OBJECTIVE To evaluate a fluorescence resonance energy transfer quantitative PCR (FRET-qPCR) assay for detection of gyrA mutations conferring fluoroquinolone resistance in canine urinary Escherichia coli isolates and canine urine specimens. SAMPLE 264 canine urinary E coli isolates and 283 clinical canine urine specimens. PROCEDURES The E coli isolates were used to validate the FRET-qPCR assay. Urine specimens were evaluated by bacterial culture and identification, isolate enrofloxacin susceptibility testing, and FRET-qPCR assay. Sensitivity and specificity of the FRET-qPCR assay for detection of gyrA mutations in urine specimens and in E coli isolated from urine specimens were computed, with results of enrofloxacin susceptibility testing used as the reference standard. RESULTS The validated FRET-qPCR assay discriminated between enrofloxacin-resistant and enrofloxacin-susceptible E coli isolates with an area under the receiver operating characteristic curve of 0.92. The assay accurately identified 25 of 40 urine specimens as containing enrofloxacin-resistant isolates (sensitivity, 62.5%) and 226 of 243 urine specimens as containing enrofloxacin-susceptible isolates (specificity, 93.0%). When the same assay was performed on E coli isolates recovered from these specimens, sensitivity (77.8%) and specificity (94.8%) increased. Moderate agreement was achieved between results of the FRET-qPCR assay and enrofloxacin susceptibility testing for E coli isolates recovered from urine specimens. CONCLUSIONS AND CLINICAL RELEVANCE The FRET-qPCR assay was able to rapidly distinguish between enrofloxacin-resistant and enrofloxacin-susceptible E coli in canine clinical urine specimens through detection of gyrA mutations. Therefore, the assay may be useful in clinical settings to screen such specimens for enrofloxacin-resistant E coli to avoid inappropriate use of enrofloxacin and contributing to antimicrobial resistance.

OBJECTIVE To evaluate the efficacy of each of 3 incremental doses of MK-467 for alleviation of dexmedetomidine-induced hemodynamic depression in isoflurane-anesthetized cats. ANIMALS 6 healthy adult domestic shorthair cats. PROCEDURES Each cat was anesthetized with isoflurane and received a target-controlled infusion of dexmedetomidine estimated to maintain the plasma dexmedetomidine concentration at 10 ng/mL throughout the experiment. Heart rate (HR) and direct arterial pressures were measured at baseline (isoflurane administration only), during dexmedetomidine infusion, and before and after IV administration of each of 3 serially increasing doses (15, 30, and 60 μg/kg) of MK-467. Cardiac index (CI) and systemic vascular resistance (SVR) were recorded at baseline, during dexmedetomidine infusion, and at the mean arterial pressure nadir after administration of the 30- and 60-μg/kg doses of MK-467. RESULTS Compared with baseline values, the dexmedetomidine infusion significantly decreased HR and increased arterial pressures. Each dose of MK-467 caused a significant decrease in arterial pressures and a significant, albeit clinically irrelevant, increase in HR (≤ 10%). Following administration of the 30- and 60-μg/kg doses of MK-467, all cats developed clinical hypotension (mean arterial pressure, < 60 mm Hg) even though CI and SVR returned to baseline values. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated administration of small doses of MK-467 to isoflurane-anesthetized cats receiving dexmedetomidine restored CI and SVR, but caused a substantial decrease in arterial pressures and only a marginal increase in HR. Therefore, caution should be used when MK-467 is administered to alleviate dexmedetomidine-induced hemodynamic depression in isoflurane-anesthetized cats.

OBJECTIVE To evaluate acute changes of the liver by use of shear wave elastography (SWE) and CT perfusion after radiofrequency ablation (RFA). ANIMALS 7 healthy Beagles. PROCEDURES RFA was performed on the liver (day 0). Stiffness of the ablation lesion, transitional zone, and normal parenchyma were evaluated by use of SWE, and blood flow, blood volume, and arterial liver perfusion of those regions were evaluated by use of CT perfusion on days 0 and 4. All RFA lesions were histologically examined on day 4. RESULTS Examination of the SWE color-coded map distinctly revealed stiffness of the liver tissue, which increased from the normal parenchyma to the transitional zone and then to the ablation zone. For CT perfusion, blood flow, blood volume, and arterial liver perfusion decreased from the transitional zone to the normal parenchyma and then to the ablation zone. Tissue stiffness and CT perfusion variables did not differ significantly between days 0 and 4. Histologic examination revealed central diffuse necrosis and peripheral hyperemia with infiltration of lymphoid cells and macrophages. CONCLUSIONS AND CLINICAL RELEVANCE Coagulation necrosis induced a loss of blood perfusion and caused tissue hardening (stiffness) in the ablation zone. Hyperemic and inflammatory changes of the transitional zone resulted in increased blood perfusion. Acute changes in stiffness and perfusion of liver tissue after RFA could be determined by use of SWE and CT perfusion. These results can be used to predict the clinical efficacy of RFA and to support further studies, including those involving hepatic neoplasia.

