key: cord-0041803-t57l3ec7
authors: nan
title: 2013 ACVIM Forum Research Abstracts Program
date: 2013-05-09
journal: J Vet Intern Med
DOI: 10.1111/jvim.12100
sha: ea87658c8817c8e09e649b5b9b6dde8bc2eeb11a
doc_id: 41803
cord_uid: t57l3ec7

nan

Most commonly dogs with sinus node dysfunction (SND) that require pacing are treated with VVI pacing using a single transvenous lead in the right ventricular apex. Two potential problems exist with this pacing method: (1) Loss of atrioventricular (AV) synchrony and (2) Exacerbation of coexisting AV valve degeneration. The latter disease is prevalent in the breeds most commonly afflicted with SND. Pacing the right atrium would permit normal depolarization of the ventricle; however, placement of the lead in the right auricle is difficult in these small dogs. Moreover, assurance of adequate AV nodal conduction (AVNC) is required for atrial pacing alone. While dual chamber pacing offers ideal pacing with backup when AVNC is abnormal or questionable, cranial vena caval syndrome is a risk and synchronized ventricular contraction is lost. Therefore, atrial pacing with a single lead in a location that is technically feasible, after assurance of adequate AVNC, would be preferred in dogs with SND.

The objectives of this pilot study were to determine the Wenckebach point (WP) for the assessment of AVNC during a commonly used anesthesia protocol, and to assess the interatrial septum as an alternative site for atrial pacing in small dogs. Six beagles (8.3-12.9 kg) were given propofol for induction and isoflurane for maintenance of anesthesia. The WP was tested via atrial pacing up to 220 bpm. Using single plane fluoroscopic guidance, Medtronic â 3830 Select Secure active fixation leads were placed in the interatrial septum via a Medtronic â Attain Select â II standard 90 LV delivery catheter. Pacing threshold and lead impedance were assessed at implantation. Biplane thoracic radiographs were taken immediately following implantation to identify the lead position.

In 5 of 6 dogs the WP was not reached at the maximum rate of 220 bpm. In one dog the WP was 110 bpm. Atrial capture thresholds and lead impedances were evaluated at implant. Lead impedance ranged from 583 to 1421 ohms. Lead dislodgement was detected in 2 dogs 24 hours after implantation and in 3 dogs at one month. The lead remained in position in only one dog. Following consultation with Medtronic implantation specialists, the problems identified were directed specifically to requirements of the 3830 lead: (1) Inadequate lead slack for this lead. (2) Low impedance recorded at implant (<900 ohms indicates inadequate helix fixation). (3) Specific location in the cranial right atrial septum should be assessed via biplane imaging during implantation. The single dog in which the lead remained in place followed these requirements.

A learning curve exists with any new technique. This pilot study provides initial data concerning the WP in normal dogs and information to optimize the method for atrial lead positioning. Symptomatic bradyarrhythmias unresponsive to medical therapy remain the most common indication for artificial cardiac pacing in canines. While the right ventricular apex (RVA) allows for easy endocardial placement of leads, there is mounting evidence of the detrimental effects of long term RVA cardiac pacing. During RVA pacing, the action sequence deviates significantly from normal and is associated with considerable depression of left ventricular (LV) function. One human study has shown that the LV adverse remodeling and deterioration of systolic function in patients with 3 rd -degree AV block is prevented by BiV pacing. Prior studies in dogs using tissue Doppler imaging (TDI) did not identify a statistical difference in LV synchronization between pacing sites, although there was a trend for more synchronization during BiV pacing. A newer 2D imaging modality, speckle tracking strain analysis, is more effective in detecting LV dyssynchrony in humans. This study aimed to evaluate the effect of pacing site on LV synchronization, using 2D speckle tracking strain.

Six canine patients with complete AV block and five purpose bred research dogs implanted with transvenous BiV pacing systems were included. Electrocardiograms and echocardiograms were assessed during pacing from the: Right Atrial Appendage/ RVA(RAA/RVA), RAA/LV, and RAA/BiV in each dog. Radial (RS) and circumferential strain (CS), using right parasternal short axis at the level of the papillary muscles were calculated. Synchrony was assessed by measuring the difference in the QRS onset time-to-peak strain between the interventricular septum and left ventricular free wall segments. A one-way ANOVA was performed to evaluate differences in pacing modality. LV synchronization was also assessed at regular recheck intervals for dogs with complete AV block and BiV pacing systems. Differences in LV synchronization during BiV pacing were assessed over the 2 year study period with a paired T-test. A p < 0.05 was considered significant.

Significant differences were found in the time to peak RS (P = 0.022) between groups. Post-Hoc tests showed a significant difference between dogs paced from the RV and the BiV, with a mean RS of 74 ms and 34 ms , respectively. CS did not exhibit significant differences between groups (p = 0.495). The paired T-test did not show any differences on RS or CS on the BiV pacing dogs over time (p = 0.94 and 0.18, respectively) .

The results of this study show that there is dyssynchrony created with RVA pacing and that BiV, but not LV pacing appears to preserve LV synchronization in dogs with complete AV block treated with artificial cardiac pacing. The significance and long term functional effects of BiV pacing in dogs with complete AV block is unknown but currently under evaluation by our group. This study evaluated the clinical findings, success rate, and complications for dogs undergoing radiofrequency catheter ablation (RFCA) of accessory pathways (APs). Medical records, electrophysiologic (EP) data, echocardiographic data, and long-term follow-up by owner contact were reviewed retrospectively for dogs undergoing RFCA of APs from 1997 -2012.

Fifty-eight dogs with a median age of 1 yr 9 m (5 m-10 yr) and a mean weight of 28 kg (11 kg-50 kg) were evaluated. Twenty breeds were represented with the most common being Labrador retriever (36%), golden retriever (10%), and English Bulldog (7%).

All dogs were diagnosed with orthodromic atrioventricular reciprocating tachycardia (OAVRT). Median maximum heart rate was 303 bpm (240-400 bpm). Common presenting complaints included weakness / lethargy (66%), signs of congestive heart failure (CHF) (36%), vomiting / diarrhea (36%), decreased appetite (33%), and syncope (18%). Twenty-one dogs (36%) had confirmed CHF. Eight (14%) had left-sided CHF, eight (14%) had right-sided CHF and five (9%) had biventricular CHF. Six additional dogs (10%) had tachypnea with suspected CHF. Twenty-four dogs (41%) had decreased systolic function defined as fractional shortening of less than 25%.

Of the 58 dogs undergoing EP study for APs, 53 dogs (91%) had successful ablation procedures. Three dogs (5%) required a second procedure for long-term success. Three dogs (5%) died in the peri-operative period. Of the 55 dogs (95%) surviving surgery, two had pathways that were not successfully ablated (3%). One dog required pacemaker implantation secondary to atrioventricular nodal damage (2%).

A total of 65 APs were identified in 57 dogs with AP mapping (one dog died prior to mapping). Forty-nine (86%) dogs had a single pathway, while 8 dogs (14%) had 2 APs. The majority of the APs were located in the right posteroseptal region (55%).

Other locations included the right free wall (28%), midseptal region (8%), left posteroseptal region (6%), and left free wall (3%). Sixty-three (97%) of APs were successfully ablated. The APs were evenly distributed between retrograde conduction (49%) and bidirectional conduction (49%). One AP conducted only antegrade (2%). Median cycle length during RFCA was 214 msec (146 -298 msec).

Long-term follow-up by owner contact was available in 50 (91%) of the 55 dogs that survived the initial procedure. Follow-up data in all dogs ranged from 0 months to 11.5 years. There are 27 dogs still living. Post-procedural survival times in deceased dogs ranged from 1 year to 11 years with a median survival of 5 yr 10 m. None of the dogs with successful AP ablation needed antiarrhythmic therapy after the procedure. Only 1 dog has passed away from suspected cardiac disease. This Boxer dog died suddenly at age 13, which was 6 years after successful ablation.

In conclusion, RFCA is a well-tolerated procedure with low incidence of major complications. This is an effective cure for OAVRT secondary to an AP, eliminating clinical signs and the need for antiarrhythmic therapy. Orthodromic atrioventricular reciprocating tachycardia (OA-VRT) is a supraventricular tachycardia caused by an accessory pathway. It can lead to tachycardia-induced cardiomyopathy (TICM) and congestive heart failure. Treatment options include anti-arrhythmic drug therapy and radiofrequency ablation (RFA).

The purpose of this study was to determine long term clinical and echocardiographic outcome in cases of OAVRT in dogs in the UK undergoing RFA.

Case records of the first 20 dogs to undergo RFA at the R(D) SVS (2006) (2007) (2008) (2009) (2010) (2011) were reviewed. TICM was determined by the presence of dilation of the left or right ventricle, decreased contractility and/or clinical manifestation of congestive heart failure.

Follow-up ranged from 92 to 1922 days (mean 867 days) in all dogs and from 350 to 1922 (mean 943 days) in the dogs where ablation was completely successful. RFA resolved clinical signs and prevented recurrence of OAVRT in 18/20 dogs. One dog died immediately post operatively and one dog's left-sided pathway could not be ablated. Echocardiographic follow-up was available in 16/20 dogs. M-mode and 2D variables indexed according to allometric scaling revealed none of the dogs showed progression of TICM. Comparing pre and post ablation LVIDs, LVIDd, left atrial diameter and LA:Ao ratio significantly decreased, while IVSs, LVPWs, LVPWd, aortic diameter and weight significantly increased (p < 0.045).

In conclusion, to date this study is the most comprehensive clinical and echocardiographic assessment of long-term outcome after RFA in dogs and shows that RFA is effective in permanently resolving OAVRT, preventing recurrence of clinical signs and results in reversal of TICM. Although it is well known that the prominent rhythm of the canine heart is a sinus arrhythmia, only recently has a nonsingular distribution (NSD) of beat intervals been documented leaving a region with a paucity of intervals (zone of avoidance, ZOA). This observation may be associated with the multicentric origin of the sinus node impulse; however, the latter cannot explain the mechanism of the NSD. Spontaneous discharge of the canine sinus node is under the influence of the autonomic nervous system, voltage clock (I f channel), and calcium clock (spontaneous release of calcium from the sarcoplasmic reticulum). Alterations in these factors have been implicated in sick sinus syndrome. The purpose of this study was to deconstruct specific components that influence the sinus node discharge in the ex vivo canine sinus node specifically to determine if (1) acetylcholine (ACH), (2) blockade of the calcium clock (via ryanodine) or (3) blockade of the voltage clock (via I f channel blockade with ZD7288) result in NSD with a ZOA.

Right atrial monophasic action potential (MAP) recordings and 2-site unipolar epicardial electrograms were made from perfused canine preparations with intact sinus nodes. Baseline recordings were made initially and during washout periods after ACH. Coronary perfusions with ACH (2-3uM) (n = 12), ryanodine (2-3uM) (n = 5) and ZD7288 (3uM) (n = 5) were performed. Each of the preps could only be treated with either ryanodine or ZD7288 due to permanent binding. Data were transferred from the AcqKnowledge acquisition program to MatLab. The dV/dt maximum was used to select beats and the S n S n (sinus node beat) intervals were determined. These S n S n intervals were plotted as a function of their duration (S n S n interval histogram), as a beat to beat relationship to the next interval (Poincar e plot), and as a time series maintaining the order of the intervals over time (tachogram) . Data was examined for beat distribution and proprietary software was then used to define and detect single, bimodal, and trimodal Gaussian distributions of S n S n intervals. The mean AE SD and area of the identified interval distributions were calculated after iterations to identify those fitting the estimated distribution modes.

Ex vivo studies showed singular distribution (one area > 95%) of sinus node discharge in all baseline preps (interval mean AE sd) (565 AE 30 ms) and all those treated with ryanodine (1137 AE 136 ms) or ZD7288 (1283 AE 233 ms) alone. However, preps treated with ACH (2097 AE 390 ms) (one area < 95% with range 19-63%; P < 0.001) demonstrated NSD and the ZOA. These results demonstrate that the non-innervated inherent rhythm of the canine sinus node has a singular distribution, but treatment with ACH reproduced the distributions as seen in vivo. Moreover, specific blockade of the voltage clock or calcium clock did not recreate the in vivo distribution. ACH is known to affect both the voltage and calcium clock, thus, further studies whereby suppression of both clocks is needed to determine if the NSD is recreated. Reconstruction of the NSD and ZOA with ACH in the ex vivo model suggests that other physiologic factors are not in play. The mechanism whereby ACH (as the mediator of parasympathetic tone) creates the unique beat intervals in the dog requires further study.

ate RV systolic function in dogs with different degrees of MMVD.

Clinical examination including echocardiography was performed in 103 privately-owned dogs (16 controls dogs (Beagles), 70 CKCSs with different degrees of MR and 17 dogs of different breeds with clinical signs of heart failure (HF) due to MMVD). Right ventricular systolic function was estimated by tricuspid annular plane systolic excursion (TAPSE) assessed by 2D guided M-mode and tricuspid annular peak systolic velocity (TDPW) evaluated by pulsed tissue Doppler imaging. Influence of MMVD disease group, age, gender, heart rate (HR), severity of tricuspid regurgitation (TR) and pulmonary hypertension on RV systolic function was evaluated using analyses of covariance.

TAPSE (P = 0.009) and TDPW (P = 0.005) were significantly associated with disease group. On post-hoc comparisons, CKCSs with heart murmur but without left atrial enlargement had significantly lower TAPSE (P = 0.004) and PWTD (P = 0.02) than control dogs. In addition, TDPW was lower in HF dogs compared to control dogs (P = 0.004). An influence of concomitant TR on TDPW was found (P = 0.02). TAPSE (P = 0.008) and TDPW (P = 0.03) increased with advancing age. TDPW increased with increasing HR (P = 0.002).

In conclusion, echocardiographic measurements of RV function indicate decreased RV systolic function in dogs with MMVD. Studies are needed to elucidate prognostic significance of the findings. Serotonin (5-HT) signaling is implicated in the pathogenesis of myxomatous mitral valve disease (MMVD) through 5-HT1B receptor (R) 5-HT2AR and 5-HT2BR-induced myxomatous pathology. Increased serotonin synthesis and decreased clearance, indicated by tryptophan hydroxylase-1 (TPH-1) and serotonin reuptake transporter (SERT), respectively, have been found in myxomatous valves. Extra-valvular lesions have also been documented in relation to MMVD, but the presence of serotoninrelated markers in extra-valvular tissue has not previously been reported. Using an experimental porcine model of chronic mitral regurgitation (MR), we investigated the gene expression of 5-HT1BR, 5-HT2AR, 5-HT2BR, SERT and TPH-1 in the mitral valve (MV), anterior papillary muscle (AP) and left ventricular free wall (LV) by means of qPCR.

Twenty-eight pigs were subjected to surgically-induced MR or sham operation resulting in three groups: Control (MR 10%, n = 12), mild (MR20-50%, n = 10) and severe MR (MR ! 80-100%, n = 6). Gene expression differences between locations were examined in the control group. Expression of all serotonin receptors was significantly higher in the MV compared to LV and AP (all P < 0.0001) while expression of TPH-1 was significantly higher in the LV compared to the MV (P = 0.002). Analysing impact of MR, up-regulation of valvular 5-HT2BR expression (P = 0.03) was found in the severe MR group compared to the control group.

In conclusion, control pigs show predominant valvular 5-HTR-expression whereas serotonin synthesis predominates in the left ventricle. Whether myocardial-derived 5-HT interacts with valvular 5-HTR needs further investigation. In severe MR, altered hemodynamics secondary to MR might increase 5-HT2BR signalling, emphasising the 5-HT2BR as potential future therapeutic target in MMVD. The purpose of this prospective study was to investigate the association of selected cardiac biomarkers with clinical estimates of patient stability.

Twenty-one client-owned dogs with ACVIM Stage C or D chronic degenerative valvular disease on appropriate combination therapy were recruited from the caseload of the clinical cardiology service at TAMU and evaluated twice within 1 to 4 weeks. Each evaluation consisted of a complete history, exam, FETCH questionnaire, thoracic radiographs, echocardiogram, ECG, biochemistry panel, CBC, blood pressure, brain natriuretic peptide (BNP), NT-proBNP, and high sensitivity cardiac troponin I (hs-cTnI). At each evaluation, dogs were classified as clinically stable (S) or unstable (US) based on all diagnostic tests except the results of the cardiac biomarkers; S-dogs had no current clinical signs related to heart failure, US-dogs had evidence of decompensation significant enough to require a change in treatment.

Median (IQR) age and weight was 10.9 years (5.4-16.2) and 8.8 kg (.03-14.2) respectively. Mean NT-proBNP [S, US, p-value] (1890 pmol/L, 2542 pmol/L, 0.02), BNP (4.52 pg/mL, 9.43 pg/ mL, 0.013), and hs-cTnI (0.086 ng/mL, 0.157 ng/mL, 0.038), using both clinical evaluations (n = 42 observations on 21 dogs), differed significantly between S and US dogs. Mean hs-cTnI was significantly different (0.049 ng/mL, 0.152 ng/mL, 0.008) in dogs that were S at both evaluations (n = 8) when compared to all other dogs that were US (n = 13) on at least 1 evaluation. This data suggests that biomarkers, in particular hs-cTnI, may be complimentary to other methods of clinical assessment of stability. The phosphodiesterase 5A gene (PDE5A) encodes for cyclic guanosine monophosphate (cGMP) specific phosphodiesterase. It is a key physiologic regulator of cGMP and pharmacologic target of drugs used to treat pulmonary hypertension and erectile dysfunction such as sildenafil. PDE5A polymorphisms in human beings have been described that may predict response to therapy with sildenafil and nitric oxide.

We hypothesized that polymorphisms in the PDE5A gene exist in dogs and that basal levels of cGMP would be significantly affected by these changes.

Genomic DNA from 15 unrelated dogs of 3 breeds (Golden Retriever, Norwich Terrier, Cavalier King Charles Spaniel) was utilized. Exonic, splice-site, and untranslated regions of the PDE5A gene were sequenced and compared to the dog and human reference sequences. Nucleotide substitutions were recorded and evaluated with commercially available software for potential damage to protein structure and function. Polymorphism frequency was evaluated in a larger cohort of 55 additional, unrelated, healthy dogs of 14 breeds. Plasma cGMP levels were measured in 40 PDE5A genotyped dogs and compared using an unpaired t test by genotype group.

An exonic, amino-acid substituting polymorphism was observed in a highly conserved region of the PDE5A gene in 61/ 70 dogs (21 homozygous, 40 heterozygous). The exonic polymorphism was evaluated and judged to be damaging based on protein modeling and change of amino acid charge and acid-base status. A statistical difference was observed with the reported amino acid substitution. Dogs that match the reference sequence amino acid structure were significantly higher in cGMP when compared to the pooled group of heterozygous and homozygous genotype dogs (P = 0.007). A series of 5 linked single nucleotide polymorphisms was observed in 44/70 dogs (11 homozygous, 33 heterozygous). The linked SNPs were evaluated and determined to result in significant changes to secondary protein structure. There were no statistical differences observed in plasma cGMP with the 5 linked polymorphisms or based upon age, breed or gender.

The PDE5A gene of dogs contains multiple polymorphisms and an exonic amino acid substitution that suggests a functional role. Further investigation into genotype specific PDE5A function and response to drug therapy may be warranted.

C-10 EFFECT OF LOW-DOSE IMATINIB THERAPY FOR PUL-MONARY ARTERIAL HYPERTENSION AND HEMODY-NAMIC DISTURBANCE CAUSED BY CHRONIC HEART FAILURE IN DOGS. S Arita 1,2 , N Arita 2 , Y Hikasa 1 . 1 Department of Veterinary Internal Medicine, Tottori University, Tottori, Japan., 2 Arita Sougo Animal Hospital, Hiroshima, Japan.

Pulmonary arterial hypertension (PAH) is a prognostic factor in intractable cases of heart failure. PAH is due to increased pulmonary vascular resistance caused by pulmonary vasoconstriction and vascular remodeling, The expression of platelet-derived growth factor (PDGF) and PDGF receptors is markedly increased in pulmonary vascular lesions of humans with PAH. The prevention of excessive PDGF-induced proliferation of pulmonary artery smooth muscle cells may result in the suppression of pulmonary vascular remodeling. Imatinib is a molecular-targeted therapeutic drug that inhibits the activation of PDGF by impeding phosphorylation of the PDGF receptor tyrosine kinase. It is reported that low-dose imatinib, 1 / 6 -1 / 3 of the anti-neoplastic dosage, is useful to improve PAH in humans. However, there are no reports available on the effectiveness of imatinib for PAH in dogs. The purpose of this study was to examine the effect of lowdose imatinib therapy on hemodynamic parameters and clinical manifestations in dogs with PAH due to advanced MR and heartworm disease.

Eight client-owned dogs with PAH (6 cases with advanced MR, and 2 cases with chronic filariasis) were prospectively recruited, and informed consent was obtained from every dog owner. All cases had been treated by polypharmacy approach with general therapeutic drugs for PAH. They were administered 3 mg/kg imatinib mesylate (Glivec) orally, every 24 hours, for 30 days. Before and after imatinib administration, all dogs continued to receive previous medications without any change. Physical examination, blood biochemical tests, blood pressure measurements, radiography, and Doppler echocardiography were performed at least twice, prior to imatinib administration (Pre) and again 30 days after administration (Post) A paired t-test was used for comparison between Pre and Post data for variables.

Systolic pulmonary arterial pressure, heart rate, maximum tricuspid regurgitation velocity, left atrium/aorta ratio, right and left ventricular Tei indexes, and early diastolic transmitral flow wave/mitral annulus velocity ratio decreased significantly after imatinib therapy. Diastolic blood pressure, stroke volume, cardiac output, and left ventricular fractional shortening increased significantly after therapy. Plasma atrial natriuretic peptide levels decreased significantly after imatinib therapy, and serum N-terminal pro-brain natriuretic peptide levels tended to decrease after therapy. Clinical signs including cough, exercise intolerance, syncope, and edema reduced after imatinib therapy. No plasma biochemical abnormalities suggesting renal and liver pathology were observed during imatinib therapy.

These results revealed that low-dose imatinib therapy led to reduction of systolic pulmonary arterial pressure without decreasing systemic blood pressure or cardiac output, and successfully improved cardiac function and hemodynamics in dogs with PAH caused by advanced MR or chronic filariasis. This study suggests that low-dose imatinib therapy is effective for chronic heart failure in dogs with PAH without any apparent adverse effect.

TO SEVERE OCCULT (ASYMPTOMATIC) FELINE HEART DISEASE. MC Machen 1 , SG Gordon 2 , JE Rush 3 , SE Achen 4 , RL Stepien 5 , PR Fox 6 , PM Lee 6 , MA Oyama 1 . 1 University of Pennsylvania, Philadelphia, PA, 2 Texas A&M University, College Station, TX, 3 Tufts University, North Grafton, MA, 4 Michigan Veterinary Specialists, Southfield, MI, 5 University of Wisconsin, Madison, WI, 6 The Animal Medical Center, New York, NY.

Detection of occult (asymptomatic) feline heart disease (OHD) is challenging. A pilot study indicates that a point-of-care second generation ELISA that utilizes SNAP â technology identifies cats with moderate to severe OHD. We sought to prospectively validate the assay in a larger population through a prospective, multicenter trial.

Asymptomatic cats "at-risk" for OHD were prospectively recruited based upon the presence of a heart murmur, gallop, arrhythmia, and/or being a predisposed breed. All cats received physical examination, non-invasive blood pressure measurement and echocardiogram. NTproBNP SNAP assay was performed at time of echocardiogram and visually evaluated by a reader blinded to the echo results.

One hundred and seventy-six at-risk cats were examined and classified as normal (n = 56), equivocal (n = 17), mild OHD (n = 55), moderate OHD (n = 37), or severe OHD (n = 11). Diagnosis in cats with OHD included HCM (n = 34), HOCM (n = 45), R/UCM (n = 7), and other (n = 17). Visual interpretation of the SNAP had a sensitivity/specificity of 87%/78%, respectively, to differentiate all cats with moderate and severe OHD and 91%/77% to differentiate cats with moderate and severe HOCM, HCM, or R/UCM. Comparison of SNAP assay vs. quantitative ELISA revealed a high degree of correlation between assays, and that a positive SNAP test result was associated with a NT-proBNP concentration~130pmol/L.

In the study population with prevalence of moderate to severe OHD of 27%, overall accuracy of SNAP to detect moderate to severe OHD was 80.1% with more false positives than false negatives and SNAP performed best in ruling out OHD in the study population. NT-proBNP is a plasma marker of increased cardiac wall stress that has utility in distinguishing cardiac and respiratory causes of dyspnoea. We hypothesised that measurement of NT-proBNP concentrations in pleural fluid obtained during therapeutic thoracocentesis would allow differentiation of cardiac from non-cardiac causes of pleural effusion in cats.

Samples of blood and pleural fluid were prospectively collected from cats presenting to the RVC for treatment of pleural effusion. Cats were categorised according to the cause of pleural effusion (cardiac vs. non-cardiac) based on diagnostic tests, including echocardiography, interpreted by a board-certified cardiologist. Blood and pleural fluid samples were separated within 30 minutes of collection and plasma and supernatant transferred into protease inhibitor (PI) tubes according to the manufacturer's instructions. PI plasma and pleural fluid samples were stored at -80°C for batched analysis. NT-proBNP concentrations were measured using a commercially-available feline-specific ELISA which has been validated for use with plasma samples. Precision and reproducibility of measurements of NT-proBNP in pleural fluid were assessed by calculation of intra-and inter-assay coefficients of variation (CV), respectively, in samples of low, medium and high NT-proBNP concentrations. Correlations were assessed using Spearman's correlation coefficient. Group-wise comparisons were performed using Mann-Whitney tests. Receiver operating characteristic curves were constructed to determine separate cutoffs to distinguish cardiac from non-cardiac causes of pleural effusion in plasma and pleural fluid samples.

Thirty-eight cats with pleural effusion, 19 of non-cardiac and 19 of cardiac causes, were studied between February 2011 and June 2012. Intra-assay CVs for samples of low (150 pmol/L), medium (609 pmol/L) and high (1332 pmol/L) NT-proBNP concentrations were 5.1%, 8.6% and 2.1%, respectively. Inter-assay CVs for samples of low, medium and high NT-proBNP concentrations were 16.5%, 10.3% and 10.0%, respectively. NT-proB-NP concentrations in plasma and pleural fluid were strongly correlated (R s =0.82, P < 0.001). NT-proBNP concentrations were significantly higher in the cardiac group in both plasma (P < 0.001) and pleural fluid (P < 0.001). In plasma samples, a cut-off of 273 pmol/L predicted a cardiac cause of pleural effusion (sensitivity=94.7%, specificity=89.5%, positive likelihood ratio (PLR)=15.2 (AUC=0.96, 95% confidence interval (CI) 0.90-1.0)). In samples of pleural fluid, a cut-off of 757 pmol/L predicted a cardiac cause of pleural effusion (sensitivity=94.7%, specificity=94.7%, PLR=18.0 (AUC=0.95, 95% CI 0.87-1.0)).

