjunctivitis, or both; and a plaque-purified field isolate of FHV-1 cultured in vitro. PROCEDURE: Vaccines and clinical samples were assessed for FHV-1 DNA by use of all 6 assays. Detection rates were calculated by assuming that any sample in which FHV-1 DNA was detected was a true-positive result. Detection limits were estimated by use of serial dilutions of DNA extracted from cultured FHV-1 and 1 clinical sample. RESULTS: Testing by use of all 6 assays resulted in detection of FHV-1 DNA in all 5 vaccines. Testing by use of all 6 assays yielded concordant results for 9 of 15 conjunctival biopsy specimens (8 with negative results and 1 with a positive result). Calculated detection rates for clinical samples ranged from 29% to 86%. Assay sensitivity was ranked similarly by use of detection rate or detection limit. CONCLUSIONS AND CLINICAL RELEVANCE: Testing by use of all assays was equally likely to detect vaccine virus. Therefore, a positive PCR result in a cat may reflect vaccine virus rather than wild-type virus. Test sensitivity as assessed by detection limits and detection rates varied greatly. Because FHV-1 can be shed in clinically normal animals, high detection rate will not necessarily correlate with high diagnostic sensitivity.     

Evaluation of enzyme-linked immunosorbent assays performed on milk and serum samples for detection of paratuberculosis in lactating dairy cows

OBJECTIVE: To determine whether results obtained for milk and serum samples with ELISAs intended for diagnosis of paratuberculosis in dairy cows were comparable to results obtained by means of mycobacterial culture of fecal samples. DESIGN: Cross-sectional study. ANIMALS: 689 lactating dairy cows in 9 Ontario herds. PROCEDURE: Milk, serum, and fecal samples were obtained from all cows. Fecal samples were submitted for mycobacterial culture. Serum samples were tested with a commercially available ELISA for antibodies against Mycobacterium paratuberculosis, and preserved milk samples were tested with an indirect ELISA for antibodies against M paratuberculosis. RESULTS: Results were positive for 130 of the 689 (18.9%) serum samples, 77 of the 689 (11.1%) milk samples, and 72 of the 689 (10.4%) fecal samples. The level of agreement between results for milk and serum samples was only moderate. Proportions of positive results for serum and fecal samples were significantly different, but proportions of positive results for milk and fecal samples were not significantly different. In addition, results for milk samples had a higher level of agreement with results of mycobacterial culture than did results for serum samples. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that the indirect ELISA used on milk samples may be a convenient method of detecting paratuberculosis in dairy herds.     

Evaluation of the association between initial proteinuria and morbidity rate or death in dogs with naturally occurring chronic renal failure

OBJECTIVE: To determine whether urine protein-to-creatinine ratio (UP:C) > or = 1.0 at initial diagnosis of chronic renal failure (CRF) is associated with greater risk of development of uremic crises, death, and progression of renal failure in dogs. DESIGN: Prospective cohort study. ANIMALS: 45 dogs with CRF PROCEDURE: Dogs were prospectively assigned to 2 groups on the basis of initial UP:C < 1.0 or 2 > or = 1.0. The association between magnitude of proteinuria and development of uremic crises and death was determined before and after dogs with initial UP:C > or =1.0 were assigned to 3 subgroups and compared with dogs with initial UP:C < 1.0. Changes in reciprocal serum creatinine concentration were used to estimate decrease in renal function. RESULTS: Initially, dogs had similar clinical characteristics with the exception of systolic blood pressure and UP:C. Relative risks of development of uremic crises and death were approximately 3 times higher in dogs with UP:C > or =1.0, compared with dogs with UP:C < 1.0. Relative risk of adverse outcome was approximately 1.5 times higher for every 1-unit increment in UP:C. The decrease in renal function was of greater magnitude in dogs with UP:C > or =1.0, compared with dogs with UP:C < 1.0. CONCLUSIONS AND CLINICAL RELEVANCE: Initial UP:C > or =1.0 in dogs with CRF was associated with greater risk of development of uremic crises and death, compared with dogs with UP:C < 1.0. Initial determinations of UP:C in dogs with naturally occurring CRF may be of value in refining prognoses.     

