Plasma Concentrations of Oral Ondansetron in Hospitalized Dogs Exhibiting Clinical Signs of Nausea.

The purpose of this study was to evaluate plasma ondansetron (OND) concentrations in a population of dogs with naturally occurring nausea after oral OND administration. Twenty-four dogs were randomly assigned to receive one of the following doses of oral OND: 0.5 mg/kg q8h, 0.5 mg/kg q12h, 1 mg/kg q8h, or 1 mg/kg q12h. Blood samples for plasma OND measurements were collected at baseline and 2, 4, and 8 h after administration of the first dose of OND. OND concentrations averaged over an 8 h time period were not significantly different between dose groups (0.5 mg/kg group: median 8.5 ng/mL [range 1-96.8 ng/mL], 1 mg/kg group: median 7.4 ng/mL [range 1-278.7 ng/mL]). The mean maximum concentrations in the 0.5 mg/kg and 1 mg/kg groups were 35.8 ± 49.0 ng/mL and 63.3 ± 121.1 ng/mL, respectively. OND concentrations were below the lower limit of quantification (LLOQ) in 50% (18/36) of samples in the 0.5 mg/kg groups and 39% (14/36) of samples in the 1 mg/kg groups. Six dogs (6/24, 25%) did not have OND detected at any time. The mean nausea scores at baseline were similar amongst all groups and decreased over time. The bioavailability of oral OND appears to be poor. Despite low plasma OND concentrations, nausea scores improved over time.

Effects of canine influenza infection and DA2PP vaccination on the development of platelet-associated immunoglobulins and platelet counts in dogs.

BACKGROUND: Immune thrombocytopenia (ITP) is commonly associated with platelet-associated immunoglobulins (PAIg). Demonstration of PAIg can help determine etiologies for thrombocytopenia. In humans, ITP and thrombocytopenia have been associated with various vaccinations and influenza infections, respectively. OBJECTIVES: We aimed to evaluate platelet counts and PAIg in research dogs with H3N2 and in research and client-owned dogs routinely vaccinated for distemper, adenovirus-2, parainfluenza, and parvovirus (DA2PP). The hypotheses were that H3N2 infection but not DA2PP vaccination would decrease platelet counts, and neither would result in the detection of PAIg. METHODS: Three pilot studies. Platelet counts and PAIg, measured by direct flow cytometry as %IgG, were evaluated in eight research Beagles following experimental infection with H3N2 (experiment 1), nine research Beagles vaccinated for DA2PP (experiment 2), and thirty client-owned dogs vaccinated for DA2PP (experiment 3). All animals were considered healthy at the start of the experiments. RESULTS: Transient, self-resolving decreases in platelet counts and increases in %IgG occurred following H3N2 infection, and one dog became thrombocytopenic and positive for PAIg. Following DA2PP vaccination, %IgG increased in research and client-owned dogs, but only one dog was considered positive for PAIg with a concurrent increase in platelet count. Mean PAIg increased from baseline in client-owned dogs following vaccination. CONCLUSIONS: Transient PAIg and thrombocytopenia can occur following H3N2 infection, while routine vaccination for DA2PP in this group of dogs was not associated with the development of thrombocytopenia or clinically relevant formation of PAIg.

A prospective cohort study to identify clinical diagnostic and prognostic markers of primary immune thrombocytopenia in dogs.

BACKGROUND: Primary immune thrombocytopenia (pITP) in dogs presents a diagnostic challenge, and clinical markers of severity are lacking. OBJECTIVES: Identify clinicopathologic features that differentiate pITP from secondary ITP (sITP) and markers related to bleeding severity, transfusion, and survival of dogs with pITP. ANIMALS: Ninety-eight thrombocytopenic dogs (58 pITP and 40 sITP). METHODS: Client-owned dogs with platelet counts <50 000/µL were enrolled in a prospective, multi-institution cohort study. History and treatment information, through a maximum of 7 days, was recorded on standard data forms. Bleeding severity was scored daily using a bleeding assessment tool (DOGiBAT). At-admission blood samples were collected for CBC, biochemistry, C-reactive protein concentration, and coagulation panels, and to measure platelet surface-associated immunoglobulin G (PSAIg) and expression of platelet membrane proteins and phospholipids. Dogs with evidence of coincident disease were classified as sITP. RESULTS: No definitive pITP diagnostic test was found. However, pITP cases were characterized by lower platelet counts, D dimer concentrations, and platelet membrane protein expression than sITP cases. Differentiation between pITP and sITP was further enhanced using logistic regression modeling combining patient sex, coagulation profile, platelet count, D dimer, and PSAIg. A second model of pITP severity indicated that low hematocrit and high BUN concentration were associated with non-survival. Low hematocrit at admission, but not platelet count or DOGiBAT score, was associated with transfusion. CONCLUSIONS AND CLINICAL IMPORTANCE: Pending validation studies, models constructed from at-admission clinicopathologic findings may improve differentiation of pITP from sITP and identify the most severe pITP cases at the time of presentation.