In conclusion, NT-proBNP can be measured in feline pleural fluid with good precision and acceptable reproducibility. Measurement of NT-proBNP in plasma or pleural fluid allows differentiation of cardiac from non-cardiac causes of pleural effusion with similar accuracy in cats. These findings should be validated prospectively in a separate population. Dexmedetomidine (DXM) is approved for sedation of cats, and its alpha-2 agonist properties may make it preferable in cats with left ventricular (LV) concentric hypertrophy. However, effects of DXM on the echocardiogram (echo) and serum concentrations of cardiac troponin-I ( [cTnI] ) and N-terminal pro-B-type natriuretic peptide ( [NT-proBNP] ) are poorly described in cats.

Volunteered healthy pet cats (n = 14) underwent baseline (PRE) testing: high-sensitivity measurement of [cTnI] (Abbott Architect Stat Troponin-I) and [NT-proBNP] (IDEXX Laboratories), and echocardiography (ECHO; 2D, M-mode, spectral Doppler, volumetric calculations). Cats received DXM, 40 mcg/ kg IM per label and baseline tests were repeated when cats were sedated (INTRA, 15-38 minutes after DXM injection). DXM was reversed with atipamezole (ATI; 300 mcg/kg IM) and cats underwent the same tests 2 hours later (2H POST). Finally, [cTn-I] and [NT-proBNP] were analyzed on blood drawn 24 hours later (24H POST).

Emesis occurred in 14/14 cats after DXM administration, prior to the onset of sedation. DXM administration was associated with a marked increase in mean left ventricular (LV) end-systolic diameter ( Mitral valvular insufficiency was present in 1 cat (7%, trace) PRE and all cats (100%) INTRA; it resolved 2H POST in 8 cats (58%). Similarly, aortic valvular insufficiency was present in 2 cats (14%; both trace) PRE and 12/14 cats (86%; 4 trace, 6 mild, 1 mild-moderate, 1 moderate) INTRA; it resolved 2H POST in 8 cats (58%).

The following biomarker changes were statistically significant: [cTnI (ng/ml)] from 0.026 PRE to 0.196 2H POST (P = 0.002), from 0.099 INTRA to 0.196 2H POST (P = 0.02); [NT-proBNP (pmol/l)] from 59 PRE to 89 2H POST (P = 0.03), and from 59 PRE to 90 24H POST (P = 0.04).

In the healthy cats in this study, DXM markedly increased LV systolic dimensions but had no significant effects on LV diastolic dimensions or LV wall thickness during or after sedation. After ATI reversal, cardiovascular changes were slower to return toward normal than were sedative effects. Minimal valvular insufficiency was commonly caused by DXM and was reversible. Acute, DXM-induced transient loading of the LV produced changes in cTnI and NT-proBNP that may lead to greater understanding of the kinetics of these biomarkers in cats. A valid and reliable method has been developed for assessing health-related quality of life (QOL) in cats with cardiac disease. However, further research is needed to test the tool's sensitivity to changes in medical treatment (responsiveness) and its potential role as a clinical and research tool. The goals of this study were 1) to determine optimal timing for administration of the CATCH questionnaire in cats with acute congestive heart failure (CHF), 2) to determine the responsiveness of the questionnaire to changes associated with medical treatment in cats with CHF, and 3) to assess the relationship between changes in the questionnaire and N-terminal pro BNP (BNP). Cats were enrolled at the time of admission for acute CHF due to any type of cardiomyopathy. BNP was analyzed during hospitalization and the CATCH questionnaire was administered to the owner prior to discharge. The questionnaire was re-administered 3 days after discharge by telephone. Finally, cats were re-evaluated 7-14 days after discharge when BNP was measured and the CATCH questionnaire was administered. Thirty-seven cats were enrolled in the study: 5 cats died or were euthanized before discharge, 1 was euthanized after discharge but before the final study visit, and one was lost to follow up. Therefore, 30 cats completed the study and were included in the final analysis. Mean (AESD) age was 10.7 AE 4.0 yrs, with 17 males and 13 females (all neutered). Most cats had hypertrophic cardiomyopathy (n = 20), with smaller numbers of unclassified (n = 5), dilated (n = 3), restrictive (n = 1), and arrhythmogenic right ventricular (n = 1) cardiomyopathies. All cats were in International Small Animal Cardiac Health Council 3a (n = 3) or 3b (n = 27) at the time of first admission; ISACHC score improved significantly (p < 0.001). Median BNP at baseline was 655 pmol/L (range, 188 to >1500 pmol/L) and was 583 pmol/L (range, 41 to >1500 pmol/L) when reassessed 7-14 days later (p = 0.59). Median QOL was 26 at baseline (range, 0-70; possible range of 0-80, with 80 being the worst perceived QOL), 19 (range, 0-61) at the time of discharge, and 19 (range, 2-49) 7-14 days after discharge (p = 0.66). Responses on the questionnaire from owners were highly variable and scores did not correlate with either BNP or ISACHC. Although the CATCH questionnaire is valid for use in cats with cardiac disease, these results suggest that the CATCH questionnaire requires further refinement for uses requiring a relatively responsive instrument. Ragdoll cats are predisposed to hypertrophic cardiomyopathy (HCM) and are reported to have a poor prognosis. A mutation, with a prevalence of 31%, in the myosin binding protein c3 (MYBPC3) gene has been associated with HCM in Ragdolls. Anecdotal reports suggest homozygous cats are likely to develop severe HCM. No study has yet reported the outcome of a large cohort tested for the MYBPC3 mutation. We hypothesized that homozygous cats have a shorter lifespan and are more likely to be diagnosed with HCM or suffer cardiac death.

We designed an online questionnaire to determine the genotype, age, sex, status (alive/dead), circumstances of death and whether HCM had been diagnosed during life. The questionnaire was publicised to Ragdoll owners/breeders online, through mailing lists and a breed club. Kaplan-Meier and Log Rank analysis was performed to evaluate associations with survival.

Responses (n = 174) comprised 117 wild-type (WT), 49 heterozygous and 8 homozygous cats. 66 (37.9%) cats were male. There was no significant association between sex and genotype (p = 0.526). Overall median survival time was 13.6 years in WT, 12.3 years in heterozygous and 5.59 years in homozygous cats. Homozygous cats had a shorter time to a composite end-point of HCM diagnosis and/or cardiac death than WT (p < 0.001) or heterozygous (p = 0.003) cats. Although there was no association between sex and all-cause mortality, male cats had a shorter time to HCM diagnosis and/or cardiac death (p = 0.006).

Ragdoll cats homozygous for the MYBPC3 mutation are more likely to succumb to cardiac disease than WT or heterozygous cats. In cats presenting with respiratory distress it is challenging to differentiate congestive heart failure from primary respiratory disease as a cause of observed clinical signs. Studies have demonstrated that the measurement of N-terminal pro-B-type natriuretic peptide (NT-proBNP), a marker for heart stress detectable in blood, enhances the ability to distinguish between congestive heart failure and primary respiratory disease in presenting animals.

The bioanalytical method validation of a prototype secondgeneration immunoassay for the quantification of feline NT-proBNP is presented. Sensitivity was determined by measuring the limit of detection, defined as three standard deviations above background. Accuracy was measured via dilutional parallelism of six different high concentration plasma samples and calculating observed to expected ratios. Intra-assay, inter-assay and total precision was determined for six samples based upon six replicates per run x three serial lots x two runs per day x three days x three operators. The performance of the assay with 107 matched feline plasma and serum samples was determined. Finally, a Bland-Altman method comparison between the first and second-generation feline NT-proBNP assays was performed using 66 feline plasma samples.

Results establish the assay sensitivity at 10 pmol/L. The average observed to expected ratio for serial dilutions is 103.5% AE 11.1% for six different plasma samples. Coefficients of variation for intra-assay, inter-assay, and total precision for six samples are as follows: [NT-proBNP] The results of 107 matched feline plasma and serum samples did not differ (p = 0.3449), indicating that the test can support both sample types. A method comparison between the first and second-generation feline NT-proBNP assays performed using 66 feline plasma samples indicated an overall bias of 34 pmol/L that was not statistically significant (p = 0.1992) .

The results of this evaluation indicate that the performance of a second generation feline NT-proBNP immunoassay with improved precision and accuracy will more effectively measure feline NT-proBNP and serve as a valuable tool for assessing feline heart health. Cardiac cachexia, a loss of lean body mass, is a syndrome that often accompanies congestive heart failure (CHF) in dogs, cats, and people, and is associated with numerous deleterious effects including reduced strength and quality of life, impaired wound healing and immune function, and reduced survival. Due to adverse effects of cardiac cachexia and the limited methods of treating this syndrome, improved strategies to treat cardiac cachexia would be beneficial. Myostatin is a protein that inhibits muscle growth. Blocking myostatin therapeutically appears to significantly enhance muscle size and strength in rodent models and in human clinical trials. Therefore, the goal of this study is to use a dog specific myostatin antagonist [Canine ActRIIB-Fc (CAP-031)] in a pilot study to test its efficacy in dogs with CHF and cardiac cachexia. Dogs with CHF due to chronic valvular disease (CVD), cardiomyopathy (CM), or congenital cardiac disease (CD) and moderate to severe cachexia were eligible for the study. Dogs received 4 weekly subcutaneous injections of CAP-031. Endpoints were body weight, body condition score (BCS; on a 1-9 scale), muscle condition score (MCS; on a 0-4 scale, where 0 = no muscle loss, 1 = mild muscle loss, 2 = moderate muscle loss, 3 = marked muscle loss, and 4 = severe muscle loss), owner assessment of dog's appetite on a visual analog scale, plasma tumor necrosis factor concentrations, echocardiography, and quality of life (as assessed by the Functional Evaluation of Cardiac Health [FETCH] Questionnaire). To date, 7 dogs have been enrolled; 6 successfully completed the study with no side effects and 1 dog developed fever and polyarthropathy after the first injection. This dog was found to be Borrelia positive and responded to antibiotics but received no further CAP-031 injections. For the 6 dogs that completed the study, mean (AESD) age was 8.8 AE 1.7 yrs. Underlying diseases included CM (n = 4) and CVD (n = 2), and dogs were in International Cardiac Health Council stage 3a (n = 5) or 2 (n = 1). Median time since CHF onset was 326 days (range, 26-1019 days). All dogs were receiving furosemide (median dosage=4.8 mg/kg/day, range, 1.8-5.8 mg/kg/day), ACE inhibitor, and pimobendan. Other medications included diltiazem (n = 5), spironolactone (n = 4), digoxin (n = 4), torsemide (n = 3), and 1 each of carvedilol, amiodarone, and sildenafil. At baseline, median body weight = 27.0 kg (range, 17.3-62.0 kg), mean BCS = 3.8 AE 1.2, and mean MCS = 3.3 AE 0.5. Mean baseline FETCH score was 23 AE 14. There were no significant changes in body weight, BCS, appetite, or FETCH score. Mean MCS improved numerically (from mean = 3.3 at baseline to mean = 2.5 at week 4) but this change was not statistically significant (p = 0.09). All dogs are alive with median survival time = 510 days (range, 119-1215 days). Further research is needed on this and other therapies for cardiac cachexia.

balloon valvuloplasty (HPBV) in dogs with severe subaortic stenosis (SAS). Twenty-eight dogs with severe SAS (peak Doppler systolic left ventricular pressure gradient (PG) > 80 mmHg) underwent CBV and HPBV at six locations and were reexamined at one day (n = 28), one month (n = 20), three months (n = 16), six months (n = 17), twelve months (n = 13), eighteen months (n = 5), twenty-four months (n = 11), and thirty-six months (n = 2) post-procedure. CBV combined with HPBV significantly reduced the intra-procedure peak-to-peak systolic PG (n = 20) from 84.47 AE 58.42 mmHg (mean AE SD) to 38.38 AE 26.25 mmHg (P < 0.0001). Compared with baseline, median peak Doppler systolic PG across the left ventricular outflow tract (143.36 mmHg; was significantly decreased at one day (77.66 mmHg; range 43.55-149.37 mmHg; P < 0.0001), one month (84.02 mmHg, range 52.22-157.33 mmHg;P < 0.0001), 3 months (89.01 mmHg, range 49.37-154.9 mmHg;P < 0.0001), six months (91.70 mmHg, range 56.55-151.0 mmHg;P < 0.0001), twelve months (116.29 mmHg, range 54.46-176.79 mmHg;P < 0.0001), eighteen months (117.90 mmHg, range 89.91-165.66 mmHg;P < 0.0001), and twenty-four months (97.75 mmHg, range 28.0-154.31 mmHg;P < 0.0001). Six dogs (21%) are deceased. Of these six dogs, three dogs were euthanized for progressive myocardial failure and left-sided congestive heart failure, one dog was euthanized for return of syncopal episodes, and two dogs died suddenly. Including three deceased dogs, nine dogs (32%) developed syncopal episodes post-CBV and HPBV. Four dogs (14%) are currently receiving sotalol therapy (mean dose 1.69 mg/kg q12 hrs) for ventricular arrhythmias; all other dogs are currently receiving atenolol therapy (mean dose 0.73 mg/kg q12 hrs). CBV followed by HPBV resulted in a significant reduction in LVOT PG in this group of SAS dogs and gradient reduction was maintained at mid-to-long term follow-up in the majority of dogs. However, the degree to which this reduction affects long-term survival and subjective quality of life continues to be studied. Circulating microparticles (MP) are subcellular structures (0.1 to 1 lm) derived from the cells of the vascular system. MP counts are increased in many human diseases and their roles in coagulation, inflammation and endothelial function are increasingly recognised. We hypothesised that MPs would be detectable in dogs, would be higher in dogs with mitral valve disease (MVD), and would increase with increasing MVD severity. The aim of this study was to measure MPs in dogs at different stages of MVD.

Dogs with MVD and age and weight matched healthy controls (group I) were prospectively recruited through a first opinion practice in London. Dogs were classified according to the 2009 ACVIM classification system into early (B1; group II) and advanced MVD group (B2 and C; group III). Citrated blood was collected by jugular venepuncture and centrifuged at 1,000 g for 15 minutes within 30 minutes of collection to obtain plateletpoor plasma. This was then analysed using flow cytometry within 4-6 hours. Latex beads (1.1 lm) were used as size reference to determine the position of the MP gate. Having checked their cross-reactivity, samples were stained with commercially available fluorescently labelled antibodies against endothelial cells (CD31), platelets (CD41), erythrocytes (CD235) and the apoptotic marker MC540. Total and positively stained MPs were determined. Data were compared between groups using the Kruskal-Wallis multiple-comparisons test.

Samples were obtained from 13 dogs in group I, 11 in group II and 10 in group III. Group I dogs had lower overall MPs compared to groups II and III (median= 4,914 events (IQR= 4,145 -6,047) vs 8,556 events (7,590 -13,082) and 8,781 events (5,117 -13,915) respectively; p = 0.005). vs 154 events (114 -275) and 176 events (125 -226); p = 0.016) were similarly higher in the MVD groups. Group III had higher numbers of CD235 MPs compared to groups I and II (group I: 289 events (197 -428), group II: 208 events (55 -392) vs. group III: 480 events (311 -537); p = 0.028), and group II had higher CD31 MPs compared to group I but not to group III (group I: 9 events (5.5 -12), group II: 15 events (9 -29) vs. group III= 10 events (4 -23); p = 0.038). No differences were detected between groups for MPs staining positively for CD41 (p = 0.833).

In conclusion, these preliminary data show MPs are detectable in fresh plasma samples from dogs and their numbers vary depending on the stage of MVD. Further studies are warranted to determine the significance and possible role of MPs in dogs with MVD. 

Pinschers is time-consuming and remains a challenge especially in the early stages of the disease. Measurement of the myocardial performance index (MPI) could be useful to distinguish Doberman Pinschers with DCM from healthy dogs.

The aim of this study was to evaluate left ventricular MPI calculated either using conventional pulsed wave Doppler (PW-MPI) or tissue Doppler imaging (TDI-MPI) in Doberman Pinschers with and without DCM and to compare the different examination methods. 366 echocardiographic examinations from 193 purebred Doberman Pinschers were included (107 examinations from healthy dogs and 259 examinations from Doberman Pinschers with DCM in different stages of the disease). The single examinations of the Doberman Pinschers with DCM were grouped according to the results of the 24-hour-ECG and the echocardiographic examination into 3 different groups: last normal examination, only ventricular premature contractions (VPCs), or systolic dysfunction.

In general, particularly TDI-MPI increased with progression of DCM. The TDI-MPI revealed statistically significant differences between healthy Doberman Pinschers and Doberman Pinschers in all stages of DCM, including the last normal examination of those dogs. For PW-MPI significant differences between healthy Doberman Pinschers and Doberman Pinschers with VPCs or systolic dysfunction could be shown (P < 0,05). There was only a moderate correlation between the results of the two different MPI evaluation techniques.

TDI-MPI could be an useful additional test in the diagnosis of DCM in Doberman Pinschers, also in the early stages of the disease. The AliveCor â ECG device incorporates electrodes into an Apple iPhone â case allowing wireless recordings of electrocardiograms. We hypothesized that the AliveCor â would permit accurate heart rate and rhythm identification in dogs, cats and horses with normal sinus rhythm and spontaneous arrhythmias when compared to a reference ECG. Standardized 6 -lead ECGs and AliveCor â recordings were acquired simultaneously from 46 dogs and 23 cats; simultaneous base-apex ECGs and AliveCor â recordings were acquired from 18 horses. Instantaneous heart rates were obtained from identical QRS complexes where these were identified; 15-second average heart rates were obtained where identical QRS complexes were not identified. 3 observers independently evaluated the rhythm and the polarity of QRS depolarization for each recording. The results were compared within observer and between observers.

Instantaneous and average heart rates were identical in all cases where exact matches could be made for comparison between the iPhone and reference ECG, and were within 1 beat where average heart rates were calculated. Intra-observer agreement for rhythm assessment was very high, with no disagreement for equine ECGs, maximal disagreement in 2/46 canine and 4/23 feline ECGs. The polarity of depolarization was occasionally different between the AliveCor â and reference ECG in horses and dogs, but frequently different in cats. Inter-observer agreement for AliveCor â ECGs was similar to that for reference ECGs, with all observers agreeing on the rhythm analysis and polarity most of the time.

Our data suggest that the AliveCor â accurately identifies cardiac rhythms in horses, dogs and slightly less accurately in cats. Heart rate variability (HRV) is one of the most valuable noninvasive markers of autonomic nervous system and is a prominent method to assess its modulation of cardiovascular system. In human patients with sepsis, it is well known that there is a decrease in HRV and in the past few years the relationship between this decrease and the development of multiple organ dysfunction has been studied. Despite the autonomic imbalance in human patients with sepsis is well documented, it has never been investigated in dogs with naturally occurring sepsis. Thus, the aim of this study was to evaluate the HRV in bitches with sepsis due to cystic endometrial hyperplasia-pyometra complex.

For this purpose, were included in the study 10 bitches with clinical, hematological and ultrasonographic diagnosis of cystic endometrial hyperplasia-pyometra complex with scoring criteria for sepsis according to the assessment of heart rate, respiratory rate, body temperature, white blood cells and immature band forms. Prior to hysterectomy (T-0), 15 days (T-15) and 30 days (T-30) after, animals underwent 1-hour Holter digital recording with a 4 lead, 3 channel electrode system from where HRV variables were obtained. The variables in the time domain were SDNN, SDANN, SDNNidx, rMSSD, pNN50. Data were submitted to normality test of residuals and transformed using logarithmic (SDNN, SDANN, SDNNidx and rMSSD) and square root (pNN50) function to provide equal variances. Analysis between moments was performed by using One-Way ANOVA and Tukey's post hoc test (P < 0.05).

In every tested variable, there was no statistical difference between means of T-15 and T-30, but both moments had significantly higher values when compared to T-0. Means (AE standard deviation) of the variables in the moments T-0, T-15 and T-30 respectively were: SDNN, Transesophageal atrial pacing (TAP) is a non-invasive method of atrial pacing that can be performed without specialized imaging equipment. Pacing is achieved by placement of a pacing catheter within the esophagus of a patient and applying a pacing stimulus. In humans, the use of transesophageal atrial waveforms has been shown to guide the placement of the pacing catheter to the site where atrial pacing was achieved with the minimum pacing threshold (MPT). TAP without the use of fluoroscopic guidance has been described in dogs but evaluation of the use of transesophageal waveforms to guide the placement of the pacing catheter has not yet been evaluated. The purpose of this study was to determine if the transesophageal atrial (A), ventricular (V) wave amplitudes or A/V wave ratio can be used to guide optimal positioning of a transesophageal pacing catheter.

Fourteen client owned dogs weighing 16.9 AE 7.7 kg (mean AE sd) were used in this study. TAP was achieved with a 6 Fr pacing catheter under general anesthesia. The pacing catheter was inserted orally into the esophagus to a position caudal to the heart. With the pulse generator set at a rate 20 bmp above the intrinsic sinus rate using a pulse width of 10 ms and amplitude of 20 mA, the catheter was slowly withdrawn until atrial pacing was noted on surface electrocardiogram. Then the catheter was withdrawn at 1 cm increments until atrial capture was lost. MPT and transesophageal ECG were recorded at each site. Transesophageal ECG amplitudes of the A and V waves were then measured and correlated to MPT by linear regression.

TAP was achieved in all dogs. In 9/14 and in 7/14 dogs the site of MPT was the same as the site of maximal A and V waveform deflection respectively. The average distance from max A and V waveform deflection location to MPT location was 0.64 AE 0.57 cm and 0.29 AE 0.83 cm, respectively. In 6 dogs with at least four pacing sites, the relationship between maximum transesophageal atrial and ventricular amplitude was poorly correlated with MPT (R 2 = 0.2 and R 2 = 0.04, respectively). The average range of the amplitude of A and V wave deflections was 0.49 AE 0.56 mV and 0.61 AE .42 mV, respectively making visual evaluation of waveform size difficult. As such, use of transesophageal ECG waveforms to direct catheter placement was not found to be clinically useful. Myocardial fibrosis is a hallmark of hypertrophic cardiomyopathy (HCM). Human HCM patients have high serum concentrations of biomarkers of collagen synthesis, indicating a profibrotic state. This abnormality exists in sarcomere mutation carriers both with and without overt left ventricular hypertrophy, suggesting a role in early identification of HCM. Whether serum levels of biomarkers of collagen synthesis are increased in cats with HCM is unknown.

We prospectively enrolled and evaluated cats presented to a university veterinary cardiology referral service. Cats with a high serum T4 concentration, high systolic arterial blood pressure, recent or ongoing diuretic administration, and/or overt signs of hypovolemia, were excluded. The diagnosis of HCM was established using echocardiography (echo; IVS and/or LVFW ! 6 mm on M-mode and/or 2D, in the absence of other lesions). Cats with IVS and LVFW <6 mm and no other echo abnormalities were considered normal.

Forty-seven cats qualified for and participated in the study: 28 with an echo diagnosis of HCM and 19 normal controls. We evaluated serum concentrations of C-terminal propeptide of type I procollagen (PICP), as a marker of collagen synthesis, using 2 commercially available ELISAs (1 validated for use in humans, 1 in rats). We used a commercially available ELISA previously validated for use in cats for quantifying serum C-terminal telopeptide of type I collagen (CTx) as a marker of collagen degradation. We compared serum [CTx] Mitral regurgitation (MR) following chronic valvular heart disease (CVHD) is the most common acquired cardiac disease in dogs. Although neurohumoral factors play important roles in the progression of CVHD, mechanical stimuli are known to affect various cells in the body. In MR, various mechanical stimuli might be caused by high-speed blood flow. In addition, in vitro experiments show that vibration at 50 Hz increases osteoblast growth. The peak frequency of cardiac murmur due to MR is 100-200 Hz. Therefore, we hypothesized that vibration due to cardiac murmur affects cells of the mitral valve. The purpose of this study was to investigate whether there is a relationship between vibration and cells of the mitral valve.

Interstitial cell cultures derived from canine mitral valve were used for this study. The cultured interstitial cells were vibrated in an incubator for 30 days. The frequencies of vibration were 12.5 and 100 Hz. Control cell cultures without vibration were concurrently cultured in the same incubator. Cell growth curves and expressions of vimentin, a-smooth muscle actin (a-SMA), and collagen I/III were compared between the vibration group and the non-vibration group. The cell growth curves and the expressions of vimentin and collagen I/III did not differ between the 2 groups, but the expression of a-SMA was significantly higher in the 12.5 Hz-vibration group than in the control group (p = 0.0001).

In conclusion, vibration might affect interstitial cells of the mitral valve. However, further studies are needed to determine the vibration effects on the mitral valve. Transesophageal atrial pacing (TAP) is a non-invasive method of atrial pacing in people and dogs. Pacing is achieved through the use of electrical stimulation applied through pacing catheters available with variations in catheter characteristics including catheter shape (curved vs. straight), electrode size (ES) and inter-electrode spacing (IS). An ideal catheter would allow atrial pacing at low stimulus thresholds (PT) over a wide area (zone of capture, ZOC) and without inducing extraneous muscular stimulation (EMS). The optimal catheter configuration is unknown in dogs. Therefore, the purpose of this study is to determine the effects that catheter characteristics has on TAP threshold and ZOC in dogs.

Seven catheter configurations (Table 1) were tested in 10 purpose bred dogs (18.8 AE 8.9 kg, meanAESD) maintained under general anesthesia. Outcome measures consisted of PT, ZOC and EMS. Student's paired t-tests and linear regression analysis were used to determine significance. Statistical significance is set at p < 0.05.

TAP was achieved in all configurations in all dogs. The curved catheter had significantly lower PT, wider ZOC and less EMS than the straight catheter. When electrode size was evaluated, 6 mm poles produced significantly more EMS than either 2 or 4 mm poles. Although not significant, 4 mm poles generated the largest ZOC with the lowest PT. When inter-electrode spacing was evaluated, 5 mm poles produced significantly less EMS and required significantly higher PT than 15 or 25 mm spacing. Additionally, there is a significant direct correlation between inter-electrode spacing and ZOC.

The results of this study suggest that a curved catheter with multiple 4 mm electrodes to allow for variable electrode spacing would be ideal for TAP in dogs. Such a catheter would allow the operator to tailor the pacing outcome to the needs of the clinician and patient. Cardiac Troponin-I (C-TI) and N-terminal prohormone of Btype Natriuetic Peptide (NT-proBNP) are biomarker compounds for myocardial disease and left atrial overload/congestive heart failure, respectively. NT-proBNP has been shown to have high temporal variability in normal dogs. However, the degree of temporal variability of this compound is unknown in dogs with cardiac disease. The aim of this study was to estimate the degree of biological and analytical variability for ultrasensitive C-TI and NT-proBNP in clinically healthy dogs and dogs with well characterized cardiac disease of varying severities. We hypothesized that dogs with cardiac disease would show lower inter-and intra-individual variation, resulting in lower critical change values for these analytes in dogs with cardiac disease when compared to healthy control dogs.

Three groups of dogs were prospectively enrolled, clinically healthy dogs (n = 12), dogs with mitral valve disease but no evidence of congestive heart failure (n = 5), and dogs with mitral valve regurgitation and evidence of left atrial overload requiring at least diuretic therapy (n = 4). Serum and plasma samples were collected once weekly for 7 weeks in 20/21 dogs; one dog was lost to follow up after 4 weeks. Serum C-TI concentrations were measured in duplicate using a commercial ultra-sensitive troponin assay (Siemens Healthcare), while NT-proBNP was measured with the Cardiopet â NT-proBNP ELISA (IDEXX Laboratories). Data were analyzed via nested ANOVA to derive values for analytical (CV A ), intra-individual (CV I ), and within-group (CV G ) variability. These values were then used to derive Index of Individuality (IoI) and Critical Change Values (CCV) for both markers in the control dog and cardiac disease populations (initial analyses showed no difference between the two groups with cardiac disease, thus data from these groups were combined).

