The acute effects of a protein-rich meal on the urinary corticoid:creatinine ratio in healthy dogs.

The objective of this prospective crossover study was to investigate the effects of a high-protein diet on canine urinary corticoid-to-creatinine ratio (UCCR). The hypothesis was that meal-induced hypercortisolism is, as has been shown in humans, a predictable and consistent finding in healthy dogs. Eight clinic-owned beagles were randomly assigned to one of two groups. The allocation to the groups defined the sequence of a protein-enriched meal (meal A) or no meal on the first and second days, whereas on the third day all dogs again received an identical meal (meal B) to test reproducibility. Urinary corticoids were measured using a solid-phase, competitive CLIA on unextracted urine. Contrary to our expectations, consistent incremental responses of the UCCR were not observed (meal A vs. no meal [anova]: absolute increase, F = 2.546, p = 0.162; relative increase, F = 4.084, p = 0.09; AUC(UCCR) , F = 0.279, p = 0.616). Nevertheless, the robust increases in two dogs above 60% of baseline suggest that the collection of urine prior to feeding likely increases the specificity of the UCCR to discriminate between dogs with and without hypercortisolism.

Glucose concentration in capillary blood of dairy cows obtained by a minimally invasive lancet technique and determined with three different hand-held devices.

BACKGROUND: Dairy cows have a massive demand for glucose at the onset of lactation. A poor adaption to this period leads to an excessive negative energy balance with an increased risk for ketosis and impaired animal health and production. Besides the measurement of ketones, analysing the glucose concentration in blood is reported as helpful instrument for diagnosis and differentiation of ketosis. Monitoring metabolic parameters requires multiple blood sampling. In other species, new blood sampling techniques have been introduced in which small amounts of blood are rapidly analysed using electronic hand-held devices. The objective of this study was to evaluate the suitability of capillary blood for blood glucose measurement in dairy cows using the hand-held devices FreeStyle Precision (FSP, Abbott), GlucoMen LX Plus (GLX, A. Menarini) and the WellionVet GLUCO CALEA, (WGC, MED TRUST). In total, 240 capillary blood samples were obtained from dry and fresh lactating Holstein-Friesian cows. Blood was collected from the skin of the exterior vulva by using a lancet. For method comparison, additional blood samples were taken from a coccygeal vessel and analyzed in a laboratory. Glucose concentrations measured by a standard laboratory method were defined as the criterion standard. RESULTS: The Pearson correlation coefficients between the glucose concentrations analyzed in capillary blood with the devices and the reference were 73% for the FSP, 81% for the GLX and 41% for the WGC. Bland-Altman plots showed biases of -18.8 mg/dL for the FSP, -11.2 mg/dL for the GLX and +20.82 mg/dL for the WGC. The optimized threshold determined by a Receiver Operating Characteristics analysis to detect hyperglycemia using the FSP was 43 mg/dL with a sensitivity (Se) and specificity (Sp) of 76 and 80%. Using the GLX and WGC optimized thresholds were 49 mg/dL (Se = 92%, Sp = 85%) and 95 mg/dL (Se = 39%, Sp = 92%). CONCLUSIONS: The results of this study demonstrate good performance characteristics for the GLX and moderate for the FSP to detect hyperglycemia in dairy cows using capillary blood. With the study settings, the WGC was not suitable for determination of glucose concentrations.

Measurement of ß-hydroxybutyrate in capillary blood obtained from an ear to detect hyperketonemia in dairy cows by using an electronic handheld device.