Multi-institutional retrospective study investigating blood culture protocols and test positivity in 701 dogs.

Objectives: (i) To determine the influence of specimen collection protocol (timing and specimen quantity), primary disease process, and pre-existing antimicrobial or immunosuppressive therapy on blood culture (BC) positivity and (ii) To determine agreement between urine culture and BC results. Animals: 701 client-owned dogs. Methods: Multi-institutional retrospective study (2019-2022). Mixed-effect logistic regression was used to determine whether primary disease process, the number of BCs, or the timing of specimen collection was associated with BC positivity. Prediction plots were generated. Associations between urine culture and BC results were performed using logistic regression. Results: Dogs with a positive urine culture were more likely to have a positive BC (OR: 4.36, 95% CI: 2.12-8.97, p = 0.003). Dogs that had three BC specimens had the greatest odds of obtaining a positive BC result (adjusted predictive value: 0.44, 95% CI: 0.21-0.70), although this was not significant. Isolates from 38.5% of dogs with a positive BC had resistance to ≥3 antimicrobial classes. The timing between specimen collection had no significant association with BC positivity. Pre-existing antibiotic or immunosuppressive therapy had no significant association with BC positivity. Clinical relevance: Dogs with a positive urine culture were more likely to have a positive BC result.

Fanconi syndrome with hepatorenal karyomegaly in a young Sphynx cat.

Case summary: A 3-year-old male neutered Sphynx cat was referred for history of chronically increased liver enzymes and lower urinary tract signs that were first reported when the cat was 5 months old. Urine metabolic profile revealed increased amino aciduria and glucosuria despite normoglycemia, suggesting Fanconi syndrome. Urine sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a banding pattern suggestive of primary tubular damage. Serial blood work showed non-regenerative normocytic normochromic anemia, persistently elevated liver enzymes, worsening azotemia and progressive hyperchloremic metabolic acidosis. Ultrasound revealed irregular kidneys and bilaterally hyperechoic cortices and medullae with a loss of normal corticomedullary distinction. Laparoscopic kidney biopsy revealed a moderate-to-severe chronic interstitial fibrosis with chronic lymphoplasmacytic inflammation, tubular degeneration and atrophy, mild glomerulosclerosis and mild large vascular amyloidosis. Tubular epithelial cell karyomegaly was multifocally evident throughout the kidney. The liver had moderate diffuse zone 1 hepatocellular atrophy, periportal fibrosis, biliary hyperplasia, mild perisinusoidal amyloidosis and hepatocyte karyomegaly in zones 2 and 3. The patient continued to decline and developed polyuria, polydipsia, lethargy and hyporexia irrespective of rigorous management, which failed to curtail the progressive anemia and azotemia. The patient was euthanized 8 months from the onset of clinical signs. Relevance and novel information: Fanconi syndrome in cats is a rare condition, with most reports occurring secondary to chlorambucil treatment. This is the first known case of Fanconi syndrome occurring with concurrent hepatorenal epithelial karyomegaly in a young Sphynx cat.

Sino-orbital aspergillosis with obstructive cervical lymphadenopathy in a cat caused by Aspergillus viridinutans complex.

Case summary: This report describes an indoor-only cat with a rare form of sino-orbital aspergillosis (SOA) with cervical lymphadenopathy causing local obstruction. Extensive work-up on initial presentation failed to identify the underlying etiology and the diagnosis was not determined until the disease progressed during a prolonged course of glucocorticoid therapy. Relevance and novel information: SOA caused by Aspergillus viridinutans complex is increasingly recognized as a significant cause of mortality in cats in recent years, with most cases reported in Australia, Europe and Asia. Feline SOA carries a poor prognosis owing to its invasive nature and resistance to antifungal therapy. This case demonstrates the importance of clinical awareness of SOA as a differential for cats with chronic nasal signs and exophthalmos in the USA. Moreover, it demonstrates a rare form of presentation and potential difficulty in achieving a correct diagnosis.