Mycoplasma hyorhinis (MHR) causes polyserositis and lameness in grower pigs. While herd-specific vaccines for this bacterium are being marketed, there are currently no licensed, commercially available vaccines for MHR. The objective of this study was to develop a challenge model in cesarean-derived, colostrum-deprived (CDCD) pigs using cell-associated MHR that results in both severe pericarditis and lameness, in order to evaluate suitable vaccine candidates. We investigated administering MHR to 7-week-old pigs over 3 d using 3 different routes compared to administering MHR on a single day using 1 of 3 routes. Pigs were monitored for 21 d for signs of lameness and well-being. At the end of the study, pigs were examined for evidence of polyserositis and arthritis associated with Mycoplasma. Results indicate that clinical manifestation of disease depended more on the route of administration than on the total dose given. A single intravenous (IV) administration of MHR resulted in extensive polyserositis, while a single intranasal (IN) administration showed little to no signs of disease. A single intraperitoneal (IP) administration did not induce the same level of polyserositis as observed in the IV group, but did result in an increased incidence of lameness. Furthermore, pigs administered MHR by IP (Day 0), IV (Day 1), and IN (Day 2) on 3 consecutive days showed a more robust disease manifestation, which resulted in both polyserositis and lameness. Optimization of this group showed that elimination of the 3rd-day IN challenge had no detrimental effect on clinical outcomes. The consecutive day administration of cell-associated MHR will allow polyserositis and lameness to be simultaneously evaluated in future vaccine trials.

OBJECTIVE To determine the physiochemical properties and pharmacokinetics of 3 midazolam gel formulations following buccal administration to dogs. ANIMALS 5 healthy adult hounds. PROCEDURES In phase 1 of a 2-phase study, 2 gel formulations were developed that contained 1% midazolam in a poloxamer 407 (P1) or hydroxypropyl methylcellulose (H1) base and underwent rheological and in vitro release analyses. Each formulation was buccally administered to 5 dogs such that 0.3 mg of midazolam/kg was delivered. Each dog also received midazolam hydrochloride (0.3 mg/kg, IV). There was a 3-day interval between treatments. Blood samples were collected immediately before and at predetermined times for 8 hours after drug administration for determination of plasma midazolam concentration and pharmacokinetic analysis. During phase 2, a gel containing 2% midazolam in a hydroxypropyl methylcellulose base (H2) was developed on the basis of phase 1 results. That gel was buccally administered such that midazolam doses of 0.3 and 0.6 mg/kg were delivered. Each dog also received midazolam (0.3 mg/kg, IV). All posttreatment procedures were the same as those for phase 1. RESULTS The H1 and H2 formulations had lower viscosity, greater bioavailability, and peak plasma midazolam concentrations that were approximately 2-fold as high, compared with those for the P1 formulation. The mean peak plasma midazolam concentration for the H2 formulation was 187.0 and 106.3 ng/mL when the midazolam dose administered was 0.6 and 0.3 mg/kg, respectively. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that buccal administration of gel formulations might be a viable alternative for midazolam administration to dogs.

OBJECTIVE To evaluate and compare surface and cross-sectional structure as well as localized electrochemical corrosion and ion release for cast stainless steel (SS) tibia plateau leveling osteotomy (TPLO) plates retrieved from dogs with and without osteosarcoma (OSA) and to compare these findings with similar variables for forged SS TPLO plates retrieved from dogs. SAMPLE 47 TPLO plates explanted from 45 client-owned dogs (22 cast plates from dogs with OSA, 22 cast plates from dogs without OSA, and 3 forged plates from dogs without OSA). PROCEDURES Histologic evaluations of tissue samples collected from implant sites at the time of plate retrieval were performed to confirm implant site tumor status of each dog. Surfaces and metallographic cross sections of retrieved plates were examined, and the microcell technique was used to obtain local electrochemical corrosion and ion release measurements. RESULTS Findings indicated that all cast SS plates demonstrated high spatial variability of their electrochemical surface properties and inhomogeneous superficial and cross-sectional composition, compared with forged plates. Greater metal ion release was observed in cast plates than in forged plates and in cast plates from dogs with OSA than in cast or forged from dogs without OSA. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that accumulation of metal ions from implants could be a trigger for neoplastic transformation in neighboring cells. Metal ion release caused by corrosion of implants that do not comply with recommended standards of the American Society for Testing and Materials International or the International Organization for Standardization could potentially place patients at increased risk of tumor development.