Both compounds showed high individuality in all groups (IoI's of 3.14 and 3.93 for C-TI, 2.48 and 4.94 for NT-proBNP, control vs. cardiac disease, respectively), indicating that comparison to a population based reference range has limited utility. Two-sided critical change values (CCV's of 112.5% vs 64.5% for C-TI, 99.4% vs 51.8% for NT-proBNP) were lower for dogs with cardiac disease, indicating that threshold values for meaningful biological change for both CT-I and NT-proBNP are lower for dogs with established cardiac disease than in healthy controls. Together the high IoI's and lower CCV values for dogs with cardiac disease suggest that the greatest clinical utility from these Subaortic stenosis (SAS) is one of the most common congenital cardiac defects in dogs. Severe SAS is frequently treated with beta-blockers (BB), though this approach is largely empirical. The purpose of this study was to determine if BB therapy influences survival in dogs with severe SAS. The medical records from the University of Missouri Veterinary Medical Teaching Hospital and the University of Minnesota Veterinary Medical Center were reviewed. Dogs with severe, uncomplicated SAS defined as a pressure gradient (PG) ! 80 mmHg and diagnosed using 2-D and Doppler echocardiography were included in the study. Dogs with hemodynamically significant concurrent cardiac disease or dogs for which no follow up information was available were excluded from analysis.

Between 1999 and 2011, 51 dogs met the inclusion criteria. 27 dogs were treated with a BB (26 with atenolol, 1 with propranolol), and 24 received no treatment serving as a control group (CON). At time of analyses, 24 dogs in the BB group and 15 dogs in the CON group were dead. Cardiac death was defined as euthanasia or death following onset of signs of congestive heart failure, or witnessed sudden death. Cause of death could not be determined in 8 dogs (6 BB, 2 CON) and these cases were censored in the cardiac mortality analysis. Median age at diagnosis (AGE) for all dogs was 9.5 months (range: 2 months-11.6 years), and median PG was 121 mmHg (range: 80-227.6). There was no difference in PG between groups; however the AGE of the CON group was significantly older then the BB group (15.5 months Vs 6.2 months, respectively; P = 0.01). Cox proportional hazards regression was used to identify the influence of PG, AGE, and BB therapy on survival. In the all-cause multivariate mortality analysis, only AGE (P = 0.014) and PG (P = 0.019) affected survival time. The adjusted hazard ratio (HR) results indicated that increased AGE was associated with an increase in survival time (HR = 0.73, confidence limits [CL] = 0.56-0.93), whereas an increase in PG was associated with a decrease (HR = 1.009, CL = 1.002-1.017). In the cardiac mortality analysis, only PG influenced survival time (HR = 1.012; CL = 1.002-1.021, P = 0.015). Therapy with a BB did not influence survival time in either the all-cause (HR = 0.995, CL = 0.46 -2.152, P = 0.99) or cardiac cause (HR = 0.885, CL = 0.347-2.258, P = 0.798) mortality analyses.

Based on these results, we conclude that BB therapy does not influence survival in dogs with severe SAS, and a higher PG at diagnosis was associated with increased risk of death. We speculate that the benefit of increased age at diagnosis in all cause mortality may be related to early deaths of individuals with more severe disease. Age at diagnosis was not significant in cardiac mortality analysis, but that may have been due to a larger number of censored dogs. The accurate assessment of arterial blood pressure in the leporine specie could be applied not only in anesthetic monitoring and clinical practice, but mainly given its importance as experimental model of several diseases. Therefore, the present study aimed to evaluate the accuracy of two different devices of noninvasive blood pressure measurement in non-sedated rabbits, in comparison with its invasive assessment.

For this, were used 12 clinically healthy New Zealand White rabbits, nine males and three females, weighting between 3.2 and 4.4Kg. In each animal, arterial blood pressure was measured using a digital oscillometric device designed for small animals (Pet-MAP â ) and a vascular Doppler device (Parks Medical Ultrasonic Doppler 812 â ). Cuffs with approximately 40% of left forearm circumference were placed at this site. By using the Pet-MAP â device, we assessed systolic, mean and diastolic blood pressure, while only systolic blood pressure was obtained by using vascular Doppler device. Values obtained by each noninvasive method were compared with those obtained simultaneously by invasive blood pressure monitoring (Dixtal 2010 â ), accessed on auricular artery after topical application of lidocaine and prilocaine, 25/25 mg/g cream (EMLA â ).

The mean difference of paired measurements for arterial blood pressure, the correlation between paired measures and the percentage of all measurements that lay within 10 and 20 mmHg of the reference method were evaluated.

When comparing values obtained by using Pet-MAP â device with those obtained by invasive method, the mean difference of paired measurements for systolic, mean and diastolic pressures were, respectively, 38.3 mmHg, 18,6 mmHg and 13.6 mmHg, all of them underestimating arterial blood pressure. Regarding vascular Doppler device, the mean difference of paired measurement was 1.3 mmHg. By using Pet-Map â device, the correlation was 0.6 (P = 0.04), 0.52 (P = 0.08) and 0.44 (P = 0.15) for systolic, mean and diastolic blood pressure, respectively, and 0.91 (P < 0,001) by using vascular Doppler device. For the Pet-Map â device, none of the measurements of systolic pressure lay within 10 mmHg of the reference method and 8.3% lay within 20 mmHg, 33.3% of the measurements of mean pressure lay within 10 mmHg and 66.7% within 20 mmHg, 41.7% of the measurements of diastolic pressure lay within 10 mmHg and 75% within 20 mmHg. For the vascular Doppler device, 91.7% of the measurements lay within 10 mmHg of the reference method and 100% within 20 mmHg.

In lateral recumbency and non-sedated rabbits, only the vascular Doppler device accurately reflected the auricular artery pressure, and this method can be used as an alternative to the invasive one. Mitral regurgitation (MR) due to myxomatous mitral valve disease (MMVD) is a common finding in cavalier Kings Charles spaniels (CKCSs). In addition, sinus arrhythmia is often more pronounced in dogs compared to other mammal species. The aim of the study was to evaluate whether duration of the R-R interval influences degree of MR assessed by echocardiography in dogs.

Clinical examination including echocardiography was performed in 103 privately-owned dogs (16 controls dogs (Beagles), 70 CKCSs with different degree of MR and 17 dogs of different breeds with clinical signs of heart failure due to MMVD).

Severity of MR was evaluated in apical 4-chamber view using colour Doppler flow mapping (maximum% of the left atrium area) and colour Doppler M-mode (duration in msec). The influence of the ratio between present and preceding R-R interval on MR severity was evaluated in 10 consecutive R-R intervals using a linear mixed model for repeated measurements. MR severity was observed to increase when a short R-R interval was followed by a long R-R interval in CKCSs with different degrees of MR (P < 0.003 when adjusted for multiple testing). The relationship was not significant in control dogs (also including dogs with minimal MR) or in dogs with clinical signs of heart failure due to MMVD.

In conclusion, MR severity increases in long R-R intervals when these follow a short R-R interval in CKCSs with different degrees of MR due to asymptomatic MMVD. Thus, the degree of sinus arrhythmia may affect the echocardiographic grading of MR in dogs.

To investigate the utility of a single BNP measurement to predict cardiac death, we studied 137 CHF cats with all forms of cardiomyopathy that underwent complete diagnostic evaluations between 2007-2011. BNP results were separated into quartiles, and time to cardiac death was evaluated by Kaplan Meier analysis. Cat's survival clustered at BNP <265, between 265-515pmol/L, between 515-765, and >756pmol/L, suggesting 515pmol/L as a clinically relevant break-point. Bimodal comparisons made at 50pmol/L increments starting at <350 or >350pmol/L were made for all CHF cats and then, eliminating 25 DCM cats from the analysis, for the remaining 112 cats with diastolic HF.

In both cases, the greatest survival difference at 1 month post CHF relative to BNP occurred between those cats with BNP 550pmol/L vs. > 550pmol/L BNP (p = 0.04-combined population; p = 0.08-diastolic HF cohort). Predicting significant risk stratification beyond one month after CHF was not possible.

C-32 CARDIAC BIOMARKERS IN HYPERTHYROID CATS. JK Sangster, DL Panciera, JA Abbott, KC Zimmerman, AC Lantis. Virginia-Maryland Regional College of Veterinary Medicine, Blacksburg, VA.

Differentiation of hyperthyroid heart disease from primary myocardial disease is challenging. The cardiac biomarkers NT-proBNP and troponin I (cTNI) have proven useful in identifying cats with myocardial disease and may provide a method by which hypertrophic cardiomyopathy (HCM) and hyperthyroid heart disease can be discriminated. The primary purpose of this study was to compare plasma concentrations of NT-proBNP and cTNI in three groups of cats: cats with naturally occurring hyperthyroidism, cats with primary cardiomyopathy, and healthy older cats to determine if biomarkers differ between groups and if biomarker concentrations in hyperthyroid cats change after resolution of the thyroid disease.

We prospectively evaluated 61 client-owned cats: 23 hyperthyroid cats, 19 cats with HCM without congestive heart failure, and 19 euthyroid, normotensive healthy cats eight years of age or older. Fourteen of the hyperthyroid cats were re-evaluated three months after administration of 131 I. A complete history, physical examination, CBC, serum biochemistries, urinalysis, blood pressure measurement, serum T4 concentration, plasma concentrations of NT-proBNP and cardiac troponin I, and echocardiography was obtained for each cat. Hyperthyroid and HCM cats had plasma NT-proBNP and cTNI concentrations that were significantly greater than healthy older cats, but there was no significant difference between hyperthyroid and HCM cats with respect to concentration of either biomarker. Plasma NT-proBNP and cTNI concentrations decreased in each cat that was examined three months after 131 I treatment. Plasma cTNI was within the reference interval for all cats at the three month recheck. Severely thickened myocardium persisted in one formerly hyperthyroid cat at the three month recheck, and this cat's plasma NT-proBNP remained elevated. Although there may be a role for NT-proBNP in monitoring the cardiac response to treatment of hyperthyroidism, neither NT-proBNP nor cTNI can be used to distinguish hyperthyroid cats from cats with HCM. Therefore, the thyroid status of older cats should be ascertained prior to interpreting results of cardiac biomarker testing. Dynamic obstruction of the left ventricular outflow tract (DLVOTO) is a common finding in cats with hypertrophic cardiomyopathy (HCM) possibly leading to clinical signs and disease progression. We hypothesized that Ivabradine, a highly selective I f inhibitor, will reduce or eliminate DLVOTO in cats with preclinical HCM.

In this prospective, randomized, double-blind, active-control single dose study, cats with preclinical HCM and DLVOTO received one single dose of Atenolol (2 mg/kg PO) or one single dose of Ivabradine (0.3 mg/kg PO). Pre-and 3 hour post-pill heart rate and murmur assessment, and transthoracic echocardiography were performed, with specific focus on drug effect on outflow tract obstruction based on 2D and CW Doppler studies. Statistical analysis was performed using a two-way repeated measures ANOVA.

Of a total of 21 cats with HCM and DLVOTO, 14 received Ivabradine and 7 received Atenolol. Cats receiving Ivabradine (13/14) did not reveal a reduction of LVOT velocity (while the Atenolol group (7/7) did (P < 0.05)). Mean pre-pill V max LVOT was 4.30 + /-0.98 m/s for the Ivabradine group and 4.29 + /-1.14 m/s for the Atenolol group, compared to mean post-pill values of 3.97 + /-1.22 m/s and 1.69 + /-0.47 m/s, respectively. Mean pre-pill HR was 208 + /-26 bpm for the Ivabradine group and 215 + /-29 bpm for the Atenolol group versus postpill values of 146 + /-19 bpm and 158 + /-26 bpm, respectively.

This study failed to demonstrate an effect of a single oral dose of Ivabradine on DLVOTO in cats with preclinical HCM. people using similar agents; the purpose of this pilot study was to investigate the incidence of HT in dogs treated with toceranib phosphate.

The blood pressure records of the UW Veterinary Care Cardiology Service were reviewed to extract records of dogs that had received toceranib phosphate and had systolic BP (SBP) documented prior to and after drug administration. Dogs that had received anti-HT medications or had incomplete records were excluded. Data is presented as median [range] .

Forty-six dogs met the initial entry criteria; 20 dogs had preand post-treatment SBP measurements available. Use of toceranib phosphate was associated with increases in SBP in the majority of dogs in this small sample. Most previously HT dogs remained HT post-toceranib, and 42% of normotensive dogs pre-toceranib had post-toceranib SBP ! 160 mmHg. Blood pressure monitoring pre-and post-toceranib therapy may be warranted to screen for this possible side effect. Studies have demonstrated that NT-proBNP is a strong risk marker for first onset of congestive heart failure in dogs with heart disease. However, accurate determination of clinically relevant NT-proBNP concentration cut-offs in dogs for prognostic application has been challenging due to the limited dynamic range of the Cardiopet â proBNP assay currently available on the market.

The bioanalytical method validation of a prototype secondgeneration immunoassay for the quantification of canine NT-proBNP is presented. Sensitivity was determined by measuring the limit of detection defined as 3 standard deviations above background. Accuracy was measured via dilutional parallelism of 4 different high concentration plasma samples and calculating observed to expected ratios. Intra-assay, inter-assay, and total precision was determined for 4 samples distributed across the range based on 9 replicates per run x 3 runs per operator x 3 operators over a three day period. The dynamic range is established by the upper and lower limits of quantitation between which the assay demonstrates variability 20% CV and accuracy ! 80%. Finally, a Bland-Altman method comparison between the first and second-generation canine NT-proBNP assays was performed using 149 canine plasma samples.

The assay sensitivity was established at 36 pmol/L. The average observed to expected ratio for serial dilutions was104.1% AE 15.0% for 4 different plasma samples. Coefficients of variation for intra-assay, inter-assay, and total precision for 4 different samples were as follows: [NT-proBNP] A method comparison between the first and second-generation canine NT-proBNP assays performed using 149 canine plasma samples indicated an overall bias of 26 pmol/L that was statistically insignificant (P = 0.7096).

Based on precision and accuracy, the effective dynamic range of the assay is 250 pmol/L to 10,000 pmol/L. Furthermore, the high accuracy of sample dilutions through the dynamic range suggests the potential for extending the upper limit beyond 10,000 pmol/L. Results of this study indicate the improved performance of a second-generation Canine NT-proBNP ELISA to support measurement of NT-proBNP for prognostic applications. Spironolactone, an aldosterone antagonist, has been shown to decrease the risk of cardiac-related death, euthanasia, or worsening of cardiac failure in dogs with moderate to severe mitral regurgitation caused by myxomatous mitral valve disease, when added to conventional cardiac therapy. In cats, cardiomyopathy (CM) is the predominant cause of heart failure. Activation of the renin-angiotensin-aldosterone system (RAAS) occurs in cats with cardiomyopathy and signs of congestive heart failure. Spironolactone inhibits aldosterone-induced sodium retention in the kidney, leading to decrease cardiac preload, prevents aldosterone induced fibrosis and improves endothelial function. In Maine Coon cats with familial Hypertrophic cardiomyopathy (HCM), spironolactone at 2 mg/kg POq12 h for 4 months was not shown to improve the mitral annular velocity or reduce the left ventricular mass and 4 of the 13 treated cats developed severe ulcerative facial dermatitis.

To evaluate further the safety and efficacy of spironolactone in cats with congestive heart failure (CHF) secondary to a CM, a double blind, randomized placebo-controlled study is being conducted with cats receiving spironolactone at 1,7 to 3,3 mg/ kg PO sid or placebo for up to 15 months. This dosage was extrapolated from the current approved dosage in dogs (i.e. 2 mg/kg sid). As the study is still ongoing, the authors remain blinded as to which cats are receiving spironolactone so the following results relate to the whole population, receiving spironolactone or placebo, which are being administered in addition to conventional cardiac therapy (including at least furosemide and benazepril).

Sixteen cats (14 Domestic Shorthair cats, 1 Ragdoll and 1 Burmese) with CM of various types (12 hypertrophic, 2 dilated, 1 unclassified and 1 Arrhythmogenic Right Ventricular) have been enrolled so far.

To date, 7 cats have died at 6 to 228 days post inclusion, 4 cats have been euthanized at 34 to 215 days post inclusion, 2 cats were withdrawn from the study (at Day 26 and Day 67), 2 cats remain in the study and one completed the 15-month follow-up.

Any adverse event (of cardiac origin or not) was systematically recorded. Severe cardiac failure leading to death or euthanasia was recorded for 8 cats which presented either with worsening cardiac failure (4), aortic thromboembolism (2) or acute respiratory distress (2). Slight to moderate vomiting was observed in 2 cats and was followed in one of them by anorexia and loss of weight leading to euthanasia. The mean value of all hematology and serum or urine biochemistry parameters remains within the laboratory reference range. Hypokalaemia (< 3,5 mEq/l) was recorded in 6 cats at inclusion and in 4 cats at least one study visit after inclusion. Hyperkalaemia has not been recorded. No dermatitis has been recorded to date.

In conclusion, no severe adverse events except worsening of heart failure, aortic thromboembolism and one case of severe anorexia have been recorded in 16 cats with CHF secondary to CM treated with spironolactone at 1,7 to 3,3 mg/kg PO sid or placebo for a mean duration of 158 AE 146 days (6 to 427 days) in addition to conventional cardiac therapy including furosemide and benazepril. The vertebral heart scale (VHS) method is an established indicator of global heart enlargement but is relatively nonspecific for enlargement of individual heart chambers. A recent study comparing VHS and radiographic left atrial bisecting line (LABL) measurements to each other and to left atrial to aortic root ratio (LA/Ao) found that LABL was not significantly different in dogs with and without congestive heart failure (CHF) as opposed to VHS measured on the left lateral view. Researchers argued that the lack of correlation between LABL and LA/Ao measurements was due to difficulty in accurately estimating the limit of the left atrium (LA) as it is obscured by normal anatomical structures and by perihilar pulmonary edema in the setting of CHF. The aim of this prospective study was to evaluate the clinical usefulness of new radiographic measurements to detect left atrial enlargement (LAE). These measurements were compared to VHS, LABL and left atrial to aortic root ratio (LA/Ao) in healthy dogs and dogs with degenerative mitral valve disease (DMVD). Multicenter study. Healthy patients (n = 10) and dogs with DMVD (n = 24) were prospectively selected to be included in the study. New measurements were obtained as the distance from the intersection of the long (L) and short (S) axis of the heart to caudal heart limit (R-half-VHS), the distance from the intersection of the L and S axis of the heart to ventral border of carina (top-half-VHS), the sum of these measurements (Sum R-Top) and the distance from midpoint of R-half-VHS to LA roof (LA roof measurement). LABL was defined as a line arising at a 45-degree angle from the intersection of L and S axis of the heart and extending to the border of the left atrium.VHS was measured as described previously (Buchanan, 2000) . LA/Ao ratio was obtained by two-dimensional ultrasonic mode from right parasternal short axis view at the level of the aortic valves in early diastole. The clinical usefulness of plasma natriuretic peptide concentrations in dogs with right-sided heart failure (RHF) remains unclear. We investigated whether plasma levels of atrial natriuretic peptide (ANP) and N-terminal pro B-type natriuretic peptide (NT-proBNP) are useful to diagnose and assess the severity of RHF in dogs.

This retrospective study enrolled 16 healthy dogs and 56 untreated dogs with right-sided heart disease and RHF. Results of physical examination, thoracic radiography, and echocardiography were recorded. Plasma ANP and NT-proBNP concentrations were measured using commercial laboratories. According to the International Small Animal Cardiac Health Council (ISACHC) classification, 15 dogs were class I, 14 were class II, and 27 were class III. Plasma ANP and NT-proBNP levels were significantly increased in IS-ACHC class III dogs compared to the healthy control dogs. However, plasma ANP and NT-proBNP levels showed significant overlap between ISACHC class I and class II dogs. The ANP cutoff value > 47.6 pg/mL used to detect ISACHC class III dogs had a sensitivity of 84.6% and specificity of 73.8%. Similarly, the NT-proBNP cutoff value > 3003 pmol/L had a sensitivity of 91.7% and specificity of 88.6%. The area under the curve for the receiver-operating characteristic analyses of plasma ANP and NT-proBNP levels were 0.87 and 0.93, respectively.

These results suggest that plasma ANP and NT-proBNP concentrations markedly increased in dogs with RHF. Measurements of these peptides, especially NT-proBNP, can be simple and useful biomarkers to diagnose the severity of RHF.

C-39 ECHOCARDIOGRAPHIC ASSESSMENT OF CARDIAC DIMENSIONS AND SYSTOLIC FUNCTION DURING CANINE PUERPERIUM. PR Batista 1,2 , C Gobello 1,2 , YA Corrada 1,2 , DO Arias 1 , M T ortora 1 , PG Blanco 1,2 . 1 Faculty of Veterinary Science, National University of La Plata, Argentina, 2 CONICET Ventricular hypertrophy and increased systolic function occur during physiological canine gestation to guarantee adequate uteroplacental perfusion. However, it is not clear if these changes persist after parturition. Therefore, the aim of this study was to evaluate maternal echocardiographic changes in the course of normal canine puerperium.

Ten healthy pure-bred, 2-5 (3.5 AE 0.33) year-old, weighing 2.5-6 kg (3.4 AE 0.26) bitches were included in this study. All the animals whelped healthy puppies at term which were weaned on day 60 after parturition (day 0). Echocardiographic evaluations were carried out on days -3, 3, 10, 17, 24, 38, 52 and 80. Interventricular septum in diastole (VSd, mm) and systole (VSs, mm), posterior wall in diastole (LVWd, mm) and systole (LVWs, mm) and left ventricular diameter in diastole (LVDd, mm) and systole (LVDs, mm) were ultrasonographically measured in the short axis view, during M-mode tracing. Shortening fraction (SF,%) was calculated as ((LVDd-LVDs)/ LVDd) x 100. Stroke volume (SV, mL) was calculated as the product of velocity time integral and the cross sectional area of the aorta. Cardiac output (CO, L/min) was calculated as the product of SV and hearth rate (HR, bpm; obtained by electrocardiographic monitoring). Values of VSd, VSs, LVWd, LVWs, LVDd, LVDs, SF, SV, CO and HR were analyzed by repeated measures ANOVA followed by Tukey test (SPSS 19.0; SPSS, Chicago, IL, USA) .

During the study period, VSs (P < 0.05), SF (P < 0.01), SV (P < 0.01), CO (P < 0.05) and HR (P < 0.05) progressively decreased, while LVDs increased (P < 0.05). During the first postpartum week, VSs and SF diminished markedly. Conversely, LVDs showed a pronounced increase at the same time. Stroke volume, CO and HR progressively diminished in the course of the study, while VSd, LVDd, LVWd and LVWs remained unchanged (P > 0.1).

It is concluded that VSs, SF, SV, CO and HR progressively decreased, while LVDs increased in the first 80 days of puerperium, indicating reversibility of most gestational adaptive changes. A recent ACVIM consensus statement on canine chronic valvular disease (CVD) recommended that pimobendan (0.25-0.3 mg/kg PO q12 h) be used with furosemide and an ACEinhibitor as chronic therapy for heart failure (HF). For refractory HF, some panelists administer a third 0.3 mg/kg daily dose (outside the FDA-approved dosage).

This retrospective study evaluated the frequency of use of three-times daily (TID) and high-dose-pimobendan ( ! 0.9 mg/ kg/day:HDP) at NCSU, potential survival benefit when TID pimobendan was initiated as 'rescue' therapy for refractory HF, and frequency of adverse side-effects.

Medical records of all dogs prescribed pimobendan (outpatient) ! TID between 4/2007-8/2011 were evaluated. Of 118 dogs, 63 ultimately received HDP ( ! 0.9 mg/kg/day; mean 1.2; range 0.9-2.7). HF was due to CVD in 84% of cases. At the time of writing, 57 dogs had been euthanized or died, most due to cardiac disease. Median survival from initiation of TID pimobendan was 251 days (1-1103 days). This population was compared to a historical NCSU cohort, diagnosed with HF (2002 HF ( -2005 , where pimobendan, at the recommended dosage, was added as 'rescue' therapy to standard treatment (ACE-inhibitor, loop-diuretic, digoxin, etc) . HF was due to CVD in a majority of these 68 dogs and median survival from the initiation of pimobendan was 110 days (3->350 days; p < 0.05). Side-effects, including sudden death, were rare in both groups.

HDP, commonly used in the authors' clinic, may improve survival in dogs with refractory HF due to CVD and appears to be well tolerated.

JD Thomason 1 , AE Coleman 1 , AC Dixon 1 , T Fallaw 1 , ID Boothe 2 , G.S Rapoport 1 . 1 University of Georgia, Athens, GA, 2 Auburn University, Auburn, AL.

Procainamide is a class Ia antiarrhythmic agent. The reported therapeutic trough concentration for procainamide is 4-12 mcg/ml. The drug's short half-life in the dog necessitates a dosing interval of 6-8 hours. A previously available sustained-release formulation, which allowed effective dosing when administered every 8 hours, has since been removed from the pharmaceutical market. We have consulted a private compounding pharmacy which offers a delayed-release procainamide preparation (DRP). The aim of the present study was to evaluate the pharmacokinetics of this DRP in normal dogs and to determine the feasibility of its clinical use.

For the initial portion of the study, two doses of DRP (30 mg/ kg PO) were administered 12 hours apart to two healthy, purpose-bred mixed breed dogs. Jugular venous blood samples were collected at 0, 0.5, 1, 2, 4, 6, 8, 10, 12, 12.5, 13, 14, 16, 18, 20 , and 24 hours post-initial dose. Samples were centrifuged and serum harvested and frozen within 30 minutes of collection. Pharmacokinetic analysis was performed as outlined below. Based on this data, a dose of 30 mg/kg q 12 h was deemed adequate to achieve a target trough serum procainamide concentration of 4 mcg/ml.

In the second portion of the study, two doses of DRP (30 mg/ kg PO) were administered 12 hours apart to 6 healthy, purposebred mixed breed dogs. Blood samples were collected and processed in the same manner and at the same time points as described above. Pharmacokinetic analysis was performed on all samples following the 24-hour collection period.

For pharmacokinetic analysis, serum samples were thawed and mixed to assure homogeneity. Procainamide was detected using the Siemens (New York, NY) PROC Procainamide Assay on a Siemens Dimension Xpand Plus â general chemistry analyzer. The upper and lower limits of quantitation were 20 and 0.5 mcg/mL, respectively. The system was calibrated with the Siemens Drug Calibrator II. Canine serum was spiked with known concentra-tions of procainamide HCL purchased from VWR (Radnor, PA) and used as quality control samples.

No adverse clinical effects were noted in any of the dogs. The average C max , C avg , and C min were 10.17, 7.13, and 3.07 lg/mL, respectively. TimeHgh (time over a 12-hour period that procainamide concentration exceeded 12 lg/mL), TimeDur (time over a 12-hour period that procainamide concentration was between 4-12 lg/mL), and TimeLow (time over a 12-hour period procainamide concentration was less than 4 lg/mL) were 2.35, 7.19, and 2.46 hours, respectively. T max and T min were 18.67 and 12.25 hours, respectively.