Prevalence of signs of lower urinary tract disease and positive urine culture in dogs with diabetes mellitus: A retrospective study.

BACKGROUND: No recent studies have evaluated the association between clinical signs of lower urinary tract disease (LUTD) and positive urine culture in dogs with diabetes mellitus. OBJECTIVE: Determine the prevalence of subclinical bacteriuria (ie, positive urine culture without signs of LUTD) in dogs with diabetes mellitus. ANIMALS: One hundred seven dogs with diabetes mellitus were evaluated at a university veterinary hospital. METHODS: Retrospective study evaluating diabetic dogs with a single sample paired urinalysis and urine culture. Relationship between the presence of signs of LUTD, pyuria, and bacteriuria and urine culture results were compared using Fisher exact testing. RESULTS: Fifteen dogs (14%) had a positive urine culture via cystocentesis or free catch, of which 8 (53%) had pyuria, and 4 (27%) had signs of LUTD. Of the 88 dogs (82%) without signs of LUTD, 11 (13%) had a positive culture. A significant association was found between a positive urine culture and pyuria (OR infinity; 95% CI 20.34-infinity, P < .00001) and bacteriuria (OR infinity; 95% CI 164.4-infinity, P < .00001). No association was found between urine culture results and signs of LUTD (OR 1.87; 95% CI 0.59-6.85, P = .46). CONCLUSION AND CLINICAL IMPORTANCE: Subclinical bacteriuria occurred in this cohort of dogs, and our findings reinforce the recommendation that urine cultures should not be routinely performed in diabetic dogs particularly if pyuria and bacteriuria are absent.

Prevalence of aldosterone breakthrough in dogs receiving renin-angiotensin system inhibitors for proteinuric chronic kidney disease.

BACKGROUND: The influence of aldosterone breakthrough (ABT) on proteinuria reduction during renin-angiotensin system (RAS) inhibition for spontaneous proteinuric chronic kidney disease (CKDP ) has not been determined in dogs. OBJECTIVES: Determine whether ABT occurs in dogs with CKDP and if it is associated with decreased efficacy in proteinuria reduction during RAS inhibitor treatment. ANIMALS: Fifty-six client-owned dogs with CKDP and 31 healthy client-owned dogs. METHODS: Prospective, multicenter, open-label clinical trial. Dogs were treated with an angiotensin-converting enzyme inhibitor or angiotensin receptor blocker alone or in combination at the attending clinician's discretion and evaluated at 5 time points over 6 months. Healthy dogs were used to determine the urine aldosterone-to-creatinine ratio cutoff that defined ABT. The relationship of ABT (present at ≥50% of visits) and proteinuria outcome (≥50% reduction in urine protein-to-creatinine ratio from baseline at ≥50% of subsequent visits) was evaluated. Mixed effects logistic regression was used to evaluate the relationship between clinical variables and outcomes (either successful proteinuria reduction or ABT). RESULTS: Thirty-six percent (20/56) of dogs had successful proteinuria reduction. Between 34% and 59% of dogs had ABT, depending on the definition used. Aldosterone breakthrough was not associated with proteinuria outcome. Longer duration in the study was associated with greater likelihood of successful proteinuria reduction (P = .002; odds ratio, 1.6; 95% confidence interval [CI], 1.2-2.2). CONCLUSIONS AND CLINICAL IMPORTANCE: Aldosterone breakthrough was common in dogs receiving RAS inhibitors for CKDp but was not associated with proteinuria outcome.

Association between feline hyperthyroidism and thoracic radiographic evaluation of cardiomegaly and pulmonary hyperinflation.