The primary objective of the present study was to test whether capillary blood obtained by puncturing the skin of an ear with a minimal invasive lancet technique is able to detect hyperketonemia (HYK) in dairy cows. Furthermore, test characteristics of a new available handheld device, the FreeStyle Precision Neo (FSP-Neo, Abbott GmbH & Co. KG, Wiesbaden, Germany) for determination of ß-hydroxybutyrate (BHB) concentrations in bovine blood were evaluated by comparing the measurements with a laboratory reference. The BHB concentration was determined with the FSP-Neo device in 720 capillary blood samples from 3 different sampling sites (left, right ear, and repeated measurement) and in 240 samples from a coccygeal vessel. The concentration of BHB in serum harvested from the coccygeal blood samples was analyzed at the laboratory and was used as reference. The Spearman correlation coefficient (ρs) between the BHB concentrations in capillary blood measured with the handheld device and the reference test was between 0.76 and 0.81. Using capillary blood, the mean ± standard deviation BHB difference compared with the reference test was 0.20±0.47 mmol/L for all 3 sampling locations at the ears. The receiver operating characteristic analyses for the FSP-Neo device resulted in an optimized threshold for the detection of subclinical ketosis (SCK) in capillary blood of 1.3 mmol/L (left and right ear) and 1.2 mmol/L (repeated measurements). Applying these adjusted threshold sensitivities (Se) for all 3 capillary sampling sites at the ear were 100%, and specificities (Sp) ranged between 93 and 94%. Hence, we conclude that all sampling locations were suitable to identify cows suffering from SCK. The reference test compared with BHB measurements in coccygeal blood resulted in a ρs of 0.92 with a mean ± standard deviation of 0.02±0.21 mmol/L. The receiver operating characteristic analyses for the FSP-Neo device resulted in an optimized threshold for the detection of SCK in coccygeal blood of 1.1 mmol/L, with a corresponding Se and Sp of 100 and 95%, respectively. Because capillary blood is easily achievable from an ear, particularly if animals are fixed in headlocks for routine checkups, this technique is considered as an additional minimally invasive method for the identification of dairy cows suffering from HYK.

Suitability of capillary blood obtained by a minimally invasive lancet technique to detect subclinical ketosis in dairy cows by using 3 different electronic hand-held devices.

The objective of this study was to evaluate the suitability of capillary blood obtained by a minimally invasive lancet technique to detect subclinical ketosis in 49 prepartum and 191 postpartum Holstein-Friesian cows using 3 different electronic hand-held devices [FreeStyle Precision (FSP, Abbott), GlucoMen LX Plus (GLX, A. Menarini), NovaVet (NOV, Nova Biomedical)]. The ß-hydroxybutyrate (BHBA) concentration in serum harvested from coccygeal blood samples was analyzed in a laboratory and used as a reference value. Capillary samples were obtained from the skin of the exterior vulva by using 1 of 3 different lancets. In all samples, the concentration of BHBA was immediately analyzed with all 3 hand-held devices used in random order. All lancets used in the study were eligible for capillary blood collection but differed in the total number of incisions needed. Spearman correlation coefficients between the BHBA concentrations in capillary blood and the reference test were highly significant with 83% for the FSP, 73% for the NOV, and 63% for the GLX. Using capillary blood, the FSP overestimated the mean BHBA concentration compared with the reference test (+0.08 mmol/L), whereas the GLX and NOV underestimated the mean concentration (-0.07 and -0.01 mmol/L). When a BHBA concentration of 1.2 mmol/L in serum was used to define subclinical ketosis, the corresponding analyses of receiver operating characteristics resulted in optimized thresholds for capillary blood of 1.1 mmol/L for the NOV and GLX devices, and of 1.0 mmol/L for the FSP. Based on these thresholds, sensitivities (Se) and specificities (Sp) were 89 and 84% for the NOV, 80 and 89% for the GLX, and 100 and 76% for the FSP. Based on a serum BHBA concentration of 1.4 mmol/L, analyses of receiver operating characteristics resulted in optimized cut-offs of 1.4 mmol/L for the FSP (Se 100%, Sp 92%), 1.3 mmol/L for the NOV (Se 80%, Sp 95%), and 1.1 mmol/L (Se 90%, Sp 85%) for the GLX. Using these optimized thresholds for the specific hand-held meters, no significant differences between the devices in Se and Sp to detect subclinical ketosis in coccygeal blood were observed. Calculated test characteristics for analyzing capillary blood using the hand-held devices were numerically smaller compared with blood obtained from a coccygeal vessel, but overlapping confidence intervals indicate no statistical difference between the origin of the sample. Hence, this procedure seems to be suitable for ketosis monitoring in dairy cows, but further validation with more data from different farms is recommended.

Thresholds of whole-blood ß-hydroxybutyrate and glucose concentrations measured with an electronic hand-held device to identify ovine hyperketonemia.