Although specific dosing recommendations cannot be made from this data due to individual animal variability, it appears that twice daily administration of DRP could be efficacious in dogs. Due to individual patient variability, evaluation of serum trough concentrations should be considered to confirm concentrations within the reported therapeutic range. Plasma N-terminal pro-B-type natriuretic peptide concentrations ( [NT-proBNP] ) are higher in dogs with structural heart disease than in normal dogs but have not been thoroughly investigated in dogs with PE. We hypothesized that PE would restrict atrial filling, precluding the release of NT-proBNP into circulation; consequently, plasma NT-proBNP concentrations would be higher in dogs with radiographic cardiomegaly associated with structural heart disease than in dogs with PE. We further hypothesized that dogs with PE could have characteristic plasma cardiac biomarker profiles that could be useful in clarifying the cause of PE.

Dogs prospectively evaluated over a 27-month period were categorized post-hoc into 6 groups: 1) PE and a cardiac mass on echocardiography (echo); 2) PE with no mass and no evidence of structural heart disease on echo; 3) PE and structural heart disease on echo; 4) cardiac mass and no PE on echo; 5) structural heart disease without PE on echo, associated with radiographic cardiomegaly; 6) structural heart disease without PE on echo, without radiographic cardiomegaly.

Blood samples were obtained for determining [NT-proBNP] and plasma concentrations of cardiac troponin I ([cTnI] ) and Nterminal pro-atrial natriuretic peptide ( [NT-proANP] [NT-proBNP] was significantly higher in group 4 than in groups 3 (p = 0.02), 5 (p < 0.001), and 6 (p = 0.0008). [NT-proBNP] was significantly higher in dogs with structural heart disease than in those without (group 3 > 1, 2, and 6 [p < 0.001], group 3 > 4 [p = 0.0001] and group 3 > 5 [p = 0.007]; group 5 > 1, [p = 0.0007], group 5 > 2 [p = 0.0002], group 5 > 4 [p = 0.003], and group 5 > 6 [p < 0.01]). [NT-pro-ANP] was higher in groups 3 and 5 than in all other groups (p < 0.0001).

[cTnI]: [NT-proBNP] was significantly higher in dogs with radiographic cardiomegaly due to PE than in dogs with structural heart disease (p < 0.001). Dogs with right atrial masses had significantly higher [cTnI]: [NT-proBNP] than did dogs with heart base masses (p = 0.002).

These data indicate that PE is associated with high [NT-proB-NP] when it occurs with structural heart disease associated with radiographic cardiomegaly but not when it occurs with a cardiac mass or structural heart disease without cardiomegaly. As previously shown, dogs with PE have high [cTnI] , particularly in association with a cardiac mass. [cTnI] indexed to [NT-proBNP] can further discriminate between dogs with cardiac masses and those without in the absence of PE as well as between dogs with right atrial versus heart base masses. VDD pacing, in contrast with VVI pacing, preserves atrioventricular (AV) synchrony and may reduce some of the deleterious effects related to chronic ventricular pacing. In a previous shortterm study in dogs, VDD pacing was associated with favorable hemodynamic effects, neurohormonal responses, and echocardiographic variables compared to asynchronous pacing. The longterm benefits of VDD pacing have not been previously evaluated in dogs with high-grade AV block.

This retrospective study compared survival times, resolution of clinical signs, complication rates, and owner satisfaction associated with VDD and VVI pacing. Forty-nine dogs with high-grade AV block were identified as undergoing pacemaker implantation from January 2006 to October 2012 at the Oregon State University Veterinary Teaching Hospital. A VDD pacemaker was implanted in 19 dogs (39%) whereas 30 dogs (61%) were treated with VVI pacing. The median survival times after implantation were 32.7 months (range: 2.8-44.4) for the VDD group and 24.5 months for the VVI group (range: 0.4-48.3), which were not significantly different (p = 0.39). Major complication rates associated with artificial pacing were 11% within the VDD group and 20% within the VVI group and were not significantly different (p = 0.46). Minor complications were significantly higher within the VDD group than within the VVI group (47% versus 7% respectively; p = 0.002). Resolution of clinical signs, owner satisfaction, and quality of life perception were considered excellent in both groups without statistical significance noted between groups.

Therefore no evidence of long-term clinical benefit of VDD over VVI pacing could be identified in the present study. Cardiogenic embolism (CE) is a clinically important condition in cats and humans while hyperreactive platelets have been suggested to be a possible hypercoagulable state associated with cardioembolic disease in both of these species. An in vitro platelet inhibitory assay could be helpful in identifying hyperreactive platelets in individuals, guiding antithrombotic therapy for thromboprophylaxis. The platelet receptor complex Gp IIb/IIIa is integral in platelet aggregation, regardless of platelet activator, so drugs that antagonize this receptor would appear ideal to identify hyperreactive platelets. The purpose of this study was to determine if the Gp IIb/IIIa antagonists abciximab and eptifibatide reduce platelet aggregation in vitro in healthy cats.

Ten healthy adult cats were recruited from the general population. Jugular venous blood was directly collected into tubes containing the anticoagulant hirudin. Whole blood aliquots from each cat were divided into 5 treatment groups: 1) control; 2) 50 lg/ml abciximab; 3) abciximab diluent control; 4) 4 lM eptifibatide; and 5) eptifibatide diluent control. All samples were incubated at room temperature for 10 minutes. Whole blood platelet aggregation was performed on each aliquot using ADP (6.2 lM) and thrombin receptor activator peptide or TRAP (32 lM) as agonists. The degree of platelet activation was determined by measuring the AUC for the aggregation curve after 15 minutes. Repeated measures of ANOVA were performed on transformed data and significance was defined as p < 0.05.

Eptifibatide resulted in a significant reduction in platelet aggregation compared to baseline control with both ADP [50.0 (8-122) vs. 306 (130-664); p < 0.001] and TRAP [75.5 (3-148) vs. 219 (97-578); p < 0.001]. There was no significant difference in platelet aggregation with abciximab or either diluent control using ADP or TRAP.

Eptifibatide resulted in a significant reduction in platelet aggregation while abciximab did not exhibit any effect on platelet aggregation even at a very high drug concentration. Given that abciximab is the fab fragment of a monoclonal antibody directed against the human GP IIb/IIIa receptor complex while eptifibatide is a cyclic heptapeptide, we hypothesize that either binding kinetics or downstream signaling with abciximab is ineffective in cats. The use of eptifibatide in an in vitro platelet inhibitory assay may be helpful in identifying hyperreactive platelets in cats. The purposes of this study were (1) to evaluate the activation of neurohormones, cardiac biomarkers, and pro-inflammatory cytokines in canine myxomatous mitral valve disease (MMVD), and (2) to investigate whether these circulatory parameters differ between MMVD dogs with and without congestive heart failure (CHF).This study prospectively evaluated 15 healthy dogs and 26 dogs with MMVD. MMVD dogs were divided into non-CHF group or CHF group on the basis of radiographic and echocardiographic assessments. At presentation day, blood was sampled from the jugular vein for the measurement of pro-atrial natriuretic peptide (pro-ANP), brain natriuretic peptide (BNP), endothelin 1 (ET1), renin, angiotensin 1 (AT1), AT2, aldosterone, angiotensin converting enzyme (ACE), antidiuretic hormone (ADH), tumor necrosis factor alpha (TNF-a), and interleukin 6 (IL-6). Additionally, the urine sample was obtained from the cystocentesis for the measurement of urine aldosterone to creatinine (UAC) ratio. The serum, plasma, and urine samples were stored at -80°C and all samples analyzed within 2 months.

In univariate and multivariate logistic regression analysis, serum ACE showed significant difference between CHF and nCHF in MMVD dogs (P < .05), whereas other neurohormones and all of cardiac biomarkers did not differ significantly between the groups. Therefore, the diagnostic performance of ACE was evaluated using the area under the ROC curve, with a 95% confidence interval. A cut off value of plasma ACE < 33.5 uint was able to discriminate CHF group with 93% sensitivity and 60% specificity, the area under ROC curve was 0.77.

In conclusion, in the present study, serum ACE can be used as effective predictors of CHF in MMVD dogs when neurohormones could not differentiate between CHF and non-CHF groups. Congenital tricuspid valve (TV) stenosis is rare in dogs. There are reports of individual cases in the veterinary literature, but limited information regarding treatment and outcome. The aim of this retrospective analysis was to evaluate clinical signs, echocardiographic features, results of treatment, and outcome in dogs treated with balloon valvuloplasty (BV).

The University of California, Davis VMTH medical record database was searched for dogs admitted to the hospital between 2000 and 2012 for BV of TV stenosis. Five dogs were identified. All were Labrador Retrievers. Median age was 2 yr (range 1-5 yr) and mean body weight was 32.6 kg (SD +/-3.6 kg). Presenting complaints included episodic weakness/syncope (n = 4), abdominal distension (n = 4), weakness/lethargy (n = 2), and exercise intolerance (n = 2). Examination findings included ascites (n = 4), weak femoral pulse (n = 3), muffled heart sounds (n = 3), pleural effusion (n = 1), and atrial fibrillation (n = 1). On echocardiography, the right atrium was severely enlarged in all dogs and diastolic doming of the TV was appreciated in 3 dogs. The median peak trans-tricuspid flow velocity in diastole was 2.8 m/s (range 1.6-3.2 m/s).

Balloon valvuloplasty was attempted in all 5 dogs. The procedure was aborted in dog 1 (jugular vein approach) due to inability to pass a catheter through the TV. BV was successfully performed in the remaining 4 dogs. The largest balloon size used in each dog ranged from 15 mm to 25 mm. Right heart failure resolved post BV in 2 dogs (dogs 2 and 3), but recurred within 2 years in both dogs. In dog 2 (jugular vein) the balloon appeared to never fully cross the valve, but ascites resolved post-operatively. Ascites redeveloped 406d post BV. Repeat BV (16 mm, right femoral vein) resulted in resolution of the ascites. The patient was euthanized 2 months later for unrelated reasons. Dog 3 (18 mm, jugular vein) had pleural effusion and ascites that resolved post-BV. Pleural effusion redeveloped 485d post BV. Repeat BV (20 mm, right femoral vein) resulted in resolution of the pleural effusion. This patient was lost to follow-up. Of the 2 remaining dogs, dog 4 (15 mm, right femoral vein) experienced fewer episodes of episodic weakness, but the mild amount of ascites present pre-BV remained. This patient continues to do well 2 years post-operatively on no treatment. Dog 5 (25 mm balloon, left femoral vein) had moderate tricuspid regurgitation (TR) but no right heart failure at presentation. Severe TR and ascites developed post-BV and the dog was euthanized 160d post BV.

In conclusion, TV stenosis appears to be overrepresented in Labrador Retrievers. BV can be an effective treatment, however recurrence of clinical signs may occur. Severe right heart failure due to worsened TR is a potential complication of BV and in this study occurred in the one dog in which the largest balloon (25 mm) was used. Most dogs presenting with high-grade second degree or complete atrioventricular block (HGAVB) are diagnosed as having idiopathic disease. But there have been reports of AV nodal conduction disturbances being linked to myocarditis. In the dog the presumptive diagnosis of myocarditis can now be aided with the use of the cardiac Troponin-I (cTnI) blood test. Vector-borne and other infectious diseases have been reported to cause myocarditis in man and dogs. We hypothesized that dogs diagnosed with high-grade atrioventricular block would have evidence of vector-borne infectious disease.

Twenty-five dogs with HGAVB were studied. Blood was used to evaluate the prevalence of positive serological, culture, and PCR evidence for vector borne infectious disease. Serum was also used to measure cTnI to determine if there is a correlation between serologic/culture/PCR evidence of vector-borne disease.

Fourteen milliliters of blood was collected using aseptic technique from each study dog. Serum was submitted for a tick panel (E. canis, B. canis, R. ricketssii, B. henselae, B. visonii (IFA) and B.burgdorferi, E. canis, Anaplasma, heartworm antigen (Snap 4DX). Additional Bartonella koehlerae serology was also performed. Serum and whole blood were submitted for BAPGM (Bartonella alpha-Proteobacteria growth medium) blood culture and Bartonella PCR. Serum cTnI was measured by the clinical pathology laboratory and with a point-of-care analyzer (iStat). Results of both measures were evaluated by commercially available statistical software for correlation.

Four of the 25 dogs in this study were positive for vector borne disease. All dogs in the study had elevated cTnI concentrations on both the laboratory test (median 1.1, range 0.23-25.5 ng/ml) and point-of-care analyzer (median 0.44, range 0.06-10.32 ng/ml). There was no correlation between the dogs that were positive for a vector borne disease and the degree of cTnI elevation (correlation coefficient -0.08; 95% confidence interval -0.47-0.34; P = 0.7). Strong correlation was observed for serum cTnI measures by point-of-care analyzer and clinical pathology laboratory (correlation coefficient of 0.93; 95% confidence interval 0.84-0.97; P < 0.0001).

C-48 TRANSCATHETER TRISCUPID VALVE REPLACEMENT (TTVR) IN DOGS WITH TRISCUPID DYSPLASIA. G Kramer, J Schneiderman, D Ozer. Atlantic Coast Veterinary Specialists, Bohemia, NY.

Chronic atrioventricular valvular disease (acquired and congenital) is a common problem in veterinary cardiology without a curative medical treatment. Medical therapy for severe atrioventricular valvular insufficiency is palliative at best in both dogs and in man. The definitive treatment for severe atrioventricular insufficiency in man is surgical valve replacement or repair. That treatment in veterinary medicine, while it may be superior to medical therapy, has intrinsic limitations. Transcatheter valve replacement may be a new therapeutic option that avoids the limitations and complications of open heart valve replacement or repair. The purpose of this study was to demonstrate that transcatheter valve replacement is a viable therapeutic treatment in dogs with atrioventricular valvular disease. Three Labrador retrievers with severe tricuspid valve dysplasia were selected for transcatheter valvular replacement. All three dogs had severe right-sided cardiomegaly and large tricuspid regurgitations. Two of the dogs were in chronic, severe rightsided CHF, which required repeated abdominocentesis and thoracocentesis. An experimental, prototype transcatheter valve was designed and manufactured by the author. It was passed down the jugular vein under fluoroscopic and ultrasonic guidance, fixed into the apex at the right ventricle and deployed between the tricuspid leaflets. The jugular vein was litigated and closure was routine. Intra-operative echocardiography showed marked reduction of the tricuspid regurgitant jet in all three patients. Deployment of the device did not result in any increase in tricuspid inflow gradients.

The two dogs that had been in chronic right-sided CHF died several hours after surgery due to complications (severe hypotension) attributed to the anesthesia and compounded by the advanced stage of the heart disease. The other dog developed ventricular arrhythmias after surgery that responded to medical management. There was no echocardiographic evidence that the device caused a reduction in cardiac output in any of the dogs.

This study shows that transcatheter tricuspid valve replacement is possible in dogs and can reduce regurgitant fraction. It appears this may be a viable treatment and further investigation is warranted. Cardiac troponin I (cTnI) is a biomarker measured in veterinary patients with congenital and acquired cardiovascular diseases for diagnostic and prognostic purposes, but few researchers utilize high-sensitivity assays. Thus, the goal of this study was to analytically validate a high-sensitivity immunoassay, developed and marketed for measurement of serum cTnI concentrations in humans, for use in dogs.

A commercially-available high-sensitivity immunoassay, AD-VIA Centaur CP â Ultra-TnI (Siemens Medical Solutions Diagnostics, New York, NY) was analytically validated for use in dogs. Four serum samples with different concentrations of cTnI were obtained from clinical patients. Assay validation included determination of dilutional parallelism, spiking recovery, intraassay variability, and inter-assay variability.

The reported analytical working range for the assay is 0.006 to 50 ng/mL. Observed to expected ratios for serial dilutions for the 4 samples ranged from 60.4 to 94.3% at dilutions of 1 in 2, 1 in 4, and 1 in 8. Observed to expected ratios for spiking recovery ranged from 86.99 to 109.64% for spiking each of the 4 serum samples with each of the other 3 serum samples. Coefficients of variation for intra-assay variability for the 4 serum samples were 4.9, 3.6, 5.7, and 4.3%. Coefficients of variation for inter-assay variability for the 4 serum samples were 3.4, 2.4, 4.7, and 5.9%.

Based on the limited linearity observed with sample dilution, we do not recommend diluting samples for clinical analysis. We conclude that this assay is sufficiently accurate, precise, and reproducible for use in dogs. The aims of this study were to (1) compare the indices of tissue Doppler with the conventional echocardiography in healthy dogs and dogs with myxomatous mitral valve disease (MMVD), and (2) determine if the tissue Doppler derived indices over conventional echocardiography can be effectively used for accurate diagnosis of congestive heart failure (CHF) in dogs with MMVD. This study prospectively evaluated 15 healthy dogs and 26 dogs with MMVD. MMVD dogs were divided into non-CHF group or CHF group on the basis of radiographic and echocardiographic assessments. The conventional echocardiographic parameters included percentage increase in the left ventricular internal diameter in diastole (LVIDd inc%), percentage increase in the LVID in systole (LVIDs inc%), ratio of left atrial diameter to aortic diameter (LA/AO), mitral regurgitation (MR), fractional shortening (FS), ejectional fraction (EF), ratio of E wave to A wave velocity (E/A ratio), dP/dt, and -dP/dt. The tissue Doppler echocardiographic parameters included radial and longitudinal velocities, strain, and strain rate. The ratios of transmitral peak E wave velocity to peak early diastolic velocity obtained by the tissue Doppler at the interventricular septal (IVS) basal segment (E/Em) were calculated.

In univariate analysis, several indices of conventional and tissue Doppler echocardiographic indices were significant (P < .05) predictors of CHF. However, after adjusting by multivariate logistic regression analysis, E/Em was the only independently significant (P < .05) predictor of CHF. The cut-off value of E/Em >18.7 has sensitivity 56%, specificity 90%, for prediction of CHF in MMVD dogs.

In conclusion, tissue Doppler-derived E/Em can be used as effective predictor of CHF in MMVD dogs to overcome the limitation of conventional echocardiography. Doxorubicin (DOX) is one of the most effective chemotherapeutic agents currently available. However, its use has been limited by cardiotoxic effects, especially when used for a long time. The purpose of this study was to investigate the diastolic function using Tissue Doppler Imaging (TDI) in rabbits during the treatment with DOX.

Twenty male New Zealand White rabbits were randomized in two experimental groups with 10 rabbits each, named: G1: control group, receiving NaCl 0.9%, G2: receiving DOX 1 mg/kg twice a week for 6 weeks. Echodopplercardiographic evaluations were performed in awaken animals, before DOX administration (T0) and every fifteen days (T15, T30 and T45). With rabbits positioned in left lateral recumbency, apical four-chamber view images were acquired for TDI evaluation, with a probe 6-8 MHz. The sample volume was placed at the lateral and septal insertion site of the mitral annulus. Measurements included peak early diastolic (Em), late diastolic (Am) mitral annular velocities (left wall and septum) and calculation of Em/Am ratio. The statistical analysis was performed by analysis of variance followed by Tukey′s test.

There was diastolic dysfunction (Em / Am < 1) for left wall at T30 (G1 = 1.45 AE 0.3; G2 = 0.85 AE 0.35) and T45 (G1 = 1.7 AE 0.59; G2 = 0.9 AE 0.3) and for septum at T30 (G1 = 1.56 AE 0.43; G2 = 0.77 AE 0.26) and T45 (G1 = 1.35 AE 0.28; G2 = 0.9 AE 0.48). The systolic function changed later, with a decrease (P < 0,05) of fractional shortening for G2 at T45 (G1 = 38.26 AE 4.6%, G2 = 21.36 AE 2.43%) and ejection fraction at T45 (G1 = 71,86 AE 5.48%, G2 = 47.14 AE 4.24%).

The results showed that TDI was able to identify diastolic dysfunction in rabbits during treatment with doxorubicin. The diastolic dysfunction preceded the systolic dysfunction in this model and this fact should be observed in patients submitted to therapy with doxorubicin. The aim of this prospective study was to evaluate the frequency of VPCs in the Irish Wolfhound (IW) breed and determine whether the presence of a ventricular arrhythmia is associated with clinical consequences, including sudden cardiac death and the development of structural cardiac disease. We hypothesize that VPCs in IW are a benign finding and do not predispose the affected dogs to sudden cardiac death or structural cardiac disease.

Dogs were recruited based on the presence of VPCs on surface ECGs during cardiac screening clinics at regional and national breed shows. Each recruited dog underwent a cardiac-focused physical exam, echocardiogram, Holter monitor, and laboratory testing. When anti-arrhythmic therapy was recommended based on Holter monitor results, a follow-up Holter monitor was performed when possible. When greater than one year had lapsed in dogs not on anti-arrhythmic medication, a second Holter was performed when possible. IW were excluded if they had known primary myocardial disease or atrial fibrillation.

Nineteen IW were included in the study with a mean age of 4.6 years (range 1-10 years) and no clinical signs of cardiac disease at enrollment. One dog had equivocal findings of occult dilated cardiomyopathy (DCM) and 2 dogs had evidence of a mild variant of tricuspid valve dysplasia. Subsequent echocardiograms were performed in 2 dogs between 2 and 4 years after the initial echocardiogram and did not show progression toward occult DCM. Initial Holter monitors revealed mean total VPCs of 725.52 (min. 0; max. 4420) primarily occurring in singlets with 10 dogs having couplets (mean 23.3; min. 1; max. 166), 4 dogs having triplets (mean 4.5; min. 1; max. 14), and 3 dog having episodes of ventricular tachycardia (mean 21.3; min. 5; max. 53; mean number of beats 20.3; mean fastest rate 216.67 bpm). The mean overall heart rate was 70 bpm (mean HR min. 64; mean HR max. 95). Atrial ectopy was noted in 7 dogs with 2 dogs having paroxysmal supraventricular tachycardia. Five dogs were on or were started on anti-arrhythmic medications. Post-antiarrhythmic therapy Holters were available for 2 dogs and showed reasonable ventricular arrhythmia control. Thirteen dogs were deceased by the end of the study period. Ten were non-cardiac deaths (osteosarcoma n = 5, 2/5 on anti-arrhythmics; progressive neurological weakness n = 2; hemangiosarcoma n = 1; anesthesia/surgical complications n = 1, on anti-arrhythmics; acute renal failure n = 1). Two dogs had an unexplained sudden death event. One dog was euthanized for progressive weakness and had a history of equivocal findings of occult DCM 2 years previous and was on anti-arrhythmic therapy.

Ventricular arrhythmias appear less malignant in IW compared to other breeds. However, they may be associated with a low risk of sudden death and monitoring is recommended. This study was not able to determine if ventricular arrhythmias in IW are a precursor for the development of DCM due to the lack of follow-up echocardiograms. Multiple mobile ECG heart monitors have become available for use in human medicine, however these products have not been commonplace in the veterinary field. The AliveCor device is a simple, accessible, and affordable mobile ECG heart monitor for veterinarians. The purpose of this study was to evaluate the accuracy of the AliveCor device in rate and rhythm diagnosis.

10 cats were initially evaluated exclusively to establish the most accurate positioning of the device. Placement of the device on the left hemithorax, parallel to the sternum, at the level of the heart, with the cat in right lateral recumbency produced the cleanest readings most consistently. This positioning was used for the remainder of the study. A total of 25 cats, 6 normal cats and 19 cats with underlying cardiac disease, were subsequently evaluated using the AliveCor device and a standard ECG.

In 2 cases, the AliveCor readings contained too much artifact for correct interpretation, and they were excluded from further analysis. In 16 out of the 23 remaining cases, the average heart rates obtained from the AliveCor reading and the standard ECG were exactly the same. In those that did not match exactly, minimal difference was seen. An accurate rhythm diagnosis was made in 22 out of 23 cases using the AliveCor reading. Abnormalities such as premature complexes and bundle branch block were detected in 9 out of 13 cases. Chest lead recordings were obtained with the standard ECG in 11 of the cases. In 6 of the 11 chest lead recordings, the AliveCor reading closely resembled the V4 lead.

Limitations of the AliveCor tracings included decreased amplitude of the waveforms on some cats and an increase in respiratory and purring artifact. The addition of a water-based ultrasound gel may improve contact, leading to a cleaner ECG tracing. Also, the AliveCor reading was sometimes inverted, which was easily corrected with the "Invert ECG" function in the AliveCor application.

We conclude that the AliveCor device allows an accurate rate and rhythm diagnosis in cats. Further comparison of AliveCor tracings with traditional ECGs is warranted to determine the accuracy of waveform measurements. Hypertrophic cardiomyopathy (HCM) is the most common heart disease in cats and has been described as an inherited disease in the Sphynx. Causative mutations have been described for the Maine Coon and Ragdoll breeds in the cardiac myosin binding protein C gene (MYBPC3). Causative mutations are not yet described in the Sphynx breed.

We hypothesized that a causative mutation for HCM in the Sphynx cat would be identified in the exonic and splice site regions of 1 of 5 candidate genes selected based on published genetic mutations in human beings and SDS-PAGE analysis of affected Sphynx myocardium.

Five unrelated affected Sphynx cats and 2 unaffected Domestic Short-Hair cats (controls) were studied. Left ventricular myocardial protein bands were compared between an affected Sphynx and normal cat by SDS-PAGE analysis. Protein identification was determined by mass spectrophotometry for significantly different band patterns. The following candidate genes were chosen based upon these results, association with left ventricular hypertrophy and previously published human HCM studies: myomesin (MYOM1), MYBPC3, NADH dehydrogenase ubiquinone flavoprotein 2 (NDUFV2), aspartate beta-hydroxylase (ASPH) and phosphofructokinase of muscle (PFKM). Exonic and splice site regions of each candidate gene were sequenced in the 5 affected and 2 control cats. Sequences were compared for nucleotide changes between affected cats and the published cat and human sequences. Base pair changes were considered to be causative for HCM if they were present in the affected cats but not in the control cats or published sequences and if they involved a conserved amino acid and changed that amino acid to a different polarity, acid-base status, or structure.

A causative mutation for HCM was not identified, however several single nucleotide polymorphisms were detected in both affected and normal cats within each candidate gene. Additional studies are warranted to identify molecular causes of HCM in the Sphynx. The purpose of this case study is to bronchoscopically evaluate large breed dogs with significant cardiomegaly and the impact seen on the airways. There has been an association between bronchial collapse and chronic valvular heart disease seen in smaller breed dogs. A similar association has not been investigated in large breed dogs with cardiomegaly.

This was a case-controlled, observational study. Bronchoscopic evaluation of client owned large breed dogs ( ! 25 kg) was performed by a single operator. All dogs had significant cardiomegaly diagnosed by thoracic radiographs and echocardiography at Atlantic Coast Veterinary Specialists.