OBJECTIVES: Hyperthyroidism frequently affects middle-to-older-aged cats that can present with cardiorespiratory signs. The effects of hyperthyroidism on cardiac size and function have been previously documented. Anecdotally, pulmonary hyperinflation identified on thoracic radiographs may also be associated with hyperthyroidism; however, there is no literature to support this claim. The goal of this study was to determine any association between hyperthyroidism, pulmonary hyperinflation and cardiomegaly with the following hypotheses: (1) hyperthyroid cats would not have evidence of radiographic pulmonary hyperinflation compared with control cats; and (2) hyperthyroid cats were more likely to have evidence of radiographic cardiomegaly than control cats. METHODS: In this retrospective case-control study, the thoracic radiographs of 52 hyperthyroid cats and 46 non-hyperthyroid cats were evaluated for subjective and objective measurements of pulmonary hyperinflation and cardiomegaly. RESULTS: There were no statistically significant differences between hyperthyroid and non-hyperthyroid cats for any variable indicative of pulmonary hyperinflation. The mean ± SD vertebral heart score on lateral views for hyperthyroid cats was 7.75 ± 0.53 and for control cats was 7.55 ± 0.54, which was significantly different (P = 0.05). Among all cats, a more severe total elevation in thyroxine (T4) was correlated with a larger vertebral heart score on lateral views (Spearman's correlation coefficient = 0.23, P = 0.02). CONCLUSIONS AND RELEVANCE: While the results of this study suggest that hyperthyroid cats are more likely to have a larger vertebral heart score on lateral views than control cats, the clinical relevance of this finding is unclear given the large degree of overlap between hyperthyroid and non-hyperthyroid cats. In addition, among all cats, a greater total T4 elevation was weakly correlated with a larger vertebral heart score. Hyperthyroidism is not associated with radiographic pulmonary hyperinflation and is an unlikely differential for this radiographic finding.

Pharmacokinetics and anti-nausea effects of intravenous ondansetron in hospitalized dogs exhibiting clinical signs of nausea.

The purpose of this study was to evaluate the pharmacokinetics of intravenous (IV) ondansetron in a population of hospitalized dogs exhibiting clinical signs of nausea. The causes of nausea included pancreatitis, gastroenteritis, endocarditis, chemotherapy-induced nausea, diabetes mellitus and ketoacidosis, acute kidney injury with aspiration pneumonia, pyometra, uroabdomen, neoplasia, and hepatopathy. Twenty-four dogs were randomly assigned to one of the following IV ondansetron protocols: 1 mg/kg q12h, 0.5 mg/kg q12h, 1 mg/kg q8h, 0.5 mg/kg q8h. Serum was collected at 0, 0.25, 0.5, 1, 2, 4, 8, 16, and 24 h after the first dose, and nausea scores were recorded at multiple time points. Ondansetron and arginine vasopressin (AVP) concentrations were measured via high-performance liquid chromatography coupled to tandem mass spectrometry and ELISA, respectively. Noncompartmental pharmacokinetic modeling and dose interval modeling were performed. Ondansetron displayed linear pharmacokinetics. In the 0.5 mg/kg group, mean Cmax  = 214 ng/ml, AUC0-8h  = 463 ng/ml*h, and calculated half-life was 1.9 h. In the 1 mg/kg group, mean Cmax  = 541 ng/ml, AUC0-8h  = 1057 ng/ml*h and calculated half-life was 1.6 h. Serum ondansetron concentrations were not significantly different between dogs that required rescue anti-nausea medication (non-responders) and dogs that did not require rescue therapy (responders). In total, 83.3% of patients in the 0.5 mg/kg q8h, 0.5 mg/kg q12h, and 1 mg/kg q8h groups had improvement in nausea scores. In total, 66.7% of patients in the 1 mg/kg q12h group had improvement in nausea scores. In total, 33% of patients had resolution of nausea in the 0.5 mg/kg q8h, 1 mg/kg q8h, and 1 mg/kg q12h groups, and 16% of patients had resolution of nausea in the 0.5 mg/kg q12h group. AVP concentrations were highly variable and did not correlate with nausea scores. Nausea scores significantly decreased regardless of dosage protocol. AVP was not a reliable biomarker of nausea in this group of dogs.

Urine bacterial culture growth and association with urine sedimentation and clinical findings in cats with acute kidney injury.