OBJECTIVE To assess 2 human ELISA kits for measurement of angiopoietin-1 and -2 concentrations in canine plasma samples, determine whether plasma angiopoeitin-2 concentration differed between septic and healthy dogs, and determine the effect of tumor necrosis factor-α (TNF-α) stimulation on angiopoeitin-2 release from primary canine aortic endothelial cells (pCAECs) in vitro. ANIMALS 10 healthy dogs and 10 septic dogs. PROCEDURES Human angiopoietin-1 and -2 ELISAs were used to detect recombinant canine angiopoietins-1 and -2 in canine plasma samples. The angiopoietin-2 ELISA was further validated by use of plasma samples from healthy and septic dogs and supernatants of pCAEC cultures. Associations between plasma angiopoeitin-2 and C-reactive protein (CRP) concentrations were examined. RESULTS Angiopoeitin-2 but not angiopoeitin-1 was detected in canine plasma samples by the respective ELISAs. The angiopoeitin-2 ELISA had excellent dilutional linearity, parallelism, accuracy, precision, and reproducibility for measurements in canine plasma samples and pCAEC supernatants. Plasma angiopoeitin-2 concentration was significantly higher in septic dogs (median, 25.5 ng/mL) than in healthy dogs (median, 6.7 ng/mL) and was positively correlated with plasma CRP concentration (R2 = 0.60). Stimulation of pCAECs with TNF-α resulted in a significant increase in supernatant angiopoietin-2 concentration. CONCLUSIONS AND CLINICAL RELEVANCE The tested human angiopoietin-2 ELISA kit was useful for measuring angiopoietin-2 concentrations in canine plasma samples and pCAEC supernatants. Sepsis appeared to increase angiopoietin-2 concentration in dogs in vivo, whereas TNF-α stimulation caused release of angiopoietin-2 from pCAECs in vitro. These findings support the use of angiopoietin-2 as a marker of endothelial cell activation and inflammation in dogs.

OBJECTIVE To quantitatively measure the amount of pressure induced at the calcaneus and cranial tibial surface of dogs by use of 2 cast configurations. ANIMALS 13 client- or student-owned dogs. PROCEDURES Pressure sensors were placed over the calcaneus and cranial tibial surface. Dogs then were fitted with a fiberglass cast on a pelvic limb extending from the digits to the stifle joint (tall cast). Pressure induced over the calcaneus and proximal edge of the cast at the level of the cranial tibial surface was simultaneously recorded during ambulation. Subsequently, the cast was shortened to end immediately proximal to the calcaneus (short cast), and data acquisition was repeated. Pressure at the level of the calcaneus and cranial tibial surface for both cast configurations was compared by use of paired t tests. RESULTS The short cast created significantly greater peak pressure at the level of the calcaneus (mean ± SD, 0.2 ± 0.07 MPa), compared with peak pressure created by the tall cast (0.1 ± 0.06 MPa). Mean pressure at the proximal cranial edge of the cast was significantly greater for the short cast (0.2 ± 0.06 MPa) than for the tall cast (0.04 ± 0.03 MPa). CONCLUSIONS AND CLINICAL RELEVANCE A cast extended to the level of the proximal portion of the tibia caused less pressure at the level of the calcaneus and the proximal cranial edge of the cast. Reducing the amount of pressure at these locations may minimize the potential for pressure sores and other soft tissue injuries.

OBJECTIVE To compare the effects of 3 equimolar concentrations of methylprednisolone acetate (MPA), triamcinolone acetonide (TA), and isoflupredone acetate (IPA) on equine articular tissue cocultures in an inflammatory environment. SAMPLE Synovial and osteochondral explants from the femoropatellar joints of 6 equine cadavers (age, 2 to 11 years) without evidence of musculoskeletal disease. PROCEDURES From each cadaver, synovial and osteochondral explants were harvested from 1 femoropatellar joint to create cocultures. Cocultures were incubated for 96 hours with (positive control) or without (negative control) interleukin (IL)-1β (10 ng/mL) or with IL-1β and MPA, TA, or IPA at a concentration of 10(−4), 10(−7), or 10(−10)M. Culture medium samples were collected from each coculture after 48 and 96 hours of incubation. Concentrations of prostaglandin E2, matrix metalloproteinase-13, lactate dehydrogenase, and glycosaminoglycan were determined and compared among treatments at each time. RESULTS In general, low concentrations (10(−7) and 10(−10)M) of MPA, TA, and IPA mitigated the inflammatory and catabolic (as determined by prostaglandin E2 and matrix metalloproteinase-13 quantification, respectively) effects of IL-1β in cocultures to a greater extent than the high (10(−4)M) concentration. Mean culture medium lactate dehydrogenase concentration for the 10(−4)M IPA treatment was significantly greater than that for the positive control at both times, which was suggestive of cytotoxicosis. Mean culture medium glycosaminoglycan concentration did not differ significantly. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that the in vitro effects of IPA and MPA were similar to those of TA at clinically relevant concentrations (10(−7) and 10(−10)M).

The purpose of this study was to determine the pharmacokinetics of cannabidiol (CBD) in healthy dogs. Thirty, healthy research dogs were assigned to receive 1 of 3 formulations (oral microencapsulated oil beads, oral CBD-infused oil, or CBD-infused transdermal cream), at a dose of 75 mg or 150 mg q12h for 6 wk. Serial cannabidiol plasma concentrations were measured over the first 12 h and repeated at 2, 4, and 6 wk. Higher systemic exposures were observed with the oral CBD-infused oil formulation and the half-life after a 75-mg and 150-mg dose was 199.7 ± 55.9 and 127.5 ± 32.2 min, respectively. Exposure is dose-proportional and the oral CBD-infused oil provides the most favorable pharmacokinetic profile.