The results are included in the table below. Only one of the dogs studied showed significant airway collapse of the left cranial lobar bronchus (LB1), left caudal lobar bronchus (LB2) and right cranial lobar bronchus (RB1). Two other dogs showed mild airway collapse. None of the dogs studied showed collapse of the left principal bronchus (LPB). The smaller airways were normal in all dogs. This is in contrast to a previous study in small breed dogs showing moderate to severe airway collapse associated with cardiomegaly. The causes of cardiomegaly in this study were tricuspid valve dysplasia (3) and dilated cardiomyopathy (2). In conclusion, significant bronchial collapse does not appear to be a common finding in large breed dogs with cardiomegaly. This is in contrast to what has been previously shown in small breed dogs. However, a majority (60%) of dogs studied did have at least a mild degree of airway collapse. Additional studies are indicated to ascertain the different pathophysiologic processes involved in airway collapse in large versus small breed dogs with cardiomegaly. Heart rate evaluation is an important part of every exam and is routinely used in clinical decision making. In the hospital setting, heart rate is routinely assessed by auscultation. Ideally, inhospital heart rate would reflect the patient's heart rate at home. However, in patients with normal heart rate variability, the heart rate in the clinical setting is often higher than in the home environment. 24-hour ambulatory ECG (Holter monitor) is routinely used for an accurate evaluation of heart rate and rhythm in the home setting but can be cost prohibitive is some cases. The objective of this study is to evaluate several different methods of heart rate assessment in the hospital and at home to determine which is most strongly correlated with Holter monitor assessed heart rate. We hypothesized that heart rate assessed in the home by the iPhone wireless ECG would be more strongly correlated with Holter monitor assessed heart rate than in-hospital measures.

Normal dogs were recruited for enrollment in the study. Dogs with known arrhythmias, receiving heart rate altering medications or with known significant disease were excluded. All enrolled dogs had their heart rate assessed in the hospital by auscultation, standard diagnostic ECG and iPhone wireless ECG (AliveCor Inc., San Francisco, CA). All dogs also had a 24 hr ambulatory ECG (Forrest Medical, LLC, East Syracuse, NY) and three iPhone wireless ECGs performed at home. At home ausculted heart rate was not collected because the auscultation skills of participants (DVMs or DVM students) were not representative of the general population. The three at home iPhone wireless ECGs were obtained when the dog was sleeping, quiet and active. Greater than 20 hours of holter data was required for inclusion in the study.

Seventeen dogs were enrolled in the study. The median age was 4.5 years (range 0.5-12.5 years) and median weight was 28.4 kg (range 9.9-56.8 kg). A variety of breeds were enrolled. All dogs had either a sinus rhythm or sinus arrhythmia on their screening ECGs.

None of the in-hospital heart rate assessment methods (ausculted heart rate, standard ECG and iPhone wireless ECG) correlated significantly with the minimum, average or maximum Holter monitor assessed heart rate. The at home average iPhone wireless ECG heart rate did correlate with the average Holter monitor heart rate (Pearson r-0.7853, r 2 -0.6167, p-0.0002).

Unfortunately, in normal dogs, in-hospital heart rate measurements do not correlate with heart rate at home. The iPhone wireless ECG used at home does correlate with Holter monitor assessed average rates and provides an easy to obtain, less expen-sive option to evaluate heart rate at home. Further studies are needed to determine if iPhone wireless ECG measured rates are superior to owner ausculted heart rates and whether the correlations observed in this study persist in patients with clinical heart disease, arrhythmias and altered heart rate variability. QT interval represents the time of ventricular activation and repolarization and is inversely proportional to heart rate (HR). When not associated with a decrease in HR, the prolongation of this interval can facilitate reentrant arrhythmias because of heterogeneous ventricular repolarization. In humans, prolonged corrected QT (QTc) is predicitive of mortality in heart failure (HF) patients. This study aimed to compare QTc, QT interval dispersion and QT standard deviation (SD) in dogs with HF due to myxomatous mitral valve disease NYHA III and IV or ISACHC II and III (n = 30) and in a control group (n = 29), consisting of dogs of approximate size, without cardiac or other systemic disease. QT intervals were measured manually as the mean of 3 consecutive complexes, on 10-lead ECGs, using a caliper. QT dispersion was calculated as QTmax-QTmin among 10 leads. Bazett and Fridericia formulas were used for QTc. RR intervals used were the mean of 2 intervals as used for QT measurement in lead II. The groups were compared using Student t or Mann-Whitney test. Partial anomalous pulmonary venous connection (PAPVC) is a congenital disease in which one or more (but not all) pulmonary veins are connected to the right atrium or cranial venae cavae. Antemortem diagnosis of this condition in dogs has not been previously reported in veterinary medicine. The purpose of this report is to describe the case presentation and diagnosis of PACVC in dogs.

All 4 cases were miniature Schnauzers presented to Azabu University for evaluation of right heart enlargement. They were all asymptomatic, and right heart enlargement was coincidentally noticed on screening thoracic radiographs. An abnormal blood vessel-like structure connecting to the right atrium was observed on echocardiogram in all cases. Mild valvular pulmonary stenosis was concurrently noticed in one case. No other conditions resulting in right-heart dilation such as pulmonary hypertension, atrial septal defect or tricuspid valve regurgitation were found. Computed tomography angiography (CTA) and cardiac catheterization were performed in 2 cases. A step-up of oxygen saturation within the right atrium was observed in an oxymetry run. CTA revealed that the pulmonary vein of the right anterior lung lobe was connected to the right atrium in both 2 cases, which made for a definitive diagnosis as a PAP-VC. Although no familial relationship among these cases is known, the heredity of this disease in miniature Schnauzer was suspected. Few studies have evaluated cardiac troponin I (cTnI) in dogs using high-sensitivity assays. The purpose of this study was to evaluate cTnI concentrations using a high-sensitivity assay in 4 well-defined groups of dogs.

A high-sensitivity immunoassay, ADVIA Centaur CP â Ultra-TnI (Siemens Medical Solutions Diagnostics, NY), was used to measure cTnI concentrations in dogs that underwent comprehensive clinical evaluation, including echocardiography. Serum samples were obtained and stored at -80C until batch analysis. Groups included (1) 24 healthy dogs without evidence of systemic or cardiac disease, (2) 19 dogs with degenerative mitral valve disease (DMVD), (3) 19 dogs with congenital heart disease, and (4) 22 dogs clinically affected by arrhythmias (15 with ventricular arrhythmias). Serum cTnI concentrations were compared between the 4 groups using a Kruskal-Wallis test with a p-value <0.05 considered statistically significant. Data are reported as median (range).

Median age for Group 1 was 2.63 years (0.83-8.17), weight was 27.1 kg (5.1-42.5), cTnI was 0.017 ng/mL (0.005-0.128). Median age for Group 2 was 9.58 years (4.0-11.92), weight was 8.7 kg (4.8-22.6), cTnI was 0.085 ng/mL (0.005-0.414). Median age for Group 3 was 5.33 years (0.25-8.5), weight was 19.0 kg (1.6-54.8), cTnI was 0.088 ng/mL (0.035-1.405). Median age for Group 4 was 8.67 years (0.92-12.25), weight was 33.2 kg (12.3-54.2), cTnI was 0.247 ng/mL (0.02-25.211). Serum cTnI concentrations from dogs with cardiac disease were significantly higher than Group 1. Further studies are in progress to characterize the clinical utility of high-sensitivity cTnI immunoassays in dogs with suspected or known heart disease. Pimobendan is a phosphodiesterase III inhibitor that has been shown to have positive survival benefits in dogs with congestive heart failure (CHF) due to myxomatous mitral valve degeneration (MMVD). Clinical trials of pimobendan in human patients have demonstrated a trend towards sudden death, which is presumed to be due to ventricular arrhythmia. The question of whether pimobendan therapy alters the incidence of cardiac arrhythmia in dogs with CHF has not been completely addressed. The primary aim of this study is to determine whether pimobendan therapy has any effect on the incidence of arrhythmias in small breed dogs with CHF due to MMVD. The secondary aim of this study is to evaluate the effect of pimobendan therapy on quality of life scores and sleeping respiratory rates. Six small breed dogs (<15KG) with MMVD were recruited in a prospective double blinded cross over designed study. Each dog was treated with either pimobendan or placebo for two weeks followed by a one-week washout period, and then treated with the other medication for two weeks (placebo or pimobendan respectively). All dogs received standard background therapy with an angiotensin converting enzyme inhibitor (enalapril or benazepril) and furosemide as required throughout the study. Dogs requiring treatment with antiarrhythmic drugs were excluded from the study. For each dog, a 24-hour ambulatory Holter monitor was performed at baseline, after two weeks on pimobendan and after two weeks on placebo. Average heart rate, frequency of arrhythmia (ventricular and supraventricular) including single premature beats, couplets, and runs of tachycardia were recorded. Owners were instructed to complete a quality of life questionnaire at each time point, as well as record sleeping respiratory rates throughout the course of treatment. Frequency and type of arrhythmia were compared between baseline, placebo and pimobendan. No significant difference in frequency or type of arrhythmia was noted between baseline, pimobendan and placebo and there was no significant difference between quality of life or sleeping respiratory rate questionnaires between placebo and pimobendan. In conclusion, pimobendan does not appear to increase the incidence of cardiac arrhythmia in small breed dogs with CHF secondary to MMVD when compared to placebo. LASSBio897 is a novel derivative of the compound LASS-Bio294, which exhibited an even more potent vasodilatory activity. Recently, a pharmacokinetic study in dogs identified two metabolites from LASSBio897. Thus, the present pilot study carried out in vivo tests in order to evaluate the potential vasodilatory effect of those metabolites on the cardiovascular system of dogs.

We used one adult Beagle, female and healthy. Capsules containing the prototype were given (1 mg/kg of LASSBio-897) and serum samples were collected after fifteen (T15 m), thirty (T30 m), sixty (T60 m), ninety (T90 m) minutes and two (T2), three (T3), four (T4), six (T6), eight (T8), twelve (T12) and twenty four (T24) hours. Oscillometric blood pressure screening was performed before oral administration and after two (T2), four (T4), six (T6), twelve (T12) and twenty four (T24) hours. Another Beagle dog received 0.5 mg/kg of Benazepril as a control and its parameters were evaluated following the same periods of time described above.

Based on pharmacokinetic study, the behavior of both compounds was analyzed. LASSBio 897 decreased rapidly and progressively until disappearance in thirty minutes (T30 m). Its first metabolite's peak occurred in T30 m and the second peak in T90 m, disappearing three hours after oral administration. The second metabolite appeared three hours after oral administration (T3), showed higher serum concentration in one hour (T4) and decreased until disappearance in T6. Concomitantly, blood pressure went down twice in different times (T2 and T6), both after serum peaks of the metabolites. After T6, blood pressure went back up. Benazepril decreased progressively until its disappearance in T6, and its metabolite (Benazeprilat) increased until achieving its higher concentration in T2 and initiated progressive decrease. Concomitantly, blood pressure went down gradually after two hours (T2) of oral administration, until the last measurement taken at T24. In evaluation of the average obtained from blood pressure values, no statistical difference was observed (p = 0.3) when comparing Benazepril and LASS-Bio897. The prototype proved to be safe, since no hypotension occured.

Considering the results obtained and the conditions under which this preliminary experiment was conducted, it can be concluded that the oral administration of LASSBio-897, in doses of 1 mg/kg, seemed promising due to similar vasodilatory effects when compared to Benazepril. Thus, further studies will be analyzing higher doses of this new compound, in a group of dogs, and their effects on the cardiovascular system. Although myxomatous mitral valve disease (MMVD) is the most common acquired heart disease of dogs, the accurate diagnosis of congestive heart failure (CHF) can be difficult because clinical signs are nonspecific. Generally evaluated echocardiographic indices including ejection fraction, fractional shortening, LA/Ao ratio, and LA/Ao surface area have the limitation because of their load-dependence. By contrast, isovolumic phase index of LV function, such as dP/dt max, is less load-dependent and are theoretically a more accurate reflection of LV function. Therefore, the purpose of this study was to evaluate the ability of maximum dP/dt index in diagnosis of CHF in MMVD of dogs.

Fifty-one dogs with MMVD were classified by ISACHC scale (Ia=5 dogs, Ib=7 dogs, II=16 dogs, IIIa=10 dogs, IIIb=13 dogs). The maximal MR jet velocity (dP/dt=32/time between 1 and 3 m/ sec) was determined using CW Doppler imaging with a sweep speed of 100 mm/sec. LA/Ao ratio and FS as well as dP/dt max were also calculated with 51 dogs ( Table 1 ). The patient dogs also were classified into non-CHF group (Ia and Ib) and CHF group (II, IIIa, and IIIb). Interestingly, two dogs of IIIb class were very severe CHF condition and died in 12 hours since first presentation. However, dP/dt maxs of two dogs were markedly increased compared to other dogs of IIIb class (4050 mmHg and 4741 mmHg each). The authors think that dP/dt index can be valuable indicator for survival of CHF. LASSBio294 is a new cardioactive prototype produced from safrole substrate, found in Brazilian plants like "canela branca" (Ocotea pretiosa), which showed positive inotropic and vasodilatory effects in in vitro and in vivo tests. LASSBio897 is a novel derivative of this lead compound, which exhibited an even more potent vasodilatory activity. In a previous study two metabolites were identified (Hydroxylated LASSBio294 and LASSBio294 Sulfoxide), where LASSBio294 Sulfoxide was the major metabolite found in serum of dogs. Recently, two metabolites were identified from the analogue LASSBio897. Thus, the present pilot study carried out in vivo tests in order to ascertain potential positive inotropic effects of those metabolites on the cardiovascular system of dogs.

We used five adult Beagles, female and healthy. Capsules containing LASSBio294 were given, in a dose of 2 mg/kg, and serum samples were collected after fifteen (T15 m), thirty (T30 m), sixty (T60 m), ninety (T90 m) minutes and two (T2), three (T3), four (T4), six (T6), eight (T8), twelve (T12) and twenty four (T24) hours. Echocardiographic examinations were performed before oral administration and after two (T2), four (T4), six (T6), twelve (T12) and twenty four (T24) hours. Another Beagle dog received 1 mg/kg of LASSBio897 and its parameters were evaluated following the same periods of time described above.

LASSBio294 higher serum concentration was observed in T4 and its disappearance in T8. The serum peak of its metabolite occurred in the first hour, and it dropped continuously until disappearance in T24. An increase in EF% and SF% of 13.1% and 19.1%, respectively, was noticed at T24. LASSBio897 decreased rapidly and progressively until disappearance in thirty minutes. Its first metabolite's peak occurred in T30 m and the second peak in T90 m, disappearing three hours after oral administration. The second metabolite appeared at this time (T3), showed higher serum concentration in one hour (T4) and decreased until disappearance in T6. Concomitantly, echocardiographic examinations demonstrated no difference between EF% and SF% levels, for different times, obtained during a twenty four hours period.

Considering the results obtained, it can be concluded that the oral administration of LASSBio294 in doses of 2 mg/kg increases myocardial contractility, mainly due its metabolite's activity. However, the same was not observed with its derivative LASS-Bio897 in doses of 1 mg/kg in this pilot study. Further studies will be analyzing higher doses of this derivative, in a group of dogs, and its effects on the cardiovascular system. The purpose of this study was to compare the level of agreement in identifying left atrial enlargement (LAE) between left atrial to aortic root ratio (LA:Ao) and left atrial volume using the area-length method. Sixty dogs affected with chronic degenerative valve disease (CDVD) and 22 normal dogs were prospectively studied with 2-D echocardiography. Left atrial volume was indexed to body weight (LA Vol/BW). To define overall disease severity, each dog was assigned a mitral regurgitation severity score (MRSS) based on echocardiographic parameters that did not include left atrial size. The upper limit of normal for LA Vol/BW was 1.1 ml/kg, as determined by control group data. LA:Ao was deemed normal if 1.5. Of the 60 affected dogs, there were 20 dogs in ACVIM Stage B1, 25 dogs in Stage B2 and 15 dogs in Stage C. LA Vol/BW identified LAE in 12 cases in which LA:Ao was normal;7 of these were in Stage B1 and 5 were in Stage B2. This diagnostic disagreement was statistically significant (P = 0.00012). Of the 12 cases in which diagnostic discrepancies were identified, 5/5 of the B2 dogs and 3/7 B1 dogs had a moderate MRSS while 4/7 B1 dogs had a mild MRSS. No diagnostic discrepancies between LA:Ao and LA Vol/BW were apparent in dogs with a severe MRSS.

This study shows evidence of diagnostic disagreement between LA:Ao and LA Vol/BW for the assessment of LAE. LA Vol/BW was superior to LA:Ao for identification of mild LAE.

T Aoki, H Sunahara, K Sugimoto, Y Fujii. Azabu University, Kanagawa, Japan.

Mitral valve repair is becoming more common in dogs with mitral regurgitation (MR) due to chronic valvular heart disease (CVHD). However, the technique is challenging and outcome depends on the surgeon's skill and experience because small-sized dogs are more frequently affected by CVHD. In humans, a loop technique is used for chordal reconstruction with a small surgical incision, which allows for minimally invasive surgery. In addition, the Kay-Reed method, in which the annulus is encircled with simple mattress sutures, has been used in children, i.e., small surgical site, for tricuspid annuloplasty. The purpose of this study was to assess these surgical techniques in dogs with MR.

Study 1: A modified loop technique was performed in 3 laboratory beagles with spontaneous CVHD. After the surgery, MR was diminished or significantly decreased in all dogs. Study 2: A modified Kay-Reed method (MKR) was performed in 4 normal laboratory beagles. The peak E-wave significantly increased (p = 0.0004), although the mean pulmonic capillary wedge pressure did not change after surgery. Study 3: One laboratory beagle with cardiomegaly due to CVHD underwent mitral valve repair using both the modified loop technique and MKR. Total bypass perfusion time was 68 min versus about 100 min with the conventional technique in our hospital. MR markedly decreased and heart size was normalized after the surgery.

In conclusion, these techniques could be applicable in dogs. Further studies are needed to evaluate long-term outcomes and whether these techniques are effective in dogs with severe MR. Left atrial enlargement (LAE) has been reported as an important radiographic finding in the diagnosis of left-sided CHF (l-CHF) in cats. We hypothesized that LA size as assessed by thoracic radiography can be normal in cats with l-CHF.

100 consecutive cats with acute l-CHF were evaluated. To be included, thoracic radiographs taken in orthogonal planes and transthoracic echocardiography performed within 24 hours of diagnosis were required. On radiographs, presence of CHF (y/n), cardiomegaly (y/n), LAE (y/n), vertebral heart score (VHS,) LA-VHS, and the cardio-thoracic ratio were assessed. On echocardiography, LAE (y/n), maximum LA dimension (LADmax) and area (LAAmax), and LV size were evaluated. Reproducibility of radiographic and echocardiographic data was done using the coefficient of variation and Cohen's Kappa from 20 randomly selected samples. Data was quantitatively evaluated using standard statistical procedures.

On echocardiography, LAE was diagnosed in 95% of cats based on visual inspection of images; 86% had LADmax>16 mm and 84% had LAAmax>2.80 cm 2 suggestive of LAE. Upon lateral and ventral-dorsal radiographs, only 50% and 54% of cats had evidence of LAE, respectively. 36% had absence of LAE in both views, 39% had presence of LAE in both views, and 25% had presence of LAE in only one of the two views. 97% of cats had cardiomegaly based on subjective assessment of radiographs, whereas VHS was >8v in 92%. Pulmonary arterial enlargement (69%) was found in more cats compared to pulmonary vein enlargement (49%).

These data suggest that LAE on thoracic radiographs may be absent in cats with l-CHF. Acquired valvular heart disease is the major cause of coughing, dyspnea, syncope, pulmonary edema, pulmonary hypertension and sudden death. Increasing the number of the geriatric dogs, it is necessary to understand of the characteristics of disease, precise diagnostic evaluation and appropriate treatment. Because the characteristics of this disease are a chronic progressive, this retrospective study was started because the need for accurate prognostic factor affecting survival rates and survival time depending on patients status.

In this study, small to middle-sized 170 dogs with acquired valvular heart disease of the Veterinary Medical Teaching Hospital (VMTH) of Konkuk University during the study period (March, 2009 to June, 2012 were included, 128 dogs had mitral valve insufficiency, 3 dogs had tricuspid valve insufficiency and 39 dogs had bivalvular insufficiency. All the dogs with acquired valvular disease were classified into survival (S) group and nonsurvival (NS) group which divided into more-than-one-year-survival NS group and less-than-one-year-survival NS group. To evaluate treatment effects, case records of 64 dogs that had undergone re-evaluation were selected from previous 170 dogs group and classified into survival group and non-survival group. The items of the study include the prevalence and characteristics of physical examinations, hematologic, radiographic, ECG, echocardiographic evaluation, treatment variables, complicated disease and reason of death.

Prevalence characteristic of this study, castrated male (41.8%), Maltese (35.5%), 11 to 15 year-old dog (51.5%) were dominant. High grade murmur and ISACHC class, dyspnea, activity were affected to survival time, also type of valvular insufficiency was prognostic factor in this study.

On blood work showed complete blood cell count and electrolyte were not affected to survival time, but some serum biochemistry profiles including blood urea nitrogen (BUN), Aspartate Aminotransferase (AST), albumin (ALB) and phosphorus (P) evaluated as a prognostic factor. Presence of pulmonary edema and severity of cardiac remodeling were affected to survival time on radiographic examination. On echocardiographic evaluation, type of affected valve, severity of mitral regurgitation and fractional shortening were prognostic factor.

In evaluation of treatment effect groups, coughing, dyspnea, activity, syncope, WBC count were improved in S group, but in NS group only improvement of coughing and increasing pulmonary pressure gradient were evaluated.

In conclusion, to predict survival time and prognosis of the dogs with acquired valvular heart disease, it is necessary to perform accurate diagnostic inspection and evaluation for multiple prognostic factor rather than just one test result. Also, evaluation for improvement of clinical symptoms those mentioned above after treatment and owner education for steady medication can be useful to improve survival times and predict prognosis. Visceral leishmaniasis (VL) is a multisystemic disease, and is not well known the pathophysiological mechanisms leading to hypertension frames. We evaluated the effects of systolic blood pressure (SBP) on the cardiac structure in dogs with VL. 37 dogs of various breeds (7 to 14 kg) with VL were stratified by SBP. 17 dogs with SBP to 129 mmHg shaped the G1, 10 dogs with SBP between 130 and 160 mmHg shaped the G2, and 10 dogs with SBP above 160 mmHg shaped the G3. Chest radiographic, ECG, echocardiographic, serum creatinine (SCr), urinary creatinine protein ratio (Up/c), electrolyte profile and renal excretion of calcium, phosphorus, sodium, phosphorus and magnesium, determination of serum aldosterone, troponin I, lactate dehydrogenase 1 (LDH1), creatine kinase MB fraction (CK-MB), and plasma renin, were measured in all dogs. Data were analyzed by Kruskal-Wallis Test followed by Dunnett's test for post-hoc comparisons; significance was set at P < 0.05. G3 dogs showed at echocardiographic examination increased of IVSd, PWD, ITEI, IVRT, FS, EF, IVS%, LVM. G3 dogs had higher SCr, Up/c, renal excretion of sodium, serum aldosterone, plasma renin, troponin I, CK-MB. Our results indicate that dogs of G3 with VL from this study considered hypertensive, develop changes in cardiac function and structure due to renal impairment. Additionally, the increase in SBP, aldosterone and renin, is suggestive activation of compensatory mechanisms in these dogs. Despite medical and surgical advances, short term survival rates in equine post-operative surgical colic patients have not sig-nificantly improved over the past 20 years. Although several markers for measuring global tissue perfusion are available, there are currently no practical measures of capillary microvascular perfusion in equine patients. In human patients with sepsis, early goal directed therapy aimed at improving microcapillary perfusion rather than global perfusion improves survival. The purpose of the study was to evaluate the technique of orthogonal spectral imaging (OPS) to visualise the capillary microvasculature in conscious normal horses and to assess the reliability of this technique. Images of the capillary microvasculature were obtained in six normal horses from the lip, gum and rectum by one operator on five days and four operators on one day and stored and analysed offline. Each video was assessed for quality and perfusion parameters to include proportion of perfused vessels (PPV), functional capillary density (FCD), microvascular flow index (MFI) and vessel density (VD). Data was normally distributed and analysed using ANOVA and ICC. All four operators were able to obtain all images at each site. Images of the gum were more difficult to obtain than the lip and rectum. Technique repeatability was average to good (ICC=0.4-0.6) for all measures except for FCD and VD measurements from the rectal mucosa (ICC<0.4). Technique reproducibility revealed 93% agreement between operators and was average to good (ICC=0.4-0.6) for all perfusion measurements except for FCD and VD from the rectal mucosa which was poor (ICC<0.2). Measurement repeatability revealed good agreement (ICC=0.6-0.88) across all measures of microvascular perfusion, Measurement reproducibility revealed agreement between analysers of 83% and was average to good for all measures of perfusion (ICC=0.4-0.73) except for MFI from the gum mucosa (ICC=0.2). Image quality was correlated with agreement; the better the image quality, the more likely the measure would be reliable. In conclusion, PPV was the most reliable technique across all sites to assess the mucosal microcirculation in normal conscious horses. Assessment of the applicability of this technique in the horse requires further invivo evaluation. Endotoxin causes myocardial injury and left ventricular dysfunction in rodent, dog and human models of endotoxemia. The specific aim of this study was to describe cardiac rhythm disturbances and echocardiographic changes in heart size and function in horses undergoing experimental endotoxin challenge. Nine adult mares free of cardiac disease received 125 ng/kg/hr Ecoli O55:B5 LPS as an IV infusion over 4 hours immediately followed by a 10L IV bolus of isotonic fluids and 1.1 ng/kg IV flunixin meglumine. Heart rate, cardiac rhythm and direct blood systemic blood pressure were monitored continuously. Two-D and M-mode echocardiographic examinations were performed at 0, 2, 3, 4, 5 and 24 hours following initiation of endotoxin infusion. ECGs were continuously recorded until hour 48. Changes in echocardiographic measurements over time were assessed using mixed effects multilevel regression analysis.

All horses completed the study and developed signs of endotoxemia. Relative bradycardia was seen in association with marked systemic hypertension. One horse developed an accelerated idioventricular rhythm approx 12 hours after discontinuing the endotoxin infusion. Six horses had fractional shortening < 30% and spontaneous contrast within the left ventricle at one or more time points during LPS infusion. Left ventricular internal diameter in diastole and left atrial diameter were significantly decreased at all time points during endotoxin infusion (p 0.001). Left ventricular free wall thickness during systole was significantly decreased at hours 3, 4, and 5 (p 0.001). LPS exposure altered left ventricular size and function and induced cardiac rhythm disturbances. Angiotensin-Converting Enzyme (ACE) inhibitors are used empirically in horses with cardiovascular disorders despite the paucity of data available. The objective of the present study was to compare the pharmacodynamics of 4 different ACE inhibitors.

Eight healthy horses were fasted overnight prior to receiving 4 ACE inhibitors, each administered at 2 dosages, using a randomized Latin square design (Benazepril: 0.5 mg/kg and 0.25 mg/kg; Ramipril: 0.3 and 0.1 mg/kg; Quinapril: 0.25 and 0.125 mg/kg; Perindopril: 0.1 and 0.05 mg/kg). There was a washout period of 1 week between each administration. Serum ACE activity was measured using a kinetic spectrophotometric method. Data were analyzed using ANOVA for repeated measures.

There was a significant effect of drug and dose on maximum ACE inhibition (I max ), ACE inhibition 24 h after administration (I 24 h ), and area under the curve (AUC 0-48 h ). Benazepril at 0.5 mg/kg resulted in significantly higher I max (86.9 AE 7.0%), I 24 h (60.3 AE 7.9%), and AUC 0-48 h (2611 AE 277%•h) compared to other drugs. There was a significant decrease in indirect blood pressure over time after administration of each drug, but differences in blood pressure were not significantly different between drugs. Pharmacodynamic variables measured after administration of benazepril to horses with free access to hay were not significantly different from those obtained after fasting. Administration of benazepril orally once a day for 7 days did not result in a cumulative effect on ACE inhibition.