OBJECTIVES: A urine culture is often pursued in cats with acute kidney injury (AKI) to screen for bacterial growth in the urine, but it can be cost prohibitive. The aim of the study was to determine the ability of a urinalysis and lower urinary tract signs (LUTS) to predict urine culture results in cats with AKI. METHODS: Ninety-seven cats with AKI were included in this study. This was a retrospective, observational study. Medical records from 2008 to 2018 were reviewed to identify cats with AKI that had a paired urinalysis and urine bacterial culture. The sensitivity, specificity, positive predictive value and negative predictive values of microscopic bacteriuria, pyuria, hematuria and the presence of LUTS for predicting urine culture results was calculated. RESULTS: Thirty-two percent of cats (n = 31) had a positive urine culture. Of these, 28 (90%) had bacteriuria, 21 (68%) had pyuria, 13 (42%) had hematuria and 10 (32%) had LUTS. Of the 42 cats without hematuria or pyuria, seven had a positive urine culture (17%). Bacteriuria had a high sensitivity (90%) and specificity (92%) for predicting urine culture bacterial growth. The absence of bacteriuria had a high negative predictive value for no bacterial growth (95%). The odds of a positive urine culture were increased with bacteriuria (odds ratio [OR] 114, 95% confidence interval [CI] 29-621; P <0.001), pyuria (OR 21, 95% CI 7-70; P <0.001) and LUTS (OR 5, 95% CI 1.7-16; P = 0.004). Hematuria was not associated with a positive culture (sensitivity 42%, specificity 52%). CONCLUSIONS AND RELEVANCE: Microscopic bacteriuria and pyuria on urine sediment evaluation and LUTS can be helpful for predicting bacterial culture results in cats with AKI and in settings where submitting a urine culture may not be financially feasible.

Coagulation status, fibrinolysis, and platelet dynamics in dogs with chronic inflammatory enteropathy.

BACKGROUND: Coagulation status is poorly understood in dogs with chronic inflammatory enteropathy (CIE). Fibrinolytic activity and platelet dynamics have not been evaluated in CIE dogs. OBJECTIVES: To assess coagulation status and fibrinolysis in normoalbuminemic CIE dogs (CIE-N) and CIE dogs with protein-losing enteropathy (CIE-PLE) compared to healthy controls (HC). To evaluate thromboelastography (TEG) variable differences between groups and for correlations with clinicopathologic data. To report platelet dynamics in CIE dogs. ANIMALS: Twenty-five client-owned dogs with CIE (n = 16 CIE-N; n = 9 CIE-PLE); 14 HC beagle dogs. METHODS: All dogs had tissue factor + tissue plasminogen activator TEG. Nine of 25 CIE dogs had whole blood impedance platelet aggregometry. The TEG variables and coagulation data were compared between all CIE vs HC dogs, CIE-N dogs vs HC, and CIE-PLE dogs vs HC. Clinicopathologic and coagulation data were available for CIE dogs and assessed for correlation to TEG variables. RESULTS: Dogs with CIE had higher maximum amplitude (MA; P < .001), longer clot lysis times (CLTs; P < .001), lower % lysis after 30 minutes (LY30; P < .001), and % lysis after 60 minutes (LY60; P < .001) compared to HC, suggesting hypercoagulability and hypofibrinolysis. When separated out, both CIE-N and CIE-PLE dogs had higher MA, longer CLT, and lower LY30 and LY60 compared to HC. Serum albumin and 25-hydroxyvitamin D (25[OH]D) concentrations, and plasma antithrombin and fibrinogen concentrations moderately correlated with MA. CONCLUSIONS AND CLINICAL IMPORTANCE: Normoalbuminemic and hypoalbuminemic CIE dogs were considered hypercoagulable based on TEG compared to HC. Some CIE dogs displayed hypofibrinolytic phenotypes on TEG.

Evaluation of platelet function in cats with and without kidney disease: a pilot study.

