A Quantitative Polymerase Chain Reaction Assay for the Detection and Quantification of Epizootic Epitheliotropic Disease Virus (EEDV; Salmonid Herpesvirus 3)

Epizootic epitheliotropic disease virus (EEDV; salmonid herpesvirus [SalHV3]; family Alloherpesviridae) causes a systemic disease of juvenile and yearling Lake Trout Salvelinus namaycush. No cell lines are currently available for the culture and propagation of EEDV, so primary diagnosis is limited to PCR and electron microscopy. To better understand the pervasiveness of EEDV (carrier or latent state of infection) in domesticated and wild Lake Trout populations, we developed a sensitive TaqMan quantitative PCR (qPCR) assay to detect the presence of the EEDV terminase gene in Lake Trout tissues. This assay was able to detect a linear standard curve over nine logs of plasmid dilution and was sensitive enough to detect single-digit copies of EEDV. The efficiency of the PCR assay was 99.4 ± 0.06% (mean ± SD), with a 95% confidence limit of 0.0296 (R² = 0.994). Methods were successfully applied to collect preliminary data from a number of species and water bodies in the states of Pennsylvania, New York, and Vermont, indicating that EEDV is more common in wild fish than previously known. In addition, through the development of this qPCR assay, we detected EEDV in a new salmonid species, the Cisco Coregonus artedi. The qPCR assay was unexpectedly able to detect two additional herpesviruses, the Atlantic Salmon papillomatosis virus (ASPV; SalHV4) and the Namaycush herpesvirus (NamHV; SalHV5), which both share high sequence identity with the EEDV terminase gene. With these unexpected findings, we subsequently designed three primer sets to confirm initial TaqMan qPCR assay positives and to differentiate among EEDV, ASPV, and NamHV by detecting the glycoprotein genes via SYBR Green qPCR.

Received April 20, 2015; accepted November 10, 2015     

Neutrophil myeloperoxidase measurements in plasma, laminar tissue, and skin of horses given black walnut extract

OBJECTIVE: To compare measurements of myeloperoxidase (MPO) in plasma, laminar tissues, and skin obtained from control horses and horses given black walnut heartwood extract (BWHE). ANIMALS: 22 healthy 5- to 15-year-old horses. PROCEDURES: Horses were randomly assigned to 4 groups as follows: a control group given water (n = 5) and 3 experimental groups given BWHE (17) via nasogastric intubation. Experimental groups consisted of 5, 6, and 6 horses that received BWHE and were euthanatized at 1.5, 3, and 12 hours after intubation, respectively. Control horses were euthanatized at 12 hours after intubation. Plasma samples were obtained hourly for all horses. Laminar tissue and skin from the middle region of the neck were harvested at the time of euthanasia. Plasma and tissue MPO concentrations were determined via an ELISA; tissue MPO activity was measured by use of specific immunologic extraction followed by enzymatic detection. RESULTS: Tissues and plasma of horses receiving BWHE contained significantly higher concentrations of MPO beginning at hour 3. Laminar tissue and skin from horses in experimental groups contained significantly higher MPO activity than tissues from control horses. Concentrations and activities of MPO in skin and laminar tissues were similar over time. CONCLUSIONS AND CLINICAL RELEVANCE: In horses, BWHE administration causes increases in MPO concentration and activity in laminar tissue and skin and the time of increased MPO concentration correlates with emigration of WBCs from the vasculature. These findings support the hypothesis that activation of peripheral WBCs is an early step in the pathogenesis of acute laminitis.     

Response of healthy dogs to infusions of human serum albumin

OBJECTIVE: To evaluate the clinical and immunologic response in healthy dogs to infusions of human serum albumin (HSA). ANIMALS: 9 healthy purpose-bred mixed-breed dogs. PROCEDURES: Each dog was administered a 25% HSA solution once or twice. Various physical examination and laboratory variables were serially evaluated. Antibody against HSA was assayed before and after infusion by use of an ELISA. Intradermal testing was also conducted. A repeated-measures ANOVA or Friedman repeated-measures ANOVA on ranks was used to compare results for the variables. RESULTS: Adverse clinical reactions were observed after the first or second infusion in 3 dogs. Anaphylactoid reactions were observed in 1 of 9 dogs during the first infusion and in 2 of 2 dogs administered a second infusion. Two dogs developed severe edema and urticaria 6 or 7 days after an initial infusion. All dogs developed anti-HSA antibodies. Positive responses for ID tests were observed in 8 of 9 dogs. Short-term increases were detected in blood protein, total bilirubin, and calcium concentrations after HSA infusion. Serum cholesterol concentrations and platelet counts decreased after HSA infusion. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of HSA resulted in profound reactions in 2 of 9 dogs administered a single infusion and in 2 of 2 dogs administered a second infusion. This indicates that there is risk of life-threatening adverse reactions to HSA infusion in healthy dogs.     