Of the ACE inhibitors tested, benazepril is the most effective at inhibiting serum ACE in horses. Multiple immunoassays are available for measurement of circulating insulin concentrations in the horse, including radioimmunoassays and enzyme-linked immunosorbent assays. The chemiluminescent assay is a more convenient, technically simple assay that has recently been used to measure equine insulin concentrations. The purpose of this study was to evaluate the performance of a chemiluminescent assay (Siemens Immulite 1000) as compared to a radioimmunoassay (Siemens Coat-a-Count) that has previously been validated in the horse.

Both assays were evaluated in serum samples from 26 horses, with insulin concentrations of <3.5 to 216 lIU/mL (median, 21.6 lIU/mL), as measured by radioimmunoassay. A standard curve was generated using a commercially available equine insulin standard (12 ng/mL, Shibayagi Company, Ishahara, Japan) serially diluted in reconstituted insulin-depleted non-human serum (Siemens). Dilutional parallelism was assessed in pooled serum from a horse with high endogenous serum insulin concentrations. Agreement between the two immunoassays was assessed by Bland Altman analysis and the Lin's concordance coefficient was calculated.

Detection of the equine insulin standard was poor in both assays. On average, the RIA detected 27% (range, 22-29%) and the Immulite detected 22% (range, 10-41%) of the expected standard concentration. Both assays had excellent dilutional parallelism (r = 0.99) of the high endogenous insulin serum. Bland Altman analysis indicated that 95% limits of agreement ranged from -106.9 to 77.4. The concordance coefficient (q c = 0.58) indicated poor agreement between the two assays. Disagreement was primarily due to three discordant samples; when these were removed the coefficient was q c = 0.97 indicating substantial concordance. Additional samples collected from these three horses on multiple subsequent dates showed results from individual animals to be consistently higher when measured by chemiluminescent assay. In conclusion, the two methods for measuring equine insulin are discordant. Reasons for the poor agreement between assays remain to be determined. Glucose and insulin dysregulation are components of Equine Metabolic Syndrome (EMS) that can only be detected with diagnostic tests. Here we compared two recently introduced field tests for assessing blood glucose responses to oral sugars and insulin resistance, respectively. Sixteen adult horses (14 mares, 2 geldings) previously diagnosed with EMS underwent an oral sugar test (OST) and then two insulin tolerance tests (ITTs) one week later, under fasting conditions. For the OST, horses received corn syrup orally at a dosage of 0.15 mL/kg. Two ITTs were then performed, first with 30 mU regular (soluble) insulin per kg and then 48 hours later with a higher (60 mU/kg; n = 15) or lower insulin dosage (10 mU/kg; n = 1), depending upon initial results. Blood glucose concentrations were measured at 60, 90, 120, 150, 180, and 210 min for the OST and 15, 30, 60, 90, 120, 150 min for the ITT using a hand-held glucometer.

Area under the glucose curve (AUCg) for the OST and the slope of maximal reduction in blood glucose concentration during the ITT were significantly correlated (Pearson r = -0.74; p = 0.001). A ratio (ITT glucose slope/AUCg) was calculated and compared to individual time points in the OST. The 90 min time point (Pearson r = -0.62; p = 0.010) was the most closely correlated with ITT results in this group of EMS horses. Both the OST and ITT are easy to perform and can be used to characterize EMS. Blood insulin responses to the OST should also be assessed. There may be breed-related differences in the prevalence of insulin resistance (IR), obesity and laminitis. It is unclear whether obesity is a cause or a consequence of IR in equids. The aim of this study was to characterize insulin responses to a glucose containing meal in different breeds of horses and ponies in moderate body condition. These insulin responses were compared with insulin sensitivity (SI), as determined by an insulin-modified frequently sampled intravenous glucose tolerance test (FSIGT).

8 Standardbred horses, 8 mixed-breed ponies and 6 Andalusian horses in body condition score (median) 5.0 out of 9 (range 4.3-6.0) were used in the study. Median age was 8.5 years (range 5-19 years) and there was no significant difference between the groups in body condition score or age. Animals were each fed a meal containing 1.5 g/kg glucose. Serial blood samples were collected over 6 hours and analyzed for plasma insulin and glucose concentrations. The peak insulin value and insulin area under the curve (AUC) were compared between breed groups by a Kruskal-Wallis test. Each animal also underwent a FSIGT with minimal model analysis to determine SI. A Pearson correlation was performed to compare peak insulin concentration following the glucose meal (log transformed) with SI.

There were no significant differences in blood glucose curves or time taken to consume the meal between groups. However, the peak insulin concentration following the glucose meal was significantly lower in Standardbreds (mean AE SEM; 21.2 AE 3.5 lIU/mL) than ponies (94.1 AE 29.1; P = 0.003) and Andalusians (85.3 AE 18.6; P = 0.004). The insulin AUC was also significantly lower in Standardbreds (3.1 AE 0.6 IUÁminÁL À1 ) than ponies (13.5 AE 3.6; P = 0.009) and Andalusians (15.0 AE 2.7; P = 0.004). Insulin sensitivity, as determined by the FSIGT was significantly lower in Andalusians (0.99 AE 0.18 x10 -4 /[mUÁmin]) than Standardbreds (5.43 AE 0.94; P < 0.001); and in ponies (2.12 AE 0.44) compared with Standardbreds (P = 0.003). Peak insulin concen-tration following the glucose meal was negatively correlated with SI (P < 0.001; r = -0.73).

These results indicate that there are significant and marked breed-related differences in insulin responses to oral glucose in horses and ponies, even in moderate body condition. The lower SI of ponies and Andalusian horses may be associated with the increased susceptibility of these breeds to the development of obesity and laminitis. The equine metabolic syndrome (EMS) is characterized by obesity, insulin resistance and a predisposition to laminitis in affected horses and ponies. Dyslipidemia may be an important component of EMS and warrants further investigation. The purpose of this study was to apply a minimal model of non-esterified fatty acid (NEFA) kinetics to horses undergoing an insulinmodified frequently sampled intravenous glucose tolerance test (FSIGT). This NEFA model (the Boston model) has been previously described in dairy cows and is based on the inhibitory effect of plasma glucose on NEFA production. The aim was to demonstrate that the changes in NEFA concentrations fitted well to the model and could be used to determine the initial rates of NEFA production and utilization.

Six Standardbred horses (median age 9.5 years old; range 5-19) in moderate body condition score (mean AE SD; 4.8 AE 0.5 out of 9), underwent a standard FSIGT. Serial plasma samples were analyzed for glucose (hexokinase method) and NEFA concentrations (Free Fatty Acid Quantification Kit, Abcam, Cambridge, UK). The glucose and NEFA data was fitted with the Boston model using WinSAAM computer software (version 4.0.7).

The data fitted well to the model and the mean (AE SEM) fractional standard deviation for initial rate of NEFA production was 2.2 AE 0.3%. Mean (AE SEM) initial plasma NEFA concentration was 216.3 AE 53.2 lmol/L. Initial rate of NEFA production was 129.6 AE 40.2 lmol/L/min and rate of utilization was 3.0 AE 0.6%/min.

These data indicate that the Boston model describes very well the NEFA kinetics of horses undergoing an FSIGT. Future studies will aim to further characterize the role of dyslipidemia in EMS by comparing the NEFA kinetics of horses and ponies of different insulin sensitivities and body condition scores. Hypertriglyceridemia is a common consequence of negative energy balance associated with significant mortality in equids, especially ponies, miniature horses, and donkeys. This retrospective study included inpatients admitted to New Bolton Center (University of Pennsylvania) from 1997-2011 that were >1 year of age and had a plasma triglyceride concentration of >250 mg/dL at any point. Data tabulated included signalment, presenting complaint, plasma biochemical values, treatment, clinical parameters, and survival outcome. Data were analyzed using the Wilcoxon rank sum and Chi squared or Fisher's Exact test, and by logistic regression to investigate associations with survival to discharge.

Twenty cases were identified, including 11 ponies/miniature horses, 5 donkeys, and 4 horses. Seven of the cases (35%) died or were euthanized (median peak triglyceride 499 mg/dL; range 304-1490), and 13 (65%) survived to discharge (median triglyceride 478 mg/dL; range 291-4082). Fifteen cases (75%) had elevated serum gamma-glutamyl transferase activity (median 149 U/L; range 22-783). All cases received intravenous fluids with dextrose supplementation. Seven cases (35%) were treated with insulin CRI or intermittent dosing, 1 was treated with heparin, and 7 were treated with both. Eight cases (25%) received forced enteral nutrition, 4 received total/partial parenteral nutrition, and 3 received both. In the multivariable analysis only age OR 1.32 (95% CI 0.98-1.77, p = 0.068) and gender OR 56.32 (95% CI 1.15-2748.29, p = 0.042) were retained in the final model; the odds of not surviving were associated with increasing age and being male.

In conclusion, unlike previous studies, no significant relationships were found between triglyceride concentration and survival. While bacterial sepsis remains a leading cause of morbidity and mortality in foals, reliable diagnostic and prognostic markers are lacking. Adrenomedullin (AM) is a polypeptide with diverse biological effects on the cardiovascular system. Certain conditions, including sepsis, are associated with robust increases in plasma AM concentration (p[AM]) in humans and laboratory animals. We hypothesized that p[AM] would be increased in septic and sick non-septic neonatal foals compared to healthy controls, and that p [AM] would be predictive of survival and the presence of sepsis.

Critically ill foals less than one week of age were categorized as septic or sick non-septic based on admission blood culture results and sepsis score. Jugular venous blood was collected from septic and sick non-septic foals at admission and from healthy control foals at 24 hours of age. Plasma [AM] was measured using a commercially available equine ELISA. Additional data obtained from medical records included history, clinical and clinicopathologic variables, treatments, complications and outcome. Data were analyzed using the Mann-Whitney U test and p < 0.05 was considered significant.

Sixty-six healthy, 41 septic, and 49 sick non-septic foals were enrolled in the study. Septic and sick non-septic foals had significantly increased p[AM] compared to healthy control foals (p < 0.0001). In septic and sick non-septic foals, elevated p[AM] was predictive of non-survival (p = 0.028) but not predictive of sepsis (p = 0.71).

Plasma [AM] increases non-specifically during perinatal illnesses and appears to be a potential predictor of survival in critically ill neonatal foals. The equine intestinal microbiota is a rich and complex environment that may be disrupted by factors like changes in diet and/or administration of antimicrobials. This study aimed to characterize changes in the fecal bacterial populations of healthy horses associated with the administration of frequently used antimicrobials.

Fourteen adult mares were assigned to receive procaine penicillin: 20,000 UI/kg IM, q12 h (n = 5), ceftiofur sodium: 2.2 mg/kg IM, q12 h (n = 5) or trimethoprim sulfamethoxazole 30 mg/kg PO q12 h (n = 4) for 5 days. Fecal samples were collected before drug administration, on the last day of treatment and 25 days after. High throughput sequencing of the 16S rRNA gene PCR products was performed. Sequences were classified into operational taxonomic units (OTUs) and bacterial structures compared by the Classical Jaccard measure of dissimilarity and the inverse Simpson's diversity index. A total of 156,790 sequences were obtained.

Overall, Firmicutes and Bacteroidetes were the main phyla found before (64.7% and 30.2%, respectively) and after 5 days of antimicrobials (63.6% and 29.4%, respectively). At the class level Negativicutes and Citophagia significantly decreased (P = 0.011 and P = 0.008, respectively). The average number of OTUs per horse was 832, 681 and 795 before, 5 and 25 days after treatment, respectively. Initial diversity significantly decreased after use of antimicrobials (P = 0.001) and significantly increased 25 days after drug administration (P = 0.005) to a level that was no different than baseline (P = 0.261). There were significant differences in bacterial community structure between baseline and day 5 (P < 0.001) and days 5 and 30 (P = 0.013), but not between baseline and day 30 (P = 0.460). Despite similarities, differences in phylogenetic tree structure were evident before and 25 days after treatment, as day 30 samples did not cluster with the corresponding horse's baseline sample. Horses treated with ceftiofur and TMS clustered by drug used indicating that changes may be predictable depending on the antimicrobial used; however, penicillin treated horses were more variable.

It is concluded that the bacterial species are highly diverse in feces of healthy horses and the use of parenteral antimicrobials leads to a decrease in diversity and changes in composition, even at higher taxonomic levels. Changes on bacterial communities caused by antimicrobial administration were specific for each drug and may be predictable. TMS caused the most marked changes on fecal microbiome. Twenty-five days after treatment bacterial communities are more similar to pre-treatment levels, indicating a recovery from changes caused by antimicrobials, but differences are still apparent. This study could form a basis for a better understanding of the pathogenesis of antimicrobial associated colitis in horses. A recent study has demonstrated that the duration of acid suppression with omeprazole in the horse may be as short as 12 hours and suggested that administration prior to exercise may result in better healing than administration at other times. The objective of this study was to determine whether administration prior to exercise improved healing compared to administration post exercise. Thoroughbred racehorses in training with grade ! 2/4 squamous gastric ulceration were identified on gastroscopic examination. The glandular mucosa was also scored. Horses were randomly assigned to one of two groups. Both groups received 2.0 g (equivalent to 4 mg/kg for a 500 kg horse) of a commercially available omeprazole PO SID with it administered 1-4 hours prior to exercise in the pre-exercise group (PRE) and 1-4 hours post exercise in the post-exercise group (POST). Gastroscopy and EGUS scoring was repeated at 20-28 days. Data was both normally and non-normally distributed and analysed using Student's T test, Mann Whitney U test and Wilcoxon Paired test. Significance was assumed when p < 0.05.

Twenty-five horses met the inclusion criteria and were randomly allocated into two groups (13 and 12 horses in the PRE and POST groups, respectively). There were no differences between the groups at enrolment. In addition to the squamous ulceration, 9 and 10 horses in the PRE and POST groups, respectively, had ! 2/4 glandular ulceration. Mean squamous (PRE 3.0 AE 0.7 vs. POST 3.3 AE 0.7; p = 0.23) and glandular (PRE 1.9 AE 1.3 vs. POST 2.3 AE 1.1; p = 0.38) ulcer scores were not different between the groups at enrolment. Mean ulcer score significantly decreased for the squamous (PRE 3.0 AE 0.7 to 0.9 AE 0.9, p < 0.001: POST 3.3 AE 0.7 to 1.1 AE 0.8, p < 0.001) but not the glandular (PRE 1.9 AE 1.3 to 1.5 AE 1.1, p = 0.14: POST 2.3 AE 1.1 to 1.8 AE 0.6, p = 0.08) mucosa for both groups. There was no difference between the two groups with regards to squamous (p = 0.14) and glandular (p = 0.42) healing or squamous (p = 0.48) and glandular (p = 0.2) improvement. When compared with the glandular mucosa, a higher rate of healing was seen in the squamous mucosa overall (p < 0.001) and in both the PRE (p = 0.0001) and POST (p = 0.038) groups. Worsening of ulcer grade was observed in the glandular mucosa of 13% of horses overall. In conclusion; timing of administration does not appear to affect the efficacy of omeprazole under clinical conditions however the risk of a type II error is relatively high and should be considered. The response of the glandular mucosa to omeprazole therapy is inferior to that of the squamous mucosa. A previous study has demonstrated a dose response in the healing of squamous and glandular EGUS and suggested that the response of glandular EGUS (EGGUS) to omeprazole therapy is inferior to that of squamous EGUS (ESGUS). The purpose of this study was to further investigate these findings. Thoroughbred racehorses in training with grade 2-4/4 ESGUS or EGGUS were identified on gastroscopic examination. Horses were randomised to receive 0.5, 1.0 or 2.0 grams of enteric coated omeprazole paste PO SID, equivalent to 1, 2 and 4 mg/kg, respectively for a 500 kg horse. Omeprazole was administered 1-3 hours prior to morning exercise. Repeat gastroscopy was performed at 28 AE 2 days. Data was both non-normally and normally distributed and analysed using analysis of variance (ANO-VA) and post-hoc Bonferoni test, Kruskal Wallis with post-hoc Dunn's, Wilcoxon Paired Test, Mann Whitney U test, Friedman and Spearman correlation. Significance was assumed at p < 0.05.

Sixty horses met the inclusion criteria and were randomised into three equal groups. Three horses were lost from each of the 2 and 4 mg/kg group for unrelated reasons. Mean ESGUS score decreased in all groups (p < 0.0001). In contrast, no significant change in mean EGGUS score was seen over time for the 1 (p = 0.50), 2 (p = 0.49) or 4 (p = 0.65) mg/kg groups. No difference between the doses was observed over time. Improvement in ESGUS score was observed in 100%, 100%, and 88% of horses in the 1, 2 and 4 mg/kg groups, respectively. In contrast, improvement in EGGUS score was observed in only 31%, 45% and 27% of horses in the 1, 2 and 4 mg/kg groups, respectively. Worsening of EGGUS score was observed in 35%, 50% and 27% of horses in the 1, 2 and 4 mg/kg groups, respectively. Healing of ESGUS was observed in 89%, 94% and 81% of horses in the 1, 2 and 4 mg/kg groups, respectively. Healing of EGGUS was observed in 23%, 9% and 9% of horses in the 1, 2 and 4 mg/kg groups, respectively. No differences between doses were observed for improvement, worsening or healing of the squamous or glandular mucosa. Overall, healing and improvement were more likely to occur in squamous than glandular ulcers (p < 0.0001). In conclusion, the results of the study contradict previous findings and suggest that, under conditions that favour omeprazole efficacy, lower doses as are efficacious as 4 mg/kg. Consistent with previous findings, the results of study clearly demonstrate that the response of EGGUS to omeprazole therapy is inferior to that of ESGUS. Immunoglobulins, cytokines and leukocytes in colostrum play a key role in neonatal immunity and protection against pathogens. The purpose was to characterize leukocytes in equine colostrum and compare those to the maternal peripheral blood mononuclear cells (PBMCs).

Colostrum and heparinized blood was collected directly after birth from 35 mares in 2009, 2010 and 2012. Colostrum was centrifuged and the cell pellet washed while the PBMCs were isolated by density gradient centrifugation. Cells were fixed with 2% formaldehyde and stained with cell surface markers LFA1, CD4, CD8, IgM, CD14, IgE, CD23, CD26, CD16 and NKp46 and flow cytometric analysis performed. An aliquot of colostral cells (2012; n = 13) was isolated by density gradient centrifugation. These colostral cells and paired mare PBMC were cultured with medium or stimulated with PMA. After 48-h, supernatants were harvested and T-cell cytokines (IL-4, IL-10, IL-17 and IFN ) were determined by multiplex analysis. Wilcoxan-signed rank tests were performed.

Leukocytes in equine colostrum are composed of CD8 + (median 47 AE 10%) and CD4 + (median 43 AE 9%). Colostral cells had significantly higher proportions of CD8 + T-cells whereas Bcells and monocytes were significantly reduced (p's<0.05) compared to mare PBMCs. Lower IL-4 and IL-10 production from colostral cells (p's=0.002) compared to mare PBMCs were detected by cell stimulation.

Equine colostral leukocytes are almost exclusively composed of CD8 + and CD4 + T-lymphocytes and upregulation of CD8 + cells are seen in colostrum. Colostral T-lymphocytes differ from maternal cells in their response to stimulation. Further work is necessary to determine the function of these cells in equine neonatal immunity. Besnoitiosis is an emerging infectious disease of donkeys in the United States caused by infection with the protozoan parasite Besnoitia bennetti. Clinical disease is characterized by miliary dermatitis with parasitic cysts in the skin, nares, and sclera. Currently the only method of diagnosis is histopathologic identification of Besnoitia organisms within tissues.

The objective of this study was to evaluate the utility of clinical examination and three serologic assays for the diagnosis of besnoitiosis in donkeys.

A prospective study of 416 donkeys in 6 states was performed. Donkeys were examined for clinical lesions suggestive of besnoitiosis and evaluated for antibodies against B. bennetti using an immunofluorescent antibody (IFA) test and 2 immunoblot assays specific for bradyzoite and tachyzoite antigens, respectively. Donkeys confirmed to be infected with B. bennetti by histopathology (cases; n = 32) were compared to those with no clinical signs of besnoitiosis (controls; n = 384).

Identifying clinical lesions in 2 or more locations correctly identified infected donkeys 80% of the time. Donkeys with besnoitiosis had significantly higher IFA titers (P < .001) and numbers of bradyzoite (P < .001) and tachyzoite (P < .001) immunoblot bands than control donkeys. The sensitivity and specificity of the serologic assays for detecting besnoitiosis was 88% and 100% for IFA, 81% and 96% for bradyzoite immunoblot, and 90% and 100% for tachyzoite immunoblot, respectively. Immunofluorescent antibody and immunoblot assays are effective at identifying donkeys with besnoitiosis and provide a more efficient and less invasive diagnostic alternative to histopathology.

diagnosis and when determining response to treatments. Accurate and repeatable assessment is crucial for any clinical decision making, but is especially important when human safety and horse retirement or euthanasia are considerations. Despite general acceptance of a standard neurologic examination in horses, its reliability has never formally been tested. The aim of this study was to evaluate the inter-rater agreement of the modified Mayhew ataxia grading scale and to determine the degree to which veterinarians agree when deciding an animal is normal or abnormal. The same group of six board-certified medicine and surgery clinicians and two residents graded horses on a scale from 0 to 5 where 0 is non ataxic and 5 is recumbent. A median of 5 raters evaluated 25 horses of mixed breeds and sex. A two-way intraclass correlation coefficient (ICC) derived from ataxia scores in a mixed model with random effect of horse and rater and a kappa statistic was computed. The kappa value for agreeing on normal or neurologic gait was 0.42. The ICC for all ratings was 0.74. The ICC for grade 0 to 2 (n = 12) was 0.00 compared to 0.42 for grades >2. We conclude that the overall inter-rater agreement of the modified Mayhew ataxia grading scale, although good, is poor for the lower grades and moderate on whether the gait is normal or neurologic which emphasizes the need for better grading scales. The purpose of this study was to identify temporal trends in prevalence and antimicrobial susceptibility patterns of bacteria isolated from septic foals between 1979 and 2010.

All foals 30 days of age presented to the VMTH at UC Davis between 1979 and 2010, with a diagnosis of septicaemia confirmed by culture of bacteria from blood or internal organs (ante mortem or at necropsy), and which had a sepsis score of ! 12, were included in the study. Conventional microbiologic methods were used to identify isolated organisms. Susceptibility testing was performed using the microdilution Sensititre â procedure. Jonkheere-Terpstra and Cochran-Armitage trend tests were used for statistical analysis.

A total of 1091 bacteria were isolated from 588 foals. The percentage of gram-positive isolates increased significantly over the years (from 25.9% to 35.9%). The percentage Enterobacteriaceae decreased over time (from 49.1% to 41.1%). Enterococcus spp. isolates were cultured more often in recent years (from 5.5% to 12.1%).

Enterobacteriaceae, Actinobacillus spp. and beta Streptococci showed a decrease in susceptibility to gentamicin and amikacin. Enterobacteriaceae, Enterococcus spp. and Pseudomonas spp. showed decreased susceptibility to ceftiofur. Enterobacteriaceae became more resistant to ceftizoxime and Enterococcus spp. became more resistant to imipenem and ticarcillin/ clavulanic acid. In contrast, Actinobacillus spp. became more susceptible to ampicillin, erythromycin, penicillin and rifampin. Enterobacteriaceae showed increased susceptibility to tetracycline and Staphylococcus spp. became more susceptible to chloramphenicol. Pseudomonas spp. was the only bacterial species that became more susceptible to gentamicin.

The emergence of gram-positive organisms as an increasingly important cause of neonatal sepsis may reflect increased prophylactic or therapeutic use of antimicrobials that have a gramnegative spectrum of activity in foals. Duration of hospitalisation has increased with advances in equine neonatal medicine and therefore an increase in the number of nosocomial infections likely also contributes to the increased prevalence of gram-positive infections.

The results of this study show that the combination of amikacin and ampicillin remains a sound choice for initiating treatment while awaiting culture results, although increasing development of resistance to amikacin was demonstrated. The decrease in activity of ceftiofur against gram-negative enteric organisms is also of concern and suggests that ceftiofur cannot be considered to be a consistently reliable option when an alternative to aminoglycosides is needed. Similarly, the increase in prevalence of Enterococcus spp. and its development of resistance to imipenem are disturbing and make the treatment of neonatal sepsis more challenging. Restrictive and well-considered use of antimicrobials in all situations is therefore crucial. Rhodococcus equi, a facultative intracellular pathogen of foals, is highly susceptible to killing by gentamicin in vitro. However, gentamicin does not appear to be effective in vivo, due to its poor cellular penetration. Encapsulation of drugs in liposomes enhances cellular uptake. The objective of this study was to compare the pharmacokinetics of liposomal and free gentamicin in foals.

Eight healthy foals were given a single IV or nebulized dose (6.6 mg/kg) of liposomal or free gentamicin in a balanced Latin square design. Concentrations of gentamicin were measured in plasma and in bronchoalveolar cells using HPLC-MS. Effect of drug and administration route on each pharmacokinetic parameter was assessed using a two-way ANOVA for repeated measures.

Administration of liposomal gentamicin IV resulted in significantly lower initial plasma concentrations and significantly higher mean (AE SD) half-life (16.3 AE 3.5 vs. 6.2 AE 1.8 h) and volume of distribution (2.00 AE 1.03 vs. 0.72 AE 0.32 L/kg) compared with IV administration of free gentamicin. Plasma concentrations after administration of nebulized liposomal or free gentamicin were 0.57 lg/mL at most time points. Peak gentamicin concentrations in bronchoalveolar lavage cells were significantly higher for liposomal gentamicin compared with free gentamicin after administration by both the IV (5.3 AE 2.7 vs. 3.0 AE 1.7 lg/mL) and the nebulized (4.5 AE 2.7 vs. 1.5 AE 0. 6 lg/mL) routes.

Administration of liposomal gentamicin by the IV route or by nebulization results in significantly higher gentamicin concentrations in bronchoalveolar cells compared with administration of free gentamicin. Exercise-induced pulmonary hemorrhage (EIPH) is associated with impaired short term performance but the long term consequences to the health and well being of horses are unknown. One way of determining the importance of EIPH is to assess the association between EIPH and career performance. Tracheobronchoscopic examinations were performed during 2003 on 744 Thoroughbred horses within 2 hours after completing a race and the severity of exercise-induced pulmonary hemorrhage recorded (Grades 0-4). Race records of horses as of May 1st, 2011 were then reviewed to evaluate career performance. The association between EIPH and career starts, wins and places were analysed via negative binomial regression and career earnings (log 10 (career earnings + $1) were analysed using linear regression.