Metabolic disorders, especially hyperketonemia, are very common in dairy sheep. The whole-blood concentrations of ß-hydroxybutyrate (BHBA) and glucose can be determined by commercially available electronic hand-held devices, which are used in human medicine and for the detection of ketosis in dairy cows. The aim of this study was to evaluate the suitability of the hand-held device Precision Xceed (PX; Abbott Diabetes Care Inc., Abbott Park, IL) to detect hyperketonemia in ewes. An additional objective of this study was to evaluate the agreement between samples obtained by minimal invasive venipuncture of an ear vein and measurements of whole-blood samples from the jugular vein (vena jugularis, v. jug.). Blood samples taken from the v. jug. were collected from 358 ewes on 4 different farms. These samples and a blood drop obtained from an ear vein were analyzed simultaneously on farm with the PX. For method comparison, the samples obtained from the v. jug. were also analyzed by standard methods, which served as the gold standard at the Central Laboratory of the University of Veterinary Medicine Vienna, Austria. The correlation coefficients between the serum BHBA concentration and the concentrations measured with the hand-held meter in the whole blood from an ear vein and the v. jug. were 0.94 and 0.96, respectively. The correlation coefficients of plasma and whole-blood glucose concentration were 0.68 for the v. jug. and 0.47 for the ear vein. The mean glucose concentration was significantly lower in animals classified as hyperketonemic (BHBA ≥ 1.6 mmol/L) compared with healthy ewes. Whole-blood concentrations of BHBA and glucose measured with the PX from v. jug. showed a constant negative bias of 0.15 mmol/L and 8.4 mg/dL, respectively. Hence, a receiver operating characteristic analysis was performed to determine thresholds for the PX to detect hyperketonemia in ewes. This resulted in thresholds for moderate ketosis of BHBA concentrations of 0.7 mmol/L in blood from an ear vein and the v. jug. Cutoffs of 1.0 mmol/L (ear vein) and 1.1 mmol/L (v. jug.) BHBA were determined to detect animals at greater risk to develop severe hyperketonemia. Applying these thresholds, excellent test characteristics, with sensitivities of 1.00 for both samples and specificities of 0.98 for the ear vein and 0.97 for the v. jug. were determined. These results demonstrate that the PX is a useful tool for detection of hyperketonemia in ewes.

Short communication: evaluation of an automated in-house hematology analyzer for bovine blood.

The objective of this study was to evaluate the suitability of the V-Sight hematology analyzer (A. Menarini Pharma GmbH, Vienna, Austria) for bovine blood by a comparison with a reference device (Advia 2120i, Siemens AG, Erlangen, Germany). In total, 97 blood samples were obtained from 75 dairy cows. Analyzed parameters included counts of white blood cells (WBC), lymphocytes, monocytes, granulocytes, red blood cells (RBC), and platelets (PLT), as well as hemoglobin concentration (HGB), hematocrit (HCT), mean corpuscular volume, mean corpuscular hemoglobin (MCH), MCH concentration (MCHC), mean platelet volume (MPV), and plateletcrit (PCT). Based on Passing-Bablok regression, the V-Sight provided accurate and precise results for MCH and MCHC only. The PCT results were comparable to the reference method, but precision was inconclusive. Significant proportional differences were detected for monocytes, granulocytes, HCT, and PLT. For all other analytes, significant proportional and systemic differences were observed. The WBC and lymphocyte results from the V-Sight were characterized by poor accuracy, poor precision, and a high number of false positive outliers. Bland-Altman analysis indicated negative biases for all WBC parameters, the erythrocyte indices, and PLT. Positive biases were observed for RBC, HGB, HCT, MPV, and PCT. Correlation coefficients of >0.9 between the V-Sight and the reference method were found only for RBC, HGB, HCT, and MPV. Intraassay precision of the V-Sight analyzer was acceptable (coefficient of variation <5%) for granulocytes, the erythrocyte indices, and MPV. It was unacceptable (coefficient of variation ≥5%) for WBC, lymphocytes, monocytes, as well as RBC, and inconclusive for HGB, HCT, PLT, and PCT. Sensitivity was high for all RBC counts and indices as well as PLT, but low for monocytes, granulocytes, and MPV. Specificity was high for monocytes and granulocytes, but low for RBC, HCT, MCH, and MCHC. With accurate and precise results for only 2 out of 13 parameters, the V-Sight cannot be recommended for analysis of bovine blood.

Evaluation of 2 electronic handheld devices for diagnosis of ketonemia and glycemia in dairy goats.