OBJECTIVES: The aims of this study were to determine if stable chronic kidney disease (CKD) cats and uremic crisis cats have altered platelet function, and to determine the prevalence of positive fecal occult blood in CKD cats. METHODS: Platelet function in normal cats, clinically stable International Renal Interest Society (IRIS) stage 2-4 CKD cats and CKD cats experiencing a uremic crisis were evaluated using impedance aggregometry. Area under the curve (AUC) at 6 mins was calculated for saline, adenosine diphosphate (AUCADP) and arachidonic acid (AUCASPI). The AUC in addition to hematocrit, platelet count and mean platelet volume (MPV) were compared between groups using the Kruskal-Wallis test followed by Dunn's post-hoc analysis. Guaiac fecal occult blood tests were performed on fecal samples and results were compared between groups using a χ2 for trend test. RESULTS: AUCADP (P = 0.04) and AUCASPI (P = 0.05) were significantly higher in uremic crisis cats compared with normal cats at 6 mins. Hematocrit was significantly higher in normal cats when compared with IRIS stage 3 and 4 (P = 0.002) and uremic crisis (P = 0.0008) cats, with no difference among groups for platelet count or MPV. The proportion of cats with positive fecal occult blood samples was significantly different between groups (P = 0.0017); 50% uremic crisis cats, 33% IRIS stage 3 and 4 cats, and 10% IRIS stage 2 cats were positive, while no normal cats were positive. The proportion of cats with platelet clumping was significantly different between groups (P = 0.03). CONCLUSIONS AND RELEVANCE: Platelet hyper-reactivity may be occurring in CKD cats experiencing a uremic crisis. The etiology of positive fecal occult blood samples in CKD cats is unclear and did not appear to be related to decreased platelet function as measured in this study and requires further investigation.

Ehrlichia canis in dogs experimentally infected, treated, and then immune suppressed during the acute or subclinical phases.

BACKGROUND: Concerns for recrudescence of Ehrlichia canis infection arise when immunosuppressive drugs are used to treat immune-mediated diseases in dogs previously infected with E. canis. OBJECTIVES: Determine whether administration of prednisolone and cyclosporine would reactivate E. canis infection in dogs previously treated with doxycycline during the acute or subclinical phases. ANIMALS: Seven beagles previously experimentally infected with E. canis and administered doxycycline for 4 weeks were included. Three of the 7 dogs were incidentally concurrently infected with Anaplasma platys and Babesia vogeli and were administered 2 doses of imidocarb 2 weeks apart before enrollment in the current study. METHODS: Experimental study. Each dog was administered prednisolone and cyclosporine for 6 weeks. Clinical signs, complete blood cell count (CBC), polymerase chain reaction (PCR) assays for E. canis, A. platys, and B. vogeli DNA in blood, E. canis indirect fluorescent antibodies (IFA) titers, and flow cytometry for antiplatelet antibodies were monitored. RESULTS: All dogs completed the immunosuppressive protocol. No evidence for recrudescence of E. canis, A. platys, or B. vogeli were detected based on clinical signs or results of CBC, PCR, IFA, and flow cytometry for antiplatelet antibodies. E. canis IFA titers were negative in 5/7 dogs at the end of immunosuppressive protocol and were negative 6 months after the protocol in 5/5 dogs available for testing. CONCLUSIONS AND CLINICAL IMPORTANCE: Dogs administered with a 4-week course of doxycycline with or without imidocarb failed to show evidence of activation of E. canis infection after administration of a commonly used immune suppressive protocol.

Detection and dynamics of anti-platelet antibodies in thrombocytopenic dogs with and without idiopathic immune thrombocytopenia.

BACKGROUND: Antiplatelet antibodies are detected in multiple diseases including primary immune thrombocytopenia (ITP). Dynamics of how these antibodies change over time in ITP is unknown in dogs. HYPOTHESIS/OBJECTIVES: Antiplatelet antibodies (APA) will be detected in thrombocytopenic dogs with multiple etiologies and dynamics of APA in dogs with ITP can be used to evaluate response to treatment and relapse. Determine APA at the time of diagnosis in thrombocytopenic dogs and serially in primary ITP dogs. ANIMALS: Seventy-nine thrombocytopenic dogs and 28 primary ITP dogs. METHODS: Direct flow cytometry was performed in thrombocytopenic dogs at initial evaluation and serially in suspected primary ITP dogs. In primary ITP dogs, a 2-tailed Fisher's exact test was performed comparing survival to discharge between dogs with and without melena and to relate response to treatment and relapse to changes in APA and platelet count (repeated measures analysis, Spearman correlation). RESULTS: Twenty percent (16/79) of thrombocytopenic non-ITP dogs with infectious, neoplastic, or other diseases and all primary ITP dogs were positive for APA. Melena at initial evaluation was associated with decreased survival to discharge (odds ratio 0.06; P = .01). Persistence of APA was not associated with response to treatment, but recurrence of antibodies was associated with relapse (odds ratio 205.0; P < .01). There was no difference in percentage of APA or platelet count at initial diagnosis between dogs that did or did not respond to treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: Serial monitoring of APA in dogs with primary ITP appeared beneficial for determining relapse of disease.