Horses with EIPH severity grade ! 2 demonstrated significantly lower career earnings than horses with EIPH severity grade 1 (P = 0.001). Horses with EIPH grade 4 versus 0 demonstrated significantly less earnings for the duration of their career (p < 0.001). Number of starts, wins and places for horses with EIPH severity grades ! 1 versus 0 or grade ! 2 versus 1 were not different. When considered individually, horses with severe EIPH (grade 4) demonstrated fewer career starts (p < 0.001) and places (p = 0.02), but not less career wins (p = 0.24). Horses with severe EIPH (grade 4) demonstrate lower career earnings and fewer career starts. This may be attributable to early retirement of horses due to a combination of negative public perception and racing regulations. However, none of the horses demonstrating severe EIPH in this study were permanently disqualified from racing due to repeat epistaxis. This provides evidence of an important biological effect of severe (grade 4) EIPH on lifetime race performance of Thoroughbred racehorses competing without frusemide in Australia. There is little information pertaining to normal echocardiographic measurements in Norwegian Coldblooded Trotters (NCTs), a breed of racehorse that undergoes both speed and resistance training. The objectives were to establish reference values for standard echocardiographic measurements and left atrial velocity in NCTs and determine the effects of age, sex, and race performance on echocardiographic measurements.

Sixty-six NCTs in race training were assessed by two dimensional, M-mode, tissue Doppler and color Doppler echocardiography. Three horses had significant heart disease and were excluded from the reference population. Means, standard deviations and 95% confidence intervals were calculated for echocardiographic measurements. Mean wall thickness (MWT), relative wall thickness (RWT) and left ventricular mass (LVmass) were calculated. Linear regression was used to quantify the relationship between age, sex, number of race starts and echocardiographic measurements and calculations. Number of race starts was highly correlated with age and dropped from the model. Significance was set at p < 0.05.

Valvular regurgitation was present in 65% of horses. Reference ranges were established for standard echocardiographic measurements, left atrial velocity, RWT, MWT and LVmass. The sexes "gelding" and "intact male" were significant predictors of left ventricular internal diameter in diastole (p = 0.001; p < 0.001) and LVmass (p = 0.002; p < 0.001). Age was a significant predictor of left atrial diameter (p = 0.001), relative wall thickness (p = 0.015) and left ventricular mass (p < 0.001). These data provide reference values for NCTs and demonstrate the effects of age and sex. Risk factors for valvular regurgitations (VR) have been suspected in equids, but no extensive epidemiologic study has been performed in a large mixed equine population. Therefore, the aim of this study was to statistically test risk factors for VR in a large population of equids.

Hospital records were reviewed for 3.499 equids, admitted at the internal medicine department of the Liege Equine Teaching Hospital between 1994 and 2011, aged ! 2 years, and which underwent thorough cardiac clinical evaluation. Of this population, 495 cases had ECG and echocardiography performed because of a clinical suspicion of cardiac disease. Chi-square test or logistic regressions (as appropriate) were used to test if breed, gender, age, body weight (BW), and co-existence of various car-diac diseases were risk factors for each VR. Moreover, the risk of development of congestive heart failure (CHF) was tested for each VR. Significance was set at p < 0.05.

Most of the studied animals were warmbloods, and observed prevalences were 4.4% for mitral regurgitation (MR), 2.1% for aortic regurgitation (AR), 1.7% for tricuspid regurgitation (TR), and 1.0% for pulmonary regurgitation (PR). Significant risk factors were male gender and increasing age for AR (OR = 2.03, CI=1.07-4.94), and racehorses breed group and middle-age for TR (OR=4.36; CI=1.10-17.24). No effect of age or BW was demonstrated for MR. MR was the major valvular disease associated with atrial fibrillation (AF), ventricular tachyarrhythmia, PR and CHF. TR was also linked to AF, PR and CHF; but AR was not linked to CHF.

In conclusion, several previously suspected risks factors for VR were confirmed statistically in this study and should be taken into account in health and athletic monitoring of horses presenting predisposing factors. The purpose of the present research was to investigate if Orthogonal ECG lead Y could be used to evaluate air quality in horse stables. Hypothesis being that unsatisfying air quality causes subclinical pulmonary disease with extra load on the right cardiac ventricle on racehorses in training. This resulting in right ventricular enlargements that can be detected with orthogonal ECG lead Y.

Orthogonal ECG for horses was developed and thoroughly investigated for many years by prof. Holmes et al, college of veterinary medicine, Bristol, UK. Holmes et al found that right axis deviation in lead Y was correlated to right ventricular enlargement, indicating pulmonary disease. In the present investigation orthogonal ECG lead Y was recorded on 250 warm blooded trotters in training, stabled in 9 different stables. The percentage of horses showing signs of right ventricular enlargement on ECG lead Y was evaluated for each stable and ranged from 0% to 33%. All stables were examined for efficacy of ventilation and air quality concerning microorganism exposure at level of horse's nostrils. The stables were ranked blindly from the best to the worst, concerning air quality and the percentage of horses showing signs of right ventricular enlargement. A strikingly good agreement was achieved which indicates that orthogonal ECG lead Y could be used to easily and rapidly evaluate if suboptimal air quality has affected racehorses in training. Orthogonal polarization spectral (OPS) imaging provides an opportunity to assess capillary microvascular perfusion in the horse by visualisation of mucosal blood flow. This technique may provide a useful outcome for early goal directed therapy in horses with systemic inflammatory response syndrome (SIRS), which may in turn affect outcomes in horses with severe intestinal pathology. In order to validate the technique in the horse, evidence is required to demonstrate that changes in intestinal perfusion result in changes in mucosal blood flow determined using OPS. The purpose of this study is to demonstrate that changes in perfusion, caused by administration of the alpha 2 adrenoceptor agonist detomidine, results in measurable changes in microvascular blood flow in horses that mirror known aberrations in total peripheral resistance (TPR) and cardiac output(CO). Microvascu-lar blood flow was recorded using OPS placed, manually, per rectum in six normal horses (weighting 603 AE 134 kg) undergoing sedation for a range of clinical procedures. OPS recordings were made prior to and after sedation (5, 10, 20 minutes) with detomidine (10ug/kg) and butorphanol (10 mg/kg) administered by intravenous injection. Microvascular perfusion was determined using standardised methods from OPS recordings including proportion of perfused vessels (PPV), functional capillary density (FCD), microvascular flow index (MFI) and vessel density (VD). Data was normally distributed, presented as mean (AESD), and differences between time points were compared using repeated measures analysis of variance. There was a significant effect of detomidine on microvascular blood flow as demonstrated by changes in MFI, PPV and FCD five minutes after sedation (p < 0.001). Similar changes were effect demonstrated using VD 10 minutes after sedation (p < 0.02; Table 1 ). Microvascular had returned to pre-sedation values by 20 minutes as determined by all criterion. These data demonstrate that changes in organ perfusion that are known to be caused by the alpha-2 adrenoceptor agonists result in observable changes in mucosal blood flow and appear to mirror changes in cardiac output and total peripheral resistance that have previously been demonstrated after administration of detomidine in the horse. Further evaluation is required before the technique can be utilised to advise on clinical management of SIRS. Besides being an important part of anesthetic monitoring, measurement of blood pressure has also of fundamental importance in clinical and experimental studies in horses. Despite the use of devices for non-invasive measurement of blood pressure is routine practice, such devices have never been tested on standing and non-sedated horses according to ACVIM consensus statement. Therefore, the aim of this study was to test three different devices of non-invasive blood pressure measurement in standing and non-sedated horses by the validation criteria according to ACVIM Hypertension Consensus Panel and Veterinary Blood Pressure Society Recommendations (2007) .

For this purpose, were used six clinically healthy Arabian horses, four males and two females, weighing between 360 and 405Kg. Measurements were collected from each horse, during 2 sessions (morning and evening) on two consecutive days, by using a vascular Doppler device (Parks Medical Ultrasonic Doppler 812 â ), an automatic oscillometric device designed for human patients (Dixtal 2710 â ) and an digital oscillometric device designed for blood pressure measurement in small animals (Pet-MAP â ). The cuffs, with width of 40% of the tails circumference, were placed around the base of the tail with the bladder centered over the middle coccygeal artery. Values obtained by noninvasive methods were compared with those obtained simultaneously by invasive blood pressure monitoring (Dixtal 2010 â ), accessed on facial artery after topical application of lidocaine and prilocaine, 25/25 mg/g cream (EMLA â ).

During the procedure, animals were kept in the stocks and the devices were placed at the hearts base level.

Were evaluated the mean difference of paired measurements for systolic pressure, the correlation between paired measures for sys-tolic pressure and the percentage of all measurements for systolic pressure that lay within 10 and 20 mmHg of the reference method.

The mean difference of paired measurements for systolic pressure was 20.04 mmHg for the vascular Doppler device, 23.28 mmHg for the Dixtal 2710 â device and 31.95 mmHg for PetMap â device, all of them underestimated compared to invasive blood pressure. The correlation was 0.51 (P = 0.0002), 0.61 (P = 0.0001) and 0.60 (P = 0.0001) for vascular Doppler, Dixtal 2710 â and PetMap â device, respectively. For the vascular Doppler device, 24% of the measurements lay within 10 mmHg of the reference method and 43% within 20 mmHg. For Dixtal 2710 â device, 20% of the measurements lay within 10 mmHg of the reference method and 46% within 20 mmHg. For the PetMap â device, 7% of the measurements lay within 10 mmHg of the reference method and 17% within 20 mmHg.

In standing and non-sedated horses of this study, the three tested noninvasive blood pressure measurement devices did not reflect accurately the facial artery pressure, and thus did not fulfill the validation criteria proposed by the ACVIM consensus statement. Although cardiovascular complications arising from metabolic diseases are widely recognized in humans, similar reports are lacking in horses. Both dysregulated glucose uptake, which is mediated by glucose transporters (GLUT), and inflammation contribute to diabetic cardiomyopathy. In particular, pro-inflammatory cytokines are implicated in the pathogenesis of insulin resistance and impaired glucose uptake. Toll-like receptors (TLR), which mitigate the innate immune response, also play a key role during inflammation and metabolic dysfunction. This study aimed to investigate pro-inflammatory cytokines, TLR signaling and regulation of glucose transport in the equine myocardium during hyperinsulinaemia. Horses were treated (48 h) with a prolonged, euglycaemic hyperinsulinaemic clamp (p-EHC) or balanced electrolytes (controls). Left ventricular protein extracts were analyzed by Western blotting for GLUT (1, 4, 8, 12) , TLR4, TNF-a and IL-6 expression. Heart rate remained unchanged from baseline values for both groups. GLUT isoforms were not altered by hyperinsulinaemia either at the cell membrane or in total lysates. Down-regulation of TLR4 from the cell membrane (by 77%, p < 0.05) occurred in p-EHC-treated horses, without a change in TNF-a or IL-6 protein content. To further investigate cardiac TLR4 expression during hyperinsulinaemic conditions in vitro, isolated myocytes (rat) were incubated with high doses of insulin, which also resulted in down-regulation of TLR4 protein content compared to basal conditions. The results suggest that, in the absence of obesity, hyperinsulinaemia may directly decrease TLR4 activation in the heart, which may reduce immune system capability. Investigation of cardiac TLR4 signaling in insulin-resistant horses is required to further elucidate this provocative finding. Critically ill foals often present with clinical evidence of volume depletion and tissue hypoperfusion, which is reflected by hy- perlactatemia. Aldosterone and arginine vasopressin (AVP) are essential to maintain blood pressure, electrolyte balance and organ perfusion. Several studies have investigated the association between blood L-lactate concentrations with severity of disease and mortality in septic humans, horses and foals. However, information on aldosterone and AVP as it relates to hypoperfusion and L-lactate concentrations in critically ill foals are minimal. We hypothesized that septic foals will have higher L-lactate, aldosterone and AVP concentrations compared to healthy foals. We also proposed that hyperlactatemia will be associated with hyperaldosteronemia, hypervasopressinemia, clinical and laboratory markers of hypovolemia and non-survival in hospitalized foals. Blood samples were collected on admission from 96 septic (sepsis score >12), 182 sick non-septic (SNS; sepsis score <12), and 37 healthy foals of <3 days of age. Concentrations of aldosterone and AVP were determined by immunoassays.

L-lactate, aldosterone and AVP concentrations were higher in septic compared to healthy foals (p < 0.05). Non-surviving septic foals had higher L-lactate concentrations than survivors (p < 0.05). L-lactate concentration was positively correlated with aldosterone, AVP, creatinine and BUN concentrations in hospitalized foals (p < 0.01).The multivariate regression model showed a positive association between L-lactate concentrations, markers of hypoperfusion and sepsis score (p < 0.05). ROC curve identified an L-lactate cut-off value (7.9 mmol/L) that maximized sensitivity (72%) and specificity (68%) to predict non-survival in septic foals. Risk of non-survival increased with each 1 unit increase in L-lactate (OR=1.27, p = 0.001); aldosterone (OR = 1.12, p = 0.02) and AVP (OR=1.08, p = 0.023) concentrations in hospitalized foals.

Increased L-lactate, aldosterone and AVP concentrations were associated with non-survival in hospitalized foals; however, the L-lactate cut-off value to predict mortality in septic foals in our study was higher than in previous studies. Pituitary pars intermedia dysfunction can be diagnosed by performing a thyrotropin-releasing hormone (TRH) stimulation test. Use of this test is limited by the need for TRH to be purchased as a laboratory chemical and prepared on site. We hypothesized that compounded TRH (cTRH) solution from a commercial pharmacy would have the same effect on plasma ACTH concentrations as regular TRH (rTRH).

Thirty horses (mean age; 14.3 AE 3.6 years) from the same facility underwent two tests 24 apart. Ten horses were allocated to each group and received cTRH and then rTRH (Group 1), rTRH and then cTRH (Group 2), or rTRH on both days (Group 3). Plasma ACTH concentrations were measured at 0 and 30 minutes and delta ACTH values calculated.

Plasma ACTH responses were significantly lower on day 2 for groups 1 (P = 0.042) and 2 (P = 0.002). Mean (SD) delta ACTH values were 37.4 (28.9) and 18.9 (12.5) pg/mL for group 1, and 69.8 (54.1) and 27.3 (32.4) pg/mL for group 2 on days 1 and 2, respectively. Values were 30.9 (24.6) and 16.2 (14.6) pg/ mL, respectively for group 3 (P = 0.068). Delta ACTH values were 32.4 (30.3) and 53.6 (45.3) pg/mL, respectively for all cTRH and rTRH tests performed, and differed significantly (P = 0.006), yet the interpretation of results only differed in 3 of 20 horses.

An interval > 24 hours should be selected for test comparisons. Compounded TRH elicits ACTH responses in horses and further studies are required to establish cut-off values. Anhidrosis is a condition of substantial importance to working horses housed in hot, humid climates. Despite significant interest in the condition, an effective evidence-based treatment has not been identified. Forty-four recently anhidrotic horses were enrolled in the study on the basis of clinical signs and response to a quantitative intradermal terbutaline sweat test (QITST). Study horses were assigned randomly to treatment and control groups. Treated horses were given four acupuncture treatments at weekly intervals and herbal medication was topdressed on feed; control horses received sham acupuncture and hay powder. Horses were retested twice by QITST after completion of treatments (within two days and at four weeks).

In treated but not control anhidrotic horses, sweating responses to intradermal terbutaline increased significantly (P < 0.05) from baseline by completion of treatment but then returned to baseline within four weeks. Because there was also a nonsignificant increase in sweating in control horses after sham treatment, significant difference between groups was not achieved (P = 0.104).

Acupuncture and herbal medication may have improved sweating in recently anhidrotic horses, although the effect lasted less than four weeks after discontinuing treatment. This presumptive effect of acupuncture and herbal medication requires further investigation to fully validate its usefulness.

A RANDOM-IZED, BLINDED STUDY. BW Sykes 1 , KM Sykes 1 , GD Hallowell 2 . 1 Upper Orara, NSW, 2 University of Nottingham, Sutton Bonington, UK.

Omeprazole is widely used in the prevention of equine squamous gastric ulcer syndrome (ESGUS). Experimentally, doses of omeprazole as low as 0.7 mg/kg PO SID have been shown to result in potent suppression of both baseline and stimulated acid production. Doses as low as 0.5 mg/kg are registered in several countries around the world for the prevention of ESGUS yet, to date, the efficacy of such low doses has not been validated in the clinical setting. Thoroughbred racehorses in training without clinically significant (grade 2/4) squamous gastric ulceration were identified on gastroscopic examination. Horses were randomly assigned to one of two groups. Horses in the low and high dose groups received 250 and 500 mg (equivalent to 0.5 and 1.0 mg/ kg for a 500 kg horse), respectively, of enteric coated omeprazole orally SID. The omeprazole was administered 1-4 hours prior to exercise. Gastroscopy and ESGUS scoring was repeated at 21 -28 days. Data was both normally and non-normally distributed and analysed using Student's T test, Mann Whitney U test and Friedman test. Significance was assumed when p < 0.05.

Thirty-four horses met the inclusion criteria and were randomly allocated into two groups of 17 horses. One horse from the high dose groups was lost to follow-up for unrelated reasons and was excluded from data analysis. Eleven horses in each group had been treated with omeprazole previously and were identified on follow-up examinations, whereas the remaining horses, n = 6 and n = 5 in the low and high dose groups respectively, were identified as being ulcer free on their first endoscopic examination. There were no differences between the groups at enrolment regarding age (p = 0.31), weight (p = 0.47), time to follow-up (p = 0.42) or the number of race starts (p = 0.11). Mean squamous ulcer score (low dose 0.24 AE 0.6 vs. high dose 0.18 AE 0.5) was not different at enrolment between groups (p = 0.82). Mean ulcer score did not change over time in either the low (p = 0.83) or high (p = 0.78) dose groups. No effect of dose was observed over time (p = 0.49). Worsening of the ulcer score, to a maximum of grade 2/4, was observed in 2/17 and 1/16 horses in the low and high dose groups respectively; no difference was present between the groups (p = 0.52). In conclusion; both doses were equally efficacious in the prevention of ESGUS. The results of this study suggest that, under the conditions studied, doses of omeprazole as low as 0.5 mg/kg PO SID are as effective as higher doses in the prevention of ESGUS. Equine proliferative enteropathy (EPE) in horses is caused by L. intracellularis, an obligate intracellular bacterium. Clinical signs of this disease can include a decreased appetite, weight loss, diarrhea, and fever. Folate (FOL) and cobalamin (COB) play an important role in amino acid metabolism and in RNA and DNA synthesis and altered serum concentrations of these vitamins have been associated with gastrointestinal signs and systemic complications. Low serum COB concentrations are associated with increased serum methylmalonic acid (MMA) concentrations, which reflect the availability of COB on a cellular level. The study goal was to evaluate serum FOL, COB, and MMA concentrations in horses with EPE and a positive L. intracellularis titer.

Serum samples from horses (between postweaning and one year of age) with EPE (n = 41), in which L. intracellularis infection was confirmed by immunoperoxidase monolayer (IPMA) assay were evaluated. Also, serum samples from 40 horses without EPE were analyzed as negative controls (NC). Concentrations of serum FOL, COB, and MMA were measured using an automated immunochemiluminescence assay and gas chromatography-mass spectrometry, respectively. Results were compared between the two groups of horses using a Mann-Whitney U test. Also, a Kruskal-Wallis test was used to compare FOL, COB, and MMA concentrations based on L. intracellularis titers ( ! 480, 240, and no titer). Serum FOL concentrations were not different between the two groups (EPE horses: Horses with EPE had lower COB concentrations than control horses, but this difference did not reach significance. Horses with EPE also had significantly higher MMA concentrations than control horses. Further studies are needed to evaluate whether the lower cobalamin and higher MMA concentrations in EPE horses leads to metabolic consequences and/or systemic complications and whether cobalamin supplementation is indicated in these horses. Intra-abdominal hypertension (IAH) is the result of sustained increases in intra-abdominal pressure (IAP). Risk factors for IAH identified in people may be comparable to horses with colic due to abdominal/visceral distension, shock and hypoperfusion. IAH has been documented for individual horses, but this area remains minimally studied in cases of equine colic. The purpose of this study was to investigate abdominal pressures in horses with colic using a routine ventral abdominocentesis.

Abdominocentesis at the most ventral abdomen was performed in 53 horses with colic on admission to a referral hospital from 2011-2012 and 9 control horses without abdominal disease. The abdominocentesis cannula was connected to a saline primed line and direct IAP was measured using electronic manometry. IAP was measured in triplicate at the end expiration and mean values were used for statistical analysis. Differences in IAP between horses grouped according to their type of colic, treatment and outcome were assessed using unpaired t-tests, one-way ANOVA and Kruskall Wallis tests. P < 0.05 was significant.

Ventral IAP was significantly higher in horses with abdominal pain compared to controls (29 mmHg and 25 mmHg respectively); P = 0.025. 15/53 (28.3%) horses with colic had a ventral IAP greater than two standard deviations above the mean IAP of control horses and were considered to have IAH. 7/15 IAH horses had an obstruction of the large intestine and 5/15 IAH horses had an obstruction of the small intestine. No difference in IAP was found overall between survivors and non-survivors (28 mmHg and 29 mmHg respectively); P = 0.873. IAP was not different in horses receiving medical versus surgical management for colic (28 mmHg and 29 mmHg respectively); P = 0.587. IAP associated with small intestinal lesions was not different to large colon lesions (28 mmHg and 29 mmHg respectively); P = 0.829. Horses with large intestinal disease receiving medical management had a significantly higher IAP than horses with either small intestinal lesions receiving medical management or large intestinal lesions receiving surgical management (36 mmHg, 25 mmHg and 27 mmHg respectively); P = 0.0003.

Acute abdominal disease in horses is associated with increased IAP when obtained from the ventral abdomen. Horses diagnosed with medical large intestinal disease had the highest IAP measured, which may be attributable to impacted feed and gaseous distension within the colon. Further investigation is required to determine the true prevalence of IAH in horses with colic, whether alternative locations more accurately reflect IAH development and whether the magnitude of increase in IAP is associated with a need for therapeutic intervention. Among the different causes of colic, intestinal obstructions have been identified as the major cause of hospitalization and death in horses around the world. One cause of intestinal obstructions are enteroliths. Even though the survival has improved over time, the intestinal distension caused by the enteroliths has also been associated with poor prognostic and with postoperative complications. On account of this, over the last twenty years, several experimental models of intraluminal intestinal obstruction in horses were developed but many also have strangulating obstruction. Therefore, the aim of this study was to assess the effectiveness of small colon distension using a surgically implanted latex ball in the lumen.

For this purpose, eight healthy adult mixed-breed horses were subjected to a retro-umbilical celiotomy (Ethical approval: Comissão de Etica e Bem Estar Animal (CEBEA) -Protocol n o 007568-09). The selection of the manipulated segment was based on the presence of the mesenteric artery. After placing the area to be obstructed an incision in the antimesenteric surface was performed, the latex ball inserted oral to the incision and inflated to a pressure of 80 mmHg, as measured by a sphygmomanometer. After 240 minutes of distension, the ball was deflated and removed through a new incision. The enterotomy site was sutured and the bowel was then relocated in abdominal cavity. Throughout the procedure, animals were maintained with inhalatory anesthesia and controlled ventilation.

This model induced significant alterations in the small colon. At the time that ball was inflated thinning of the bowel wall was observed. After 240 minutes of distension, the presence of cyanotic regions due to enteric vessel stenosis was visualized, showing decreased mural perfusion. When the incision was made on the intestinal wall to remove the ball, it was thin and friable. Forty-five minutes after decompression serosa was still congested and intestinal wall thicker and more edematous in appearance.

The monitoring of physical parameters in the postoperative showed a significant increase in heart rate (55.25 AE 11.80 bpm) and body temperature (38.72 AE 0.52°C) after twelve hours of the ball removal. In peritoneal fluid analysis revealed an increase in total leukocyte (6.65 AE 6.69 x10 3 /lL) with four hours of obstruction, significantly increasing up to twelve hours (144.31 AE 100.14 x10 3 /lL). Also it was observed the kinetics of interleukin-6 in peritoneal fluid in this experimental model and results are similar to reported cases of obstructions in veterinary hospitals (significant increase after four hours of obstruction).

The implementation of a latex ball with pressure of 80 mmHg showed to be effective to mimic a moderate intestinal intraluminal obstruction, allowing more detailed research of this disease. Coagulopathy has been described in horses suffering from endotoxemia, and in LPS animal models. The purpose of this study was to evaluate the effect of a continuous rate infusion (CRI) of LPS on viscoelastic and standard coagulation parameters.

Ten healthy horses were administered Ecoli O55:B5 LPS intravenously at 125 ng/kg/hr for 4 hours. Platelet count, fibrinogen, d-dimer levels, PT, aPTT, and antithrombin, as well as viscoelastic parameters SonACT, clot rate (CR), and platelet function (PF) were collected prior to infusion, hourly during CRI, and at 1 and 24 hours post-infusion. Multi-level mixed model linear regression was used to evaluate the effect of LPS administration; a significance level of P 0.05 was used for all comparisons.

All horses survived LPS infusion, but developed tachycardia, tachypnea, fever, hyperemic mucous membranes, and leucopenia. LPS administration did not significantly change fibrinogen, ddimer level, antithrombin, PT, or aPTT. LPS decreased platelet count 2 hrs (P = 0.001) into the CRI; significant decline continued through 1 hour post-infusion. A decrease in PF was also observed, starting at 2 hrs (P = 0.002) and persisting through 5 hours. A significant decrease in SonACT was also observed, starting at 2 hrs (P = 0.006). CR significantly increased by 2 hrs (P = 0.001), and remained increased through 5 hrs.

Hemostatic dysfunction, characterized by decreased platelet number and PF developed as a result of LPS CRI. Viscoelastic parameters SonACT decreased and CR increased with LPS CRI, possibly suggestive of hypercoagulability. Viscoelastic monitoring of horses with suspected endotoxemia may be more clinically useful than standard coagulation monitoring. It is estimated that 8,000 human snakebites by venomous snakes occur annually in the United States. Of these snakebites, only five or six result in death. In horses, mortality rates are reported between 9-25%. Several retrospective studies exist that describe the complications, treatment and outcome of snake envenomation in horses. Complications described in these studies include cardiac disease, colic, colitis, laminitis, pneumonia, neuropathy and wound complications. The objective of this study was to describe clinical signs, clinicopathologic changes, treatment modalities and clinical outcomes in horses with evidence of colic associated with rattlesnake envenomation in the Rocky Mountain region of the United States by prairie rattlesnakes (Crotalus viridis viridis).

A retrospective, computer-generated search of the medical record database of equine cases evaluated for colic and/or snakebite envenomation from 2000 to 2012 was performed. Ninety-two horses with rattlesnake envenomation were identified based on history and physical examination findings. Of the 92 horses, eight horses exhibited signs of colic during hospitalization. Two horses presented for acute signs of severe colic and underwent an exploratory celiotomy. Both of the horses undergoing surgery and an additional horse developed nasogastric reflux. All three of these horses were euthanized or died prior to discharge.

Colic is an uncommon but life-threatening complication following prairie rattlesnake envenomation. Severe, acute colic or the presence of nasogastric reflux following envenomation may suggest a poor prognosis. In regions where the prairie rattlesnake is endemic, veterinarians should be aware of the possibility of colic secondary to rattlesnake envenomation, as clinical signs may mirror those of other forms of colic. Further studies are warranted to determine the pathophysiology involved in its occurrence and to identify ways to prevent and/or treat it. Horses are sensitive to the effects of endotoxemia and experience high mortality from diseases associated with bacterial sepsis. Endotoxin initiates an intense inflammatory response, yet antiinflammatory drugs have not shown clinical efficacy. Endotoxin is now known to enhance platelet reactivity to cytokine stimulation and activated platelets may in turn up-regulate inflammation.