The objective of this study was to evaluate the suitability of the electronic handheld devices FreeStyle Precision (FSP; Abbott Germany, Wiesbaden, Germany) and GlucoMen LX Plus (GML; A. Menarini GmbH, Vienna, Austria) for the measurement of ß-hydroxybutyrate (BHBA) in whole blood in dairy goats. Additionally, glucose concentration was analyzed with the FSP device. For method comparison, the samples were also analyzed in the laboratory by standard methods, which served as the gold standard in our study. A further objective was to evaluate the agreement between samples obtained by minimal invasive venipuncture of an ear vein and measurements of whole blood samples from the jugular vein (vena jugularis). In total, 173 blood sample pairs collected from 28 goats were obtained from an ear vein and from the jugular vein. The Spearman correlation coefficients (rsp) for BHBA concentrations determined with the FSP or GML and the gold standard were 0.95 and 0.85 for the ear vein and 0.98 and 0.88 for the jugular vein, respectively. Bland-Altman plots of differences showed a positive bias of 0.12 (ear vein) and 0.21 (jugular vein) when determination was performed with the FSP and a negative bias of 0.21 (ear vein) and 0.24 (jugular vein) when using the GML. For the FSP, applying the adjusted thresholds determined by ROC analysis of 0.9 (ear vein) and 1.0 mmol/L (jugular vein) allowed to distinguish between healthy goats and animals with hyperketonemia with sensitivities (Se) and specificities (Sp) for samples from the ear vein of 0.98 and 0.85, and from the jugular vein of 0.99 and 0.94, respectively. For the GML, adjusted thresholds were 0.5 mmol/L for the ear vein (Se=0.94, Sp=0.75) and 0.6 mmol/L for the jugular vein (Se=0.88, Sp=0.91). Repeated analyses of defined whole blood samples showed average inter- and intraassay coefficients of variation of 6.6 and 7.3% for FSP, and of 35.6 and 35.4% for GML, respectively. Test characteristics for determining glucose concentration with the FSP compared with the gold standard were poor (ear vein: rsp=0.41; jugular vein: rsp=0.51), with low validity to distinguish between hypo- and normoglycemia (Se=0.71, Sp=0.66). The present study showed good test characteristics for the FSP and moderate for the GML device for determining BHBA concentrations in whole blood. Additionally the results demonstrate the suitability of testing BHBA concentration in a blood drop obtained from an ear vein. Based on the results of this study, the FSP device is not suitable to differentiate normo- from hypoglycemia in goats.

Comparison of 2 electronic cowside tests to detect subclinical ketosis in dairy cows and the influence of the temperature and type of blood sample on the test results.

The objective of this study was to determine the suitability of 2 electronic hand-held devices [FreeStyle Precision (FSP), Abbott GmbH & Co. KG, Wiesbaden, Germany and GlucoMen LX Plus (GLX), A. Menarini GmbH, Vienna, Austria] for measuring ß-hydroxybutyrate (BHBA) in dairy cows. Three experiments were conducted to evaluate (1) the diagnostic performance of the devices, (2) the effect of the type of blood sample, and (3) the influence of the ambient temperature on the determined results. A total of 415 blood samples from lactating Holstein and Simmental cows were collected and analyzed with both devices (whole blood) and in a laboratory (serum). Correlation coefficients between whole-blood and serum BHBA concentrations were highly significant, with 94% for the FSP and 80% for the GLX device. Based on thresholds for subclinical ketosis of 1.2 and 1.4 mmol of BHBA/L, results obtained with the hand-held devices were evaluated by receiver operating characteristics analyses. This resulted in adjusted thresholds of 1.2 and 1.4 mmol/L for the FSP and 1.1 and 1.3 mmol/L for the GLX device. Applying these thresholds, sensitivities were 98 and 100% for the FSP and 80 and 86% for the GLX device, respectively. Corresponding specificities were 90 and 97% for the FSP and 87 and 96% for the GLX device, respectively. Additionally, concentrations of BHBA were tested with both devices in whole blood, EDTA-added whole blood, and in their resulting serum and plasma, collected from 65 animals. Determined BHBA concentrations were similar within each device for whole and EDTA-added blood, and in serum and plasma, but differed between whole blood and serum and between EDTA-added blood and plasma. Blood samples with low (0.4 mmol/L), medium (1.1 mmol/L), and high (1.6 mmol/L) BHBA concentrations were stored between +5 to +32°C and analyzed repeatedly at temperature levels differing by 4°C. Additionally, devices and test strips were stored at equal conditions and used for measurement procedures. Storage temperature of the devices and test strips did not influence the differences between the results of the laboratory and the devices, whereas the temperature of the blood samples caused significant differences. Although the level of agreement between the laboratory and the GLX device was lower than for the laboratory and the FSP device, both devices are useful tools for monitoring subclinical ketosis in dairy cows. Due to their effects on the determined results, the type and temperature of the tested sample should be considered.