Platelet aggregometry testing during aspirin or clopidogrel treatment and measurement of clopidogrel metabolite concentrations in dogs with protein-losing nephropathy.

BACKGROUND: Dogs with protein-losing nephropathy (PLN) are treated with antiplatelet drugs for thromboprophylaxis but no standardized method exists to measure drug response. It is also unknown if clopidogrel metabolite concentrations [CM] differ between healthy and PLN dogs. OBJECTIVES: Assess response to aspirin or clopidogrel in PLN dogs using platelet aggregometry (PA) and compare [CM] between healthy and PLN dogs. ANIMALS: Six healthy and 14 PLN dogs. METHODS: Platelet aggregometry using adenosine diphosphate (ADP), arachidonic acid (AA), and saline was performed in healthy dogs at baseline and 1-week postclopidogrel administration to identify responders or nonresponders. A decrease of ≥60% for ADP or ≥30% for AA at 1 or 3 hours postpill was used to define a responder. At 1 and 3 hours postclopidogrel, [CM] and PA were measured in healthy and PLN dogs. Platelet aggregometry was performed in PLN dogs at baseline, 1, 6, and 12 weeks after clopidogrel or aspirin administration. RESULTS: In PLN dogs receiving clopidogrel, PA differed from baseline at all time points for ADP but not for AA at any time point. Most dogs responded at 1 or both time points except for 1 dog that showed no response. For PLN dogs receiving aspirin, no differences from baseline were observed at any time point for either ADP or AA. No differences in [CM] were found at either time point between healthy and PLN dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: Platelet aggregometry may represent an objective method to evaluate response to clopidogrel or aspirin treatment and PLN dogs appear to metabolize clopidogrel similarly to healthy dogs.

Effects of storage conditions on two platelet agonists for whole blood impedance platelet aggregometry in dogs.

BACKGROUND: Whole blood impedance platelet aggregometry (Multiplate-) can be performed with different agonists to evaluate platelet function. Although the manufacturer recommends disposal of stored reagents after 1 month at -20°C or 24 hours at 4°C, reagent integrity after reconstitution under different storage conditions is unknown. If reagent integrity is stable for longer periods, assay costs could decrease dramatically. OBJECTIVES: This study aimed to determine the stability of reconstituted arachidonic acid (AA) and adenosine diphosphate (ADP) platelet agonists stored at -20°C and -80°C for up to 6 months. METHODS: Aliquots of reconstituted AA and ADP were stored at -20°C and -80°C each month for a total of 6 months. Six healthy staff-owned dogs were enrolled in the study. A physical examination, CBC, diagnostic panel, urinalysis, and baseline platelet aggregometry assessment was performed on all of the dogs. Platelet aggregometry was performed using fresh and stored aliquots of AA and ADP reagents on the same day. The area under the curve (AUC) was recorded from each platelet aggregometry analysis. Repeated measures (RM) analysis (one-way ANOVA) was performed and subsequent time points (1, 2, 3, 4, 5, and 6 months) were compared with fresh AUC results. RESULTS: All dogs were clinically healthy, and all diagnostic tests were normal. There were no differences in AUC obtained from fresh samples at any time point or either temperature for AA or ADP. CONCLUSIONS: Whole blood impedance platelet aggregometry reagents, AA and ADP, were stable for up to 6 months when stored at -20°C or -80°C, obviating the need to discard viable reagents, and decreasing assay costs.

Validation of a clinically applicable flow cytometric assay for the detection of immunoglobulin associated platelets in dogs.