An in vitro biomechanical comparison of the breaking strength and stiffness of polydioxanone (sizes 2, 7) and polyglactin 910 (sizes 3, 6) in the equine linea alba

OBJECTIVE: To determine failure strength, stiffness, and failure mode of 4 suture materials in equine linea alba. STUDY DESIGN: Randomized complete block design. SAMPLE POPULATION: Linea albas collected from 12 adult horses (mean weight, 475 kg; mean age, 10 years). METHODS: The ventral abdominal fascia, including the linea alba, was collected and bisected along the linea alba into right and left halves. Each half was divided into four, 5-cm sections extending from the umbilicus cranially, and randomly assigned to 1 of 4 suture materials: 2 polydioxanone (2 PD), 3 polyglactin 910 (3 PG), 6 polyglactin 910 (6 PG), and 7 polydioxanone (7 PD). A single cycle to failure test was performed on each specimen at a distraction rate of 100 mm/min. Differences in failure strength and stiffness for the materials were evaluated using a mixed linear model with significance set at P<.05. RESULTS: In 94 of 96 test sections, constructs failed by suture failure. There were significant differences in failure strength (P<.0001) and stiffness (P<.001) among the suture/tissue constructs. 7 PD had the highest breaking strength (316.8 N) followed by 6 PG (281.3 N), 3 PG (229.9 N), and 2 PD (193.0 N). Six PG had the largest stiffness (14 N/mm) followed by 3 PG (12.7 N/mm), 7 PD (10.1 N/mm), and 2 PD (7.2 N/mm). Suture breaking strength and stiffness were not affected by linea or fascia thickness, individual horse, half of the linea alba, or abdominal wall position. Eighty-five (90.4%) suture loops failed adjacent the knot. CONCLUSIONS: When tested in single cycle to failure, suture material was more likely to fail than the linea alba. Larger suture materials had higher breaking strengths than smaller suture materials and stiffness was also affected by suture material and size. CLINICAL RELEVANCE: Given its high initial breaking strength and its relatively longer in vivo strength retention, 7 PD seems an appropriate choice of suture material for closure of the equine linea alba when maximal short-term failure strength is desired.     

Tuberculosis and brucellosis prevalence survey on dairy cattle in Mbarara milk basin (Uganda)

We determined the prevalence of tuberculosis and brucellosis reactors in the dairy herds in the Mbarara district of Uganda in 2002. This is one of the most important dairy-production areas of the country and includes both pastoral and agro-pastoral zones. A total of 340 (of 11,995) randomly selected herds were tested for tuberculosis, using the intradermal tuberculosis-skin test and 315 (of 10,562) herds tested for brucellosis using the serum Rose Bengal test.

The herd prevalence for tuberculosis reactors was 74.1% (95% confidence intervals 69, 78), the individual-animal prevalence was of 6.0% (5.6, 6.5) and within-herd range was 1–50% (up to 100% if suspicious reactors were included).

The herd prevalence for brucellosis was 55.6% (50, 61.2) individual-animal prevalence 15.8% (14.8, 16.7) and within-herd range 1–90%.

The reactor prevalence increased with the age of the animals for both tuberculosis and brucellosis.

Tuberculosis reactor prevalences were higher in animals from the agro-pastoral zone. However, the individual-animal and herd prevalences of brucellosis seroprevalences were higher in the pastoral zone.     

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Determination of Mycoplasma bovis susceptibilities against six antimicrobial agents using the E test method

The objective of this study was to determine the susceptibility of Mycoplasma bovis against six antibiotics using the E test methodology. Fifty-eight isolates of M. bovis originating from 55 affected cattle were evaluated. Specimen originated from: lung tissue, synovial fluid, tracheo-bronchial wash, milk, and external or inner ear discharge. Antimicrobial agents tested were azythromycin, clindamycin, erythromycin, enrofloxacin, spectinomycin and tetracycline. The E test strips were placed on the surface of Hayflick plates on which organism suspension was spread. Plates were incubated at 35 degrees C in a candle jar for 72 h. MICs were then read by determining where the growth inhibition zone intersected with the MIC scale on the strip. M. bovis Donetta isolate was used as a control. All MICs were >256 microg/ml for erythromycin. MIC50 and MIC90 obtained for azythromycin were 3 and >256 microg/ml, respectively. MIC50 and MIC90 obtained for tetracycline were 4 and 8 microg/ml, respectively. MIC50 and MIC90 obtained for spectinomycin were 2 and >1021 microg/ml, respectively. MIC50 and MIC90 obtained for clindamycin were 0.19 and >256 microg/ml, respectively. MIC50 and MIC90 obtained for enrofloxacin were 0.19 and 0.25 microg/ml, respectively. Resistance was not associated with the specimen source except for azythromycin. M. bovis susceptibilities were easily determined by the E test which demonstrated the efficacy of enrofloxacin and the acquired resistance to tetracycline, spectinomycin, azythromycin and clindamycin.