Measurement of alkaline phosphatase in canine seminal plasma--an update.

In dogs, diagnosis of incomplete ejaculation and azoospermia can be made by measuring the activity of the enzyme alkaline phosphatase (AP) in seminal plasma. However, even though upper cut-off value of 5000 IU/l is given in the literature, results by different assays may vary considerably. Furthermore, no data exist concerning the stability of the enzyme during storage of frozen seminal plasma, and no recommendations for pre-analytic dilutions can be found. During the present study, we compared results from a conventional large scale wet chemistry analyzer to a widely used dry chemistry point of care system (POC) and established a best practice for pre-analytical dilutions. Furthermore, stability of enzyme activities in seminal plasma during storage at -18 °C for 24 h was evaluated. The average activity of AP in the 2nd fraction of normal ejaculates measured by Reflotron® was 107,328 IU/l. After 24 h of frozen storage, activities did not differ significantly (96,844 IU / l, p > 0.05). Fresh and frozen samples were analysed in parallel by the POC and conventional chemistry analyser, and the results compared that did not reveal a significant difference (p > 0.05). A dilution of seminal plasma with physiologic saline 1:100 prior to analysis was sufficient for the qualitative information whether AP activity is below or above 5000 IU/l. Present data show that AP measurement by a POC dry chemistry system is sufficiently accurate in diluted seminal plasma for the diagnosis of azoospermia and that seminal plasma can be stored frozen for 24h before analysis.

Bone and cellular immune system of multiparous sows are insensitive to ovariectomy and nutritive calcium shortage.

Research in osteoporosis, which is a complex systemic disease, demands suitable large animal models. In pigs, most research has been done in growing minipigs, which probably are not ideal models for postmenopausal osteoporosis. Therefore, our aim was to analyze the effects of ovariectomy (OVX) and nutritive calcium shortage on multiparous Large White sows. 32 animals were randomly assigned to 4 groups in a cross design with OVX vs. sham and physiological calcium supplementation (0.75% calcium) vs. dietary calcium shortage (0.3% calcium). The observation period was 10 months with blood sampling every 2 months for hematological, immunological, and biochemical bone marker measurements. At the termination of the experiment, animals were sacrificed. Samples of trabecular bone of distal radius, proximal tibia, and sixth lumbar vertebra were subjected to micro-computed tomography imaging and ashed afterwards. Dual X-ray absorptiometry scans of the proximal femora were performed with prepared bones being placed in a water bath for mimicking soft tissue. Analyses of bone marker and cytokine profile kinetics, distribution of leukocyte subpopulations, and morphometrical and densitometrical analyses showed no evidence of any impact of OVX or calcium shortage. In conclusion, the skeleton of adult sows of a conventional breed is seemingly protected from effects of OVX and calcium shortage.

[Diagnostic efficacy of plasma ACTH-measurement by a chemiluminometric assay in canine hyperadrenocorticism]. / Klinische Anwendung einer chemiluminometrischen ACTH-Bestimmung beim caninen Hyperadrenokortizismus.

This retrospective study was performed to investigate the diagnostic efficacy of the chemiluminometric ACTH-measurement to differentiate between pituitary and adrenal dependent hyperadrenocorticism (HAC) in dogs. 49 dogs with pituitary HAC, 10 dogs with adrenal HAC and 1 dog with a combination of both pathologies were included. Dogs with HAC like symptoms, where HAC had been ruled out, served as controls (n = 18). All dogs with adrenal HAC, as well as 9 dogs with pituitary HAC had an ACTH concentration below the detection limit of 2.2 pmol/l (10 pg/dl) plasma. Using 2.2 pmol/l as a cut-off the sensitivity and specificity to diagnose pituitary HAC was 0.82 (95 % CI 0.686 - 0.914) and 1 (95 % CI 0.692 - 1), respectively. With the help of the chemiluminometric assay, a correct classification was possible in 85 % of patients with HAC. As an ACTH-concentration below the detection limit was found in dogs with adrenal as well as pituitary HAC, additional discriminatory tests are necessary in these cases.