Thrombocytopenia is commonly encountered in veterinary practice when evaluating canine patients. It can occur in infectious, neoplastic, inflammatory, toxic, and immune-mediated conditions. Elucidating the underlying cause for thrombocytopenia can therefore represent a challenge to veterinary practitioners. Additionally, determination of whether an immune process could be contributing to a patient's thrombocytopenia is important for refining differentials and enhancing understanding of a particular disease process. A possible candidate test for the development of a clinically applicable assay in dogs is flow cytometry. Therefore, the purpose of this study was to develop a clinically applicable direct and indirect flow cytometric assay for the detection of canine immunoglobulin associated platelets. Direct and indirect flow cytometry was performed in nine healthy beagles and twelve client-owned thrombocytopenic dogs at four time points: fresh and after 24, 48, and 72 h of storage at 4 °C. For healthy dogs, there was no significant difference between fresh and 24 and 48 h samples but there was a significant difference between fresh and 72 h samples. There was no significant difference between fresh and 24, 48, or 72 h samples in the thrombocytopenic dogs. A cut-off value of ≤ 10% antibody binding was defined to differentiate negative and positive classifications and was determined by serial direct flow evaluations in a healthy dog. Based on this cut-off value, healthy and thrombocytopenic dogs were consistently categorized at every time point. The average intra-assay coefficient of variation for the thrombocytopenic dogs was 4.32%. The indirect flow cytometric methods evaluated herein did not provide reliable or repeatable results in healthy or thrombocytopenic dogs. Direct flow cytometry represents a potentially clinically useful test for the detection of immunoglobulin associated platelets in dogs that can be processed and evaluated within a realistic amount of time which would allow for testing in a larger number of patients. Based on the findings from this study using our protocols, indirect flow cytometry was not clinically applicable in dogs.

Variability of tissue factor-activated thromboelastography and whole blood impedance platelet aggregometry in healthy dogs.

OBJECTIVE: To assess interindividual (CVG ) and intraindividual (CVI ) variability over time for results of thromboelastography (TEG) and whole-blood impedance platelet aggregometry in healthy dogs. ANIMALS: Six healthy Beagle dogs. MEASUREMENTS AND MAIN RESULTS: Tissue factor (TF)-activated TEG and adenosine diphosphate (ADP) and arachidonic acid (AA)-induced whole blood impedance platelet aggregometry were performed at 3 different time points (days 1, 4, and 6). In addition, platelet count, hematocrit, and plasma fibrinogen concentrations were recorded each study day. Activated partial thromboplastin time, one-stage prothrombin time, antithrombin activity, and D-dimer concentrations were measured on the first day of the study. For TEG, the variables reaction time (R), clotting time (K), rate of clot formation (α), and maximum amplitude (MA) were recorded. For platelet aggregometry, the areas under the curve for ADP (AUCADP )- and AA (AUCAA )-induced aggregation were measured. The CVI was lower than the CVG over time for MA, AUCADP , and AUCAA ; however, the CVI was higher than the CVG for the TEG variables R, K, and α. There were no statistical differences in the platelet count, hematocrit, and fibrinogen measurements over time. CONCLUSIONS: In healthy dogs, a subject-based reference interval for ADP- and AA-induced platelet aggregometry and the TEG variable MA provide a more sensitive method to detect changes. However, due to the high CVI , population-based reference intervals may be more appropriate for interpretation of the TEG variables R, K, and α.

Characteristics of hemostasis during experimental Ehrlichia canis infection.

BACKGROUND: Ehrlichia canis infection in dogs can cause thrombocytopenia and clinical evidence of bleeding. It is unknown why some dogs show signs of bleeding whereas others do not despite clinically relevant thrombocytopenia. HYPOTHESIS/OBJECTIVES: Activated platelets, decreased fibrinolysis or both mitigate bleeding tendency. Assess standard hemostatic variables, platelet dynamics, and specialized coagulation testing in dogs experimentally infected with E. canis to evaluate this clinical discrepancy. ANIMALS: Four healthy laboratory beagles. METHODS: Dogs were given blood infected with E. canis IV. Platelet indices of activation, platelet aggregometry, antiplatelet antibodies (percent IgG), complete coagulation panel, and thromboelastography (TEG) were measured before inoculation and on weeks 1-8. Dogs were treated with doxycycline at approximately 5 mg/kg PO q12h between weeks 3 and 4 (day 24). For each variable, 1-way repeated measures analysis (1-way ANOVA) with post-hoc analysis was performed with statistical significance set at P < .05. RESULTS: Dogs had significantly lower platelet counts, evidence of activated platelets, and antiplatelet antibodies during E. canis infection. Dogs also appeared hypercoagulable and hypofibrinolytic using TEG as compared with baseline, changes that persisted for variable amounts of time after doxycycline administration. No overt signs of bleeding were noted during the study. CONCLUSIONS AND CLINICAL IMPORTANCE: Activated platelets and a hypercoagulable, hypofibrinolytic state could explain the lack of a bleeding phenotype in some dogs despite clinically relevant thrombocytopenia. Findings from our pilot study indicate that additional studies are warranted.