Development and psychometric testing of an instrument designed to measure chronic pain in dogs with osteoarthritis

OBJECTIVE: To develop and psychometrically test an owner self-administered questionnaire designed to assess severity and impact of chronic pain in dogs with osteoarthritis. SAMPLE POPULATION: 70 owners of dogs with osteoarthritis and 50 owners of clinically normal dogs. PROCEDURES: Standard methods for the stepwise development and testing of instruments designed to assess subjective states were used. Items were generated through focus groups and an expert panel. Items were tested for readability and ambiguity, and poorly performing items were removed. The reduced set of items was subjected to factor analysis, reliability testing, and validity testing. RESULTS: Severity of pain and interference with function were 2 factors identified and named on the basis of the items contained in them. Cronbach's alpha was 0.93 and 0.89, respectively, suggesting that the items in each factor could be assessed as a group to compute factor scores (ie, severity score and interference score). The test-retest analysis revealed kappa values of 0.75 for the severity score and 0.81 for the interference score. Scores correlated moderately well (r = 0.51 and 0.50, respectively) with the overall quality-of-life (QOL) question, such that as severity and interference scores increased, QOL decreased. Clinically normal dogs had significantly lower severity and interference scores than dogs with osteoarthritis. CONCLUSIONS AND CLINICAL RELEVANCE: A psychometrically sound instrument was developed. Responsiveness testing must be conducted to determine whether the questionnaire will be useful in reliably obtaining quantifiable assessments from owners regarding the severity and impact of chronic pain and its treatment on dogs with osteoarthritis.     

Comparison of three methods of temperature measurement in hypothermic, euthermic, and hyperthermic dogs

OBJECTIVE: To assess the reliability and accuracy of a predictive rectal thermometer, an infrared auricular thermometer designed for veterinary use, and a subcutaneous temperature-sensing microchip for measurement of core body temperature over various temperature conditions in dogs. DESIGN: Prospective study. ANIMALS: 8 purpose-bred dogs. PROCEDURES: A minimum of 7 days prior to study commencement, a subcutaneous temperature-sensing microchip was implanted in 1 of 3 locations (interscapular, lateral aspect of shoulder, or sacral region) in each dog. For comparison with temperatures measured via rectal thermometer, infrared auricular thermometer, and microchip, core body temperature was measured via a thermistor-tipped pulmonary artery (TTPA) catheter. Hypothermia was induced during anesthesia at the time of TTPA catheter placement; on 3 occasions after placement of the catheter, hyperthermia was induced via administration of a low dose of endotoxin. Near-simultaneous duplicate temperature measurements were recorded from the TTPA catheter, the rectal thermometer, auricular thermometer, and subcutaneous microchips during hypothermia, euthermia, and hyperthermia. Reliability (variability) of temperature measurement for each device and agreement between each device measurement and core body temperature were assessed. RESULTS: Variability between duplicate near-simultaneous temperature measurements was greatest for the auricular thermometer and least for the TTPA catheter. Measurements obtained by use of the rectal thermometer were in closest agreement with core body temperature; for all other devices, temperature readings typically underestimated core body temperature. CONCLUSIONS AND CLINICAL RELEVANCE: Among the 3 methods of temperature measurement, rectal thermometry provided the most accurate estimation of core body temperature in dogs.     

Efficacy of bovine herpesvirus-1 inactivated vaccine against abortion and stillbirth in pregnant heifers

OBJECTIVE: To evaluate the efficacy of an inactivated bovine herpesvirus-1 (BHV-1) vaccine to protect against BHV-1 challenge-induced abortion and stillbirth. DESIGN: Prospective study. ANIMALS: 35 beef heifers. PROCEDURES: Before breeding, heifers were vaccinated with a commercially available BHV-1 inactivated vaccine SC or IM. The estrous cycle was then synchronized, and heifers were artificially inseminated 30 to 60 days after vaccination. Heifers (n = 21) were challenge inoculated IV at approximately 180 days of gestation with virulent BHV-1. Fourteen control heifers were not vaccinated. Clinical signs of BHV-1 infection were monitored for 10 days following challenge; serologic status and occurrence of abortion or stillbirth were evaluated until time of calving. RESULTS: 18 of 21 (85.7%) heifers that received vaccine were protected from abortion following challenge, whereas all 14 control heifers aborted. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that an inactivated BHV-1 vaccine can protect against abortion resulting from a substantial challenge infection, with efficacy similar to that of modified-live BHV-1 vaccines.     

Responses of dogs to dietary omega-3 fatty acids

Long-chain omega-3 fatty acids from fish oil and other marine sources appear to be capable of modifying inflammatory and immune responses in dogs. Information is provided on the capacity of dogs to metabolize omega-3 fatty acids and the effects of omega-3 fatty acids on skin and coat, inflammatory responses, and neurologic development in puppies.