Effect of altrenogest-treatment of mares in late gestation on adrenocortical function, blood count and plasma electrolytes in their foals.

REASONS FOR PERFORMING STUDY: Mares with compromised pregnancies are often treated with altrenogest to prevent abortion. However, there is only limited information about effects on the foal when altrenogest treatment is continued during final maturation of the fetus. OBJECTIVES: To determine effects of altrenogest treatment during late gestation in mares on maturity, haematology changes, adrenocortical function and serum electrolytes in their newborn foals. METHODS: Six mares were treated with altrenogest (0.088 mg/kg bwt) once daily from Day 280 of pregnancy until foaling and 7 mares served as controls. RESULTS: Foals born to altrenogest-treated mares had a significantly lower neutrophil/lymphocyte ratio on the first day after birth than control foals (P<0.05). Basal plasma cortisol concentrations immediately after birth were higher in foals of altrenogest-treated mares than in control foals (P<0.05). Cortisol release in response to exogenous adrenocorticotropic hormone (ACTH)--except for higher values 15 min after ACTH injection in foals of altrenogest-treated mares on Day 1--revealed no differences in adrenocortical function between the groups of foals. Plasma potassium concentration in foals from altrenogest-treated mares compared to control foals was significantly lower immediately after birth (P<0.05) and plasma ionised calcium concentration was significantly lower 3 h after birth (P = 0.01). CONCLUSIONS AND POTENTIAL RELEVANCE: Altrenogest treatment of pregnant mares prolonged labour had no major effects on adrenocortical function in foals. A reduced neutrophil/lymphocyte ratio in these foals may suggest either immunomodulatory effects of altrenogest or dysmaturity of the foals.

[Invasive arterial blood pressure measurement using an aneroid pressure system in cattle]. / Invasive arterielle Blutdruckmessung mittels eines aneroiden Systems beim Rind.

The aim of this study was to compare the results of invasive arterial blood pressure measurement using an electronic pressure transducer (EPT) or an aneroid pressure system (APS) in cattle. A catheter was placed in the auricular artery of 11 adult cattle and connected to a pressure transducer via pressure line. The aneroid system was connected to the same catheter using a three-way stop-cock in the pressure line. On five occasions three consecutive measurements were performed with the APS. The mean blood pressure values of the EPT were recorded before each individual measurement. Values from each device were compared using Passing and Bablok regression of agreement and a Bland and Altman difference plot. One hundred and forty-seven paired measurements were analysed. The average bias between the two methods (EPT vs. APS) was -1.6 mmHg (95 % confidence interval [CI]: -3.0 to -0.2 mmHg). The coefficient of correlation was 1.0084. The aneroid system showed an almost perfect agreement with the EPT. This study shows that it can be used in a clinical setting as well as under field conditions to measure arterial blood pressure in cattle.

Retrospective study of 82 cases of canine lymphoma in Austria based on the Working Formulation and immunophenotyping.

In human beings the prevalence of different non-Hodgkin's lymphoma (NHL) subtypes varies according to geographical region. The aim of this study was to classify canine lymphomas in Austria and to compare the results with those of similar studies in other countries. Eighty-two NHLs were classified according to their morphology (based on the Working Formulation) and their immunophenotype (determined with anti-T-cell and anti-B-cell antibodies). Forty-two (51.2%) were of B-cell subtype, 24 (29.3%) of T-cell subtype, and 16 (19.5%) remained unclassified, because of either negative labelling (9/16) or immunoreaction with both antibodies (7/16). Diffuse lymphomas predominated (99%) over follicular lymphomas, while intermediate grade lymphomas (61%) outnumbered high-grade lymphomas (23.2%) and low grade lymphomas (13.4%). The most common subtype was the diffuse large cell lymphoma (40.2%), followed by the large cell immunoblastic lymphoma (13.4%) and the diffuse small lymphocytic lymphoma (13.4%). Follicular large cell lymphoma and small noncleaved cell lymphoma were uncommon (1.2%). Generally, these findings accord with those of similar studies in Western Europe, making the existence of specific risk factors in Austria unlikely.

Expression of vascular endothelial growth factor and its receptors in canine lymphoma.

Vascular endothelial growth factor (VEGF) stimulates endothelial cell proliferation and has a pivotal role in tumour angiogenesis. The expression of VEGF and its receptors VEGFR-1 and VEGFR-2 was examined immunohistochemically in 43 specimens of canine lymphoma and in six normal lymph nodes. Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR) were performed to detect VEGF protein and mRNA, respectively. VEGF protein was expressed by 60% of the tumours with diffuse cytoplasmic labelling of the neoplastic cells. Endothelial cells, macrophages and plasma cells were also immunolabelled. VEGFR-1 was expressed by variable numbers of neoplastic cells in 54% of lymphoma specimens. VEGFR-1 was also expressed by macrophages, plasma cells, reticulum cells, and vascular endothelial cells. Macrophages and lymphocytes in germinal centres of normal lymph nodes were also immunoreactive with anti-VEGF and VEGFR-1. Most tumours did not express VEGFR-2 but in 7% of sections there was focal labelling of neoplastic and endothelial cells, with a cytoplasmic and perinuclear pattern. The observed variability in expression of VEGF and its receptors probably relates to the fact that lymphoma is a heterogeneous lymphoproliferative tumour. Individual differences in VEGF and VEGFR expression must be taken into account when VEGF and VEGFR-targeted approaches for anti-angiogenic therapy are considered in dogs.

Impression cytology as diagnostic tool in horses with and without ocular surface disease.

BACKGROUND: Cytology plays a major role in the diagnosis of ocular surface diseases. OBJECTIVE: To compare 2 cytological sampling methods for obtaining corneal and conjunctival cell samples regarding irritation for the patient, invasiveness, sample quality and diagnostic equivalence. STUDY DESIGN: Observational prospective study. METHODS: In 5 healthy horses, conjunctival and corneal samples were taken bilaterally by impression cytology sampling (ICS) and cytobrush sampling (CBS). Irritation and invasiveness were assessed with an eye irritation and an epithelial damaging score system, respectively. Sample quality was evaluated via morphometric analysis and graded by a board certified clinical pathologist. For the assessment of diagnostic equivalence, 15 eyes of 14 client owned horses with ocular surface anomalies were sampled by ICS and CBS. The methods were compared regarding the types of inflammatory cells and/or infectious agents detected and if the correct diagnosis could be achieved. Histopathology served as gold standard when available. RESULTS: ICS was significantly less invasive and less irritating for the horses. Both methods retrieved cells of overall high quality; the cell quantity was significantly higher in IC samples. ICS preserved the natural cellular layout. There was a fair, but no statistically significant agreement between the diagnostic outcomes between sampling methods although CBS resulted in a slightly greater variability of inflammatory cell types compared to ICS. MAIN LIMITATIONS: None. CONCLUSIONS: Because of its low irritability and invasiveness ICS can be recommended for cell sampling in fragile corneas and for experimental studies. ICS is especially useful in cases where preservation of cellular layout is advantageous. CBS samples are easier to obtain because of the small equine palpebral fissure. Additionally, the identification of inflammatory cells within conjunctival cell samples is easier. Overall, CBS is still an appropriate method in clinical settings.

Pretreatment leukocyte ratios and concentrations as predictors of outcome in dogs with cutaneous mast cell tumours.

Leukocyte ratios correlate with outcome in several human cancers. Little is known about their prognostic significance in mast cell tumour (MCT). The aim of the study was to evaluate the prognostic significance of pretreatment leukocyte concentrations and their ratios in dogs with MCT for survival. Medical records of 92 dogs with MCT were retrospectively reviewed. Tumour diagnosis was made by tumour biopsy or fine-needle aspirate. Only dogs without prior treatment were included. Eosinophil, lymphocyte, monocyte and neutrophil concentration were obtained by ADVIA 2120™ (Siemens Healthcare, Vienna, Austria). Neutrophil-to-eosinophil ratio (NER), lymphocyte-to-monocyte ratio (LMR) and neutrophil-to-lymphocyte ratio (NLR) were calculated from collected leukocyte concentrations. Relative eosinophil concentration (REC), NER (P < 0.001), NLR (P = 0.001) and LMR (P < 0.001) were significant prognostic factors for outcome in univariate analysis. REC (P = 0.008) and NER (P = 0.001) remained independent predictors of survival in multiple analyses. Leukocyte concentrations and ratios, especially REC and NER may serve as prognostic indicators in MCT.

Characterization of a PCR-based lymphocyte clonality assay as a complementary tool for the diagnosis of feline lymphoma.

Differentiation between resident mature lymphocyte populations and small cell lymphoma cannot be made by cytological review alone and remains challenging in histopathological review. These cases warrant application of complementary tools like PCR-based immunoglobulin (IG) and T-cell receptor (TCR) clonality testing for confirmation. In this prospective study, diagnostic sensitivity and specificity of different primer sets for routine diagnosis of feline TCR gamma (TCRG) and complete IG heavy chain (IGH) gene rearrangements were assessed. Fine needle aspirates from 20 feline lymphoma cases and lymph node material from 10 cats without hematopoietic neoplasia were subjected to clonality testing. Feline lymphoma cell lines and previously confirmed patient material served as positive control. Detection limits for clonal populations within a polyclonal background was 90% for B-cells and 50% for T-cells. Diagnostic sensitivity and specificity of the clonality assay were 70% and 90%. Overall diagnostic accuracy was 77%, positive predictive value 93% and negative predictive value 60%.

Immunophenotype of Peripheral Blood Lymphocytes in Dogs with Inflammatory Bowel Disease.

BACKGROUND: Inflammatory bowel disease (IBD) is common in dogs. Despite the known importance of intestinal lymphocytes in its pathogenesis, little is known about the role of peripheral blood lymphocytes (PBLs) in IBD. OBJECTIVES: The aims of this study were (1) comparison of PBLs analyzed by flow cytometry (FCM) in IBD dogs and healthy controls and (2) comparison of PBLs in IBD dogs at the time of diagnosis and in dogs in clinical remission. ANIMALS: Whole blood samples of 19 IBD dogs at the time of diagnosis and blood samples of 6 dogs in clinical remission were collected. Ten healthy dogs served as controls. METHODS: In this prospective observational study, PBLs were analyzed with multicolor FCM by staining with a panel of anticanine and cross-reactive monoclonal antibodies against T- and B-cell differentiation antigens, including CD45, CD3, CD4, CD8α, CD8ß, TCRαß, TCRγδ, CD79αcy, and CD21. RESULTS: The IBD patients' PBLs had significantly decreased percentages of TCRγδ+ T lymphocytes (median: healthy dogs, 3.32; IBD dogs, 0.97; P = 0.03) and CD21+ B cells (median: healthy dogs, 27.61; IBD dogs, 17.26; P = 0.04). There were no significant differences in PBLs between pretreatment and follow-up samples. CONCLUSIONS AND CLINICAL IMPORTANCE: The differences between PBLs in healthy and IBD dogs analyzed by FCM indicate an imbalance of lymphocytes with different immunologic functions and emphasize the potential value of this technique in a larger cohort of dogs. The PBLs did not differ between IBD dogs before treatment and clinically well-controlled dogs after treatment.

Biological variation, individuality and critical differences of eight biochemical blood constituents in budgerigars (Melopsittacus undulatus).

The aim of the study was to obtain basic data on individual biological variation, the required number of specimens to define the homeostatic setpoint (the aspired value of a variable adjusted to the individuals homeostasis) and critical differences of selected chemistry parameters in budgerigars (Melopsittacus undulatus). Blood from 99 healthy budgerigars was sampled 12 times at four-week intervals. Aspartate aminotransferase (ast), calcium, creatine kinase (ck), glutamate dehydrogenase (gldh), glucose, lactate dehydrogenase (ldh), total protein and uric acid were investigated. The indices of individuality obtained in the present study were relatively low (total protein 0.93, ast 1.02, gldh 1.04, ck 1.12, ldh 1.24, uric acid 1.26, glucose 1.39, calcium 1.61) and suggest that population-based reference limits might be useful. Comparison of data showed that the application of intraindividual reference values identified much greater variation in the reference values than using conventional population-based reference limits. Otherwise, the moderate to low degrees of individuality may allow the use of reference values of one individual as a standard for another individual of the same species. The critical differences that define the change needed between two serial results to indicate a biological change was highest in gldh with 120 per cent or 1.7 U/l. In calcium, a change of 30.5 per cent or 0.5 mmol/l would be significant. In the other parameters critical differences varied between 38 per cent in glucose, up to 93 per cent in uric acid.