We undertook a placebo-controlled double blinded study to determine whether administration of clopidogrel (Plavix â ), a platelet ADP-receptor antagonist, could modulate the proinflammatory effects of endotoxin in healthy horses.

Twelve healthy horses, randomized into groups of six, received clopidogrel (4 mg/kg loading dose then 2 mg/kg via nasogastric tube q24 h) for 3 days (Group 1) or a placebo (Group 2). On day 3 all horses were infused IV with 30 ng/kg endotoxin. The horses were then serially monitored to assess the following clinical and laboratory parameters: TPR, comfort score, CBC, TNFa, thromboelastogram, PFA100 closure time, platelet aggregation, and flow cytometric activation status. Regression analyses with curvilinear transformation of time was used to test for differences between groups.

Although comfort score, heart rate, rectal temperature, neutrophil and platelet count, and blood glucose changed significantly over time, we found no difference between the 2 groups in any of these parameters. Clopidogrel treatment did significantly influence lymphocyte counts (P = .005), platelet aggregation (P < .001), and P-selectin expression (P = .02).

Clopidogrel demonstrated anti-platelet activity, but did not attenuate the clinicopathologic effects of endotoxin infusion in horses. This study aimed to characterize for the first time the bacterial colonization of the intestinal tract of newborn foals by next generation high throughput sequencing. Three clinically normal foals born on a farm were enrolled. Fecal samples were collected by digital palpation after the first 24 hours of life. High throughput sequencing of the 16S rRNA gene PCR products was performed. Sequences were classified into operational taxonomic units (OTUs).

A total of 18,638 high quality sequences were obtained. Overall, Firmicutes and Bacteroidetes were the main phyla found on feces of newborn foals. There was remarkable species richness by 24 h of age, with 197, 412 and 791 OTUs detected after subsampling of 4453 reads per foal. Overall, Firmicutes and Bacteroidetes comprised the main phyla colonizing newborn foals (54% and 20%, respectively), which is similar to relative abundances found in adult horses. Unclassified bacteria at the phylum level accounted for 20% of OTUs. Five per cent of bacteria classified at the genus level had a relative abundance at or above 10% in at least one sample, but only 1% of these bacteria had the same relative abundance in all three foals. The most common organism shared between all three foals was an unclassified bacterium of the family Ruminococcaceae of the Clostridiale order.

The microbiome of the equine newborn is already complex after 24 hours of life and shows high variability among cohabitants of the same environment. The characterization of a core microbiome along with the investigation of unclassified bacteria may bring insights to the development of new therapeutic and prophylactic treatments, like development of probiotics, and a better understanding of the pathophysiology of neonatal diarrhea. Long-term management of nutrition and hydration in dysphagic horses is challenging and cost-prohibitive. In other species, percutaneous endoscopic gastrostomy (PEG) tubes are used to provide long term nutritional support. The purpose of this study was to determine the feasibility and to evaluate a technique for standing PEG tube placement in horses.

PEG tubes were placed into the air-inflated stomach using left intercostal approach in 6 adult horses. Proper location of the PEG tube was ascertained by percutaneous ultrasound and gastroscopy. Tubes were maintained for 14 days in 4 of 6 horses and were used as the sole source to provide fluid requirements. Meanwhile, horses were administered antibiotics, flunixin meglumine and omeprazole. Horses were monitored with serial physical examinations, complete blood counts and peritoneal fluid analyses. Horses were euthanized and underwent postmortem evaluation 7 days after tube removal. Quantitative data were analyzed by repeated measures ANOVA.

Placement of PEG tubes was feasible in all animals without notable complications. Vital parameters, body weight, blood and peritoneal white cell count did not change significantly during the study. Compared to baseline, median peritoneal protein (PP) and plasma fibrinogen (FIB) concentration increased by day 3, then decreased by day 14 (PP: 1.4, 3.6 and 2.8 g/dL; FIB: 232, 415 and 354 mg/dL, p < 0.05).

PEG tube placement and maintenance was well-tolerated and resulted in a mild transient non-septic peritonitis. Fluid requirements could be adequately met with this technique. Further research is warranted to evaluate the feasibility of enteral feeding using this novel approach in horses. Colic in neonatal foals is often caused by meconium retention. Routine treatment includes enema administration to soften meconium and aid resolution. In refractory cases retention enemas performed under heavy sedation can become necessary. An intravenous dose of N-Butylscopolammonium-Bromide (NBB) may increase the volume of retained enema due to its anti-spasmodic effect and hence optimize the procedure. The study aim was to evaluate the effect of NBB on retained enema volume in anesthetized foals.

Five (5) healthy foals between 5 and 21 days of age were included in a blinded, randomized, cross-over study with two treatments: 1) NBB, 0.3 mg/kg IV and 2) 0.9% saline. Abdominal CT imaging was performed under injectable anesthesia before, immediately after and 10 minutes post contrast enema (4 ml/kg 0.6% Iohexol solution per rectum via Foley catheter). Treatments were administered IV immediately prior to enema infusion. Heart rate, respiratory rate and blood pressure were monitored. Volume estimation and 3D imaging of contrast enema was performed using OsiriX software.

The percentage of retained enema volume at 10 minutes did not differ between foals receiving NBB (59.3% AE 33.4;MeanAE SD) or saline (55.5% AE 32.3). No adverse effects were noted.

The use of NBB did not increase the amount of retained enema relative to saline in this study, but variability was larger than expected. Further investigation of NBB in foals is warranted. Toll-like receptors (TLR) not only play a central role in pathogen recognition by the innate immune system, but are also activated by endogenous ligands. TLR4 signalling stimulates proinflammatory cytokine release and plays a critical role in linking inflammation and metabolic disease. Although an association between equine laminitis and metabolic dysfunction has been proven, the disease pathogenesis is poorly understood. This study aimed to investigate TLR4 signaling in two models of insulininduced laminitis. Standardbred horses were treated with a prolonged-euglycaemic, hyperinsulinaemic clamp (p-EHC), hyperglycaemic clamp (HC) or balanced electrolytes (controls). Clamp-treated horses developed clinical (p-EHC) or subclinical (HC) laminitis, whereas controls did not. Skeletal muscle and lamellar protein extracts were analyzed by western blotting for TLR4, IL-6, TNF-a and suppressor of cytokine signalling (SOCS-3) expression. Increased lamellar TLR4 and TNF-a protein expression occurred without change in IL-6, although a trend (p = 0.09) for increased SOCS-3 content was noted, in p-EHC-treated horses compared to controls. In addition, lamellar SOCS3 protein content was positively correlated with TLR4 in p-EHC-treated horses (R 2 = 0.88, p < 0.05). The p-EHC and HC did not alter TLR signaling in skeletal muscle or lamellae respectively. To our knowledge, this is the first study of equine lamellar TLR signaling. Activation of TLR4 signaling suggests that lamellar inflammatory pathways are activated by the clinical onset of insulin-induced laminitis, a finding consistent with histopathological reports. However, the lack of activated TLR signaling during subclinical laminitis does not support a role for proinflammatory cytokines in pre-clinical hyperinsulinaemic laminitis pathophysiology. Pneumonia is the most common cause of morbidity and mortality in foals less than 6 months of age; however, adult horses, unless immunocompromised, are not as susceptible to such respiratory infections including Rhodococcus equi. We speculate that the pulmonary cytokine environment differs between the neonate and adult thereby imparting differing phenotypes to pulmonary alveolar macrophages (PAMs). This could result in differences of microbicidal activity between adult and foal pulmonary-alveolar macrophages (PAMs). Bronchoalveolar lavages (BALs) were performed in foals at week 1, month 1, month 6, and month 12 of age as well as in adults >6 years of age. Aliquots of bronchoalveolar lavage fluid (BALF; n = 12) were analyzed via a 300 cell count differential by a clinical pathologist. Foals at week 1, month 1, and month 2 showed an increased number of macrophages and significantly decreased number of lymphocytes as compared to month 12 samples and adult controls (p < 0.00001). Month 12 foals and adults had very similar cell populations (p < 0.5). PAMs from week 1, month 1, month 6 and month 12 (n = 3) were stimulated with and infected in vitro. Stimulants included LPS/IFNc, opsonized zymosan, and heat-killed R.equi antigen. Infection with R. equi was enumerated by dsRed and GFP fluorescence permitting concurrent measurement of reactive intermediates and bacterial load. Foals at week 1 exhibited lower levels of peryoxynitrite and superoxide production as compared to their counterparts at month 12 (p < 0.05). Consistent with our hypothesis, PAMs derived from foals yielded a higher bacterial load than in adults (n = 2; p < 0.01) indicating increased foal PAM susceptibility to R. equi infection. BALF (n = 4) was also analyzed for IFNc, IL-4, IL-10 via Luminex and TNFa via ELISA to analyze the role of age-associated cytokine milieu on macrophage phenotype. Results indicate an age associated increase in pro-inflammatory cytokines (IFNc and TNFa) relative to anti-inflammatory cytokines (IL-4 and IL-10) (p < 0.0001 for week 1 versus adult; p < 0.01 for month 1 versus adult). To further characterize the role of cytokines in susceptibility to infection, PAMs (n = 3 for each age) were stained for extracellular CD172a and for intracellular IL-4, IL-10 & IFNc for evaluation by flow cytometry. CD172a+ gated PAMs produced more IFNc but less IL-4 and IL-10 by 12 months of age (p < 0.1). The greatest difference in intracellular cytokine production (p < 0.02) was between the month 1 vs. month 12 age groups. Taken together, these critical differences indicate that foal PAMs are more susceptible to infection with Rhodococcus equi and support the notion that the pulmonary cytokine milieu impacts the microbicidal response of the macrophage. Strenuous exercise may be accompanied by endotoxemia, systemic pro-inflammatory changes and can also influence immune response. Exercise-induced inflammation has been studied in human and horses. Therefore, the aim of this study was to determine mRNA leukocyte expression for TNF-a, IL-1b and IL-6 in healthy trained Arabian horses during prolonged exercise.

For this purpose, five trained sound Arabian horses were submitted to an 80 km endurance ride with three intermediate veterinary check points (Vet check). Blood samples were collected before exercise, at every intermediate vet check (Vet1, Vet2, Vet3) and at the end of the endurance ride (Vet4) (Ethical Committee CEUA Protocol 001624/11). Expressions of TNF-a, IL-1b and IL-6 genes were evaluated by real time qRT-PCR with Taq-Man â . Cross threshold (Ct) values for each sample were normalized to endogenous HPRT and cytokine mRNA expression in each sample expressed as relative change using the DDCt method. The fold change was evaluated for all samples relative to preexercise mRNA expression levels to assess the effects of exercise.

Expressions of IL-1b and IL-6 increased from Vet2 (2.7-fold and 3.2-fold, respectively), reaching peak values at Vet3 (3.5-fold and 5.2-fold). At Vet4 expression of this cytokines increased 3.2fold and 2.8-fold. Expression of TNF-a remained near one during most sample times.

The IL-1b is one of the most important markers of inflammatory response and with IL-6 plays a central role in host defense, stimulating neutrophils and acute phase response. These findings indicate that an 80 km endurance ride induces early pro-inflammatory condition that may predispose horses to post-exercise complications. Rhodococcus equi, a facultative intracellular pathogen and an important cause of pneumonia in foals, is highly susceptible to killing by gentamicin in vitro. However, gentamicin does not appear to be effective in vivo, due to its poor cellular penetration. Encapsulation of drugs in liposomes enhances cellular uptake. The objective of this study was to compare the efficacy of two different formulations of liposomal gentamicin to that of free gentamicin and of other antimicrobials, for the treatment of R. equi in a mouse infection model.

Athymic nude mice were infected intravenously with 5 9 10 6 CFU of virulent R. equi. On day 4 after infection, mice were treated intravenously with 2 different formulations of liposomal gentamicin (with and without polyethylene glycol (PEG) coating), empty liposomes, free gentamicin, subcutaneous rifampin and clarithromycin, or saline (5 mice per group). Mice were subjected to euthanasia 8 days post-infection. Effects of drug on CFU in spleen and liver were assessed using an ANOVA.

Mice euthanized 8 days post-infection and treated with PEGcoated liposomal gentamicin had significantly (P = 0.005) lower CFUs of R. equi in the spleen and in the liver compared to control mice or mice treated with free gentamicin. Compared to treatment with clarithromycin-rifampin, treatment with PEGcoated liposomal gentamicin resulted in a significantly (P = 0.036) greater reduction in the numbers of R. equi CFU in the liver relative to untreated controls.

These results underscore the potential of liposomal gentamicin as a new treatment for infections caused by R. equi. Equine granulocytic anaplasmosis (EGA) and Lyme borreliosis (LB) are an emerging concern in Canada. Risk of exposure is poorly understood but clinical disease has been reported from provinces without known established tick populations. The objectives of the study were to estimate seroprevalence of EGA and equine LB in Saskatchewan (SK), Manitoba (MB) and Ontario (ON) and to investigate agreement between a point-of-care ELISA and laboratory-based serologic tests. Convenience serum samples from veterinary diagnostic laboratories in SK (n = 202), MB (n = 139) and ON (n = 35) were tested using the SNAP 4Dx ELISA (IDEXX Laboratories, Inc., Westbrook, ME). Samples testing positive for EGA (n = 2) or LB (n = 6) and a randomized subset of samples testing negative (n = 92 each) were retested by indirect fluorescent antibody test (IFA) for EGA, or whole cell ELISA confirmed with Western Blot (WB) for LB, in a commercial veterinary diagnostic laboratory. Antibody titers >1:80 for IFA and >1:160 for whole cell ELISA (when WB confirmed) were considered positive. Western Blot positive samples included a subset of equivocal results (suspect antibody reactivity). Test agreement was assessed by use of the prevalence adjusted and bias adjusted kappa (PABAK) statistic. Based on the SNAP 4Dx ELISA results, overall seroprevalence of EGA was 0.5% (95% CI: 0.06 -1.9%) while seroprevalence in SK, MB and ON, respectively, was 0.5%, 0.7% and 0% (95% CI: 0-2.7%, 0-3.9% and 0-10%). Overall seroprevalence of LB was 1.6% (95% CI: 0.6-3.4%) while seroprevalence in SK, MB and ON, respectively, was 0.5%, 2.9% and 2.9% (95% CI: 0-2.7%, 0.8-7.2% and 0.07-14.9%). All SNAP 4Dx ELISA positive samples tested positive in laboratory-based tests, whereas 25/92 (EGA) and 24/92 (LB) SNAP 4Dx ELISA negative samples tested positive in laboratory-based tests. Agreement between the SNAP 4Dx ELISA and laboratory-based tests was moderate (for IFA, PABAK = 0.47, 95% CI: 0.29-0.65, for whole cell ELISA/ WB, PABAK = 0.51, 95% CI: 0.34-0.68).

Conclusions: While the SNAP 4Dx ELISA yielded expected seroprevalence estimates, results showed only moderate agreement with laboratory-based serologic tests. This may be attributable to false positive (possibly due to cross reactivity) or false negative results. The performance of the SNAP 4Dx ELISA in comparison to laboratory-based serologic tests in horses requires further investigation. Salmonella enterica is commonly recognized as a cause of hospital acquired infections as well as zoonotic infections in veterinary teaching hospitals (VTHs). Environmental contamination is frequently associated with nosocomial transmission in veterinary hospitals. The objective of this study was to determine risk factors associated with environmental contamination of a veterinary teaching hospital with S. enterica.

Environmental samples included in this research were collected monthly for routine surveillance from March 2003 through June 2011, using a commercially available electrostatic wipe as part of an infection control program. Sampling sites included floor and hand contact surfaces throughout the VTH. Risk factors for culture-positive environmental samples that were evaluated included measures describing sampling location, hospital case load, disease severity of patients, presence of culture-positive inpatients, and season. Data on risk factors were collected retrospectively from the VTH medical records database. Multivariable logistic regression was used to determine associations between hospital risk factors and veterinary hospital environmental contamination with S. enterica.

During the study period, on average, 47 samples were collected monthly, for a total of 5273 environmental samples. Of the samples collected, a total of 8.2% (n = 434) were culture positive for S. enterica using enriched culture techniques. In general, environmental samples collected in the Food Animal Hospital and from floor surfaces were more likely to be positive, as were samples collected during the warmest third of the year (July through October). Additionally, for equids, a greater number of hospitalization days in the month prior to sampling and higher density of severely ill horses were also associated with increased risk for positive environmental samples.

Risk factors identified in this study will allow for the refinement of existing infection control programs. A better understanding of the risk factors associated with environmental contamination will allow for more practical evidence based preventive measures to be implemented in veterinary hospitals experiencing epidemics of hospital acquired infections with S. enterica. E-47 EMERGING OUTBREAKS ASSOCIATED WITH EQUINE CORONAVIRUS IN ADULT HORSES. N Pusterla 1 , S Mapes 1 , C Wademan 1 , A White 1 , RL Ball 2 , K Sapp 2 , P Burns 3 , C Ormond 4 , K Butterworth 5 , J Bartol 6 , R Steere 7 , KG Magdesian 1 . 1 School of Veterinary Medicine, University of California, Davis, CA, 2 Bracken Equine Clinic, San Antonio, TX, 3 Elkhorn Veterinary Clinic, Elkhorn, WI, 4 Oak Hill Veterinary Services, Orinda, CA, 5 SRH Veterinary Services, Ipswich, MA, 6 New England Equine Medical & Surgical Center, Dover, NH, 7 Artaurus Veterinary Clinic, Petaluma, CA.

Equine coronavirus (ECoV) has been identified by electron microscopy, culture and more recently by PCR in feces of foals with and without enteric disease. A recent study reported on the isolation of ECoV from the feces of 2-to 4-year-old horses with pyrogenic and enteric disease living in stables of a racetrack in Japan. However, little is known about ECoV, especially with regard to molecular diagnostics of field samples and the clinical significance of ECoV PCR positive fecal results. The purpose of this study was to describe clinical, hematological and fecal PCR results from 231 horses involved in outbreaks associated with ECoV.

The outbreaks happened at five separate boarding facilities between November 2011 and August 2012 in the States of California, Texas, Wisconsin and Massachusetts. The population of horses per stable ranged from 28 to 70 horses. Following the molecular detection of ECoV in the feces from the initial index cases, the remaining herdmates were closely observed for the development of clinical signs. Fecal samples were collected from sick and healthy horses for the PCR detection of ECoV. Clinical pathology from sick horses was evaluated when available.

All four outbreaks involved primarily adult horses. Seventythree horses developed clinical signs with 13 to 16 sick horses per outbreak. The main clinical signs reported were anorexia (65), lethargy (56) and fever (53). Changes in fecal character and colic were observed in 15 and 6 horses, respectively. Clinical signs generally resolved within 1-4 days with supportive care. Five horses from 4 different outbreaks were euthanized or died due to rapid progression of clinical signs. The cause of death could not be determined with necropsy evaluation in 2 horses, while septicemia secondary to gastrointestinal translocation was suspected in 3 horses. Blood work was available from 13 horses with clinical disease and common hematological abnormalities were leucopenia due to neutropenia and/or lymphopenia. Feces were available for ECoV testing by real-time PCR from 49 and 141 sick and healthy horses, respectively. 42/49 (86%) horses with abnormal clinical signs tested PCR positive for ECoV, while 134/141 (95%) healthy horses tested PCR negative for ECoV. The overall agreement between clinical status and PCR detection of ECoV was 93%. The study results suggest that ECoV is associated with self-limiting clinical and hematological abnormalities in adult horses. Real-time PCR is a sensitive and fast diagnostic tool to document the presence of ECoV in feces from horses with unspecific clinical signs. An improved model to study Rhodococcus equi pneumonia in foals is needed to better investigate vaccine and other prophylactic measures. Current models lead to acute, severe pneumonia with high mortality which does not mimic natural infection.

Eighteen 1-week-old foals were challenged using intratracheal instillation of 5 different doses (10 6 , 10 5 , 10 4 , 10 3 and 10 2 cfu/foal) of VapA+ R equi (206UKVDL). Additional foals were challenged at 2 (n = 4) and 3 (n = 6) weeks of age using 10 3 cfu/foal. Foals were observed daily and physical exam was performed twice a week. Thoracic ultrasound and blood work were performed weekly. Foals were euthanized 6 weeks after challenge or sooner if pneumonia developed. Lung lesions were scored at necropsy.

There were significant (p < 0.05) changes in physical exam and blood work over time after the high dose (10 6 , 10 5 , 10 4 ) challenges. Low dose foals (10 3 ,10 2 ) survived significantly longer than those receiving the higher doses (p < 0.001). All foals challenged at 3 weeks of age survived to the end of the study. Lung lesions were significantly more severe (p = 0.002) for high dose foals and for foals challenged at 1 and 2 vs 3 weeks of age (p = 0.043).

The clinicopathological findings after a low dose challenge of 1-week old foals are similar to those observed in field cases. This model provides a more realistic approach for assessing efficacies of vaccines and other treatments. Nosocomial infections with Salmonella spp have resulted in the temporary closure of equine hospitals thus surveillance for this organism is critical. Accurate and rapid identification of Salmonella spp is necessary for adequate management and timely implementation of infectious disease control protocols. The gold standard for detection of Salmonella spp in fecal or environmental samples is bacterial culture. However, results are not available for 3-5 days. PCR results are available within 24-36 hours but there is a higher risk of false positive results. In the food industry nucleic acid hybridization is used to identify contamination with Salmonella spp.This technology targets the pathogen's ribosomal RNA by utilizing capture and detector DNA probes specific to the rRNA of Salmonella. Results are rapidly available (27 hours) and accurate (sensitivity 98.9% and specificity 99.7%). The purpose of this study was to use nucleic acid hybridization technology (GeneQuence â ) to detect Salmonella spp in fecal samples and environmental samples from an equine hospital and compare the results to bacterial culture.

Routine fecal and environmental samples collected from April 2010 -April 2011 at Rood and Riddle Equine Hospital were used. The fecal samples were enriched in selenite broth and incubated for 24 hours, then plated on Hekton agar and incubated for 24 hours. The environmental samples were incubated in peptone broth for 24 hours, then tetrathionate broth for 24 hours. Afterwards the samples were plated and incubated on Hekton agar for 24 hours. After appropriate incubation any black colonies on the Hekton agar plates were gathered and transferred to the GeneQuence â D2 automated system for nucleic acid hybridization and to BBL crystal identification system TM for the final phase of microbiological culture identification. The GeneQuence â results were available within 2 hours whereas the bacterial culture results were available within 24 hours.

1494 samples were analyzed. The results were: 1274 samples were GeneQuence â negative and culture negative, 210 samples GeneQuence â positive and culture positive, four samples Gene-Quence â negative and culture positive, and six samples Gene-Quence â positive and culture negative. The overall sensitivity was 98.1% and the specificity was 99.5%. The positive predictive value was 97.2%, whereas the negative predictive value was 99.6%.

GeneQuence â was able to detect the presence of Salmonella spp in fecal and environmental samples. The results were available 1 day sooner and were almost as accurate as the bacterial culture, which is still required to identify an antimicrobial sensitivity.When compared with PCR, GeneQuence does not require amplification because it targets genetic material (rRNA) that is more abundant and there are fewer false positive results. Gene-Quence â is a rapid and reliable test for identification of Salmonella spp in equine hospitals. Septicemia involving gram negative bacteria is a common clinical disorder in neonatal foals. Lipopolysaccharide (LPS), associated with gram negative bacteria, plays a pivotal role in host response to bacterial invasion. Possible sequelae associated with endotoxemia include overzealous activation of the inflammatory response and coagulation cascade and derangements in multiple organ systems Polymyxin B (PMB) functions as a chelating agent and binds the lipid A portion of LPS, thus neutralizing LPS and negating the interaction of LPS with cellular receptors that result in inflammation. The objective of this study was to determine if beneficial clinical and biochemical effects are observed in foals administered PMB in an endotoxin model. Fourteen healthy neonatal foals were administered 0.5 mcg/kg of LPS (055:B5 E. coli) IV over 30 minutes. The treatment group (7 foals) received 6000 U/kg of PMB IV at the end of the LPS infusion while the control group (7 foals) received 0.9% saline IV. Blood was collected at Time -24, -0.5, 0, 0.5, 1, 1.5, 2, 3, 4, 5, 6, 12, 24 and 48 hours and physical examination parameters and an attitude score were recorded. CBC, blood lactate and glucose, and serum TNF-a and TXB2 were also measured.

The treatment group had improved physical examination, attitude, biochemical and inflammatory parameters at various time points throughout the study compared to controls. These data suggests that PMB may improve some clinical and biochemical parameters in an endotoxemia model in healthy foals and may have potential benefit in clinical cases of equine neonatal septicemia. Several studies have evaluated the use of PCR for the detection of Salmonella spp. in fecal and environmental samples. Although PCR has increased sensitivity and faster turn-aroundtime, the costs associated with this testing platform are higher than microbiological culture. The purpose of this study was to evaluate the pooling of feces and environmental samples following a selective enrichment culture step for the detection of Salmonella spp. by real-time PCR.

For the purpose of this study, 140 equine fecal and 280 environmental samples were collected at three veterinary hospitals. Feces and environmental samples were inoculated into 100 ml of selenite or tetrathionate broth and incubated for 18-24 hours. Following incubation, the enrichment broth samples were subcultured on xylose-lysine-tergitol-4 or hektoen agar plates. Suspected Salmonella spp. colonies were subcultured and further identified using biochemical assays. Concurrently to the microbiological analysis, 1 ml of the enrichment broth from fecal and environmental samples was processed for DNA purification using standard protocols. Each sample was analyzed individually (420 samples) and pooled in groups of ten by sample type (42 pools) using an already established and validated Salmonella spp. realtime PCR assay targeting the invasion A gene. A panbacterial real-time PCR assay was also used in order to ensure proper DNA extraction and rule out inhibition due to contaminants.

All 280 environmental samples tested negative by culture, and single and pooled PCR. Seven fecal samples from horses with gastrointestinal diseases cultured positive for Salmonella spp. Nine out of 140 fecal samples, including the seven culture positive samples, tested PCR positive for Salmonella spp. The additional two PCR positive samples originated from one horse diagnosed with sand impaction and a follow-up sample from a horse previously diagnosed with salmonellosis. Seven pools tested PCR positive for Salmonella spp. Five pools each contained one single positive sample, while two additional pools contained two positive samples.

The pooling strategy using feces and environmental samples incubated for 18-24 hours in selective enrichment broth was able to detect all culture and individual PCR positive samples. In this study scenario, the number of microbiological cultured (420) was significantly higher than the number of PCR analyses (42 pools at 10 samples each and 70 individual samples to determine the positive sample (s) out of each positive pool). This strategy appears to be cost-and time-effective in a hospital environment with a low prevalence for Salmonella spp. Methicillin-resistant Staphylococcus aureus (MRSA) is a multiple-drug resistant organism that has become a well known nosocomial infection in both human and veterinary medicine and may have zoonotic potential. Community acquired MRSA strains have recently been identified in non-hospitalized equine populations. There have been increasing reports of MRSA colonization and clinical disease in horses and people working with animals. All studies on equine MRSA include sampling from the nasal passages or the clinical infection site. It has been shown in humans that the prevalence of MRSA infection can be much higher when examining the throat. No studies have been performed sampling the nasopharynx and guttural pouches of the horse. The purpose of this study was to determine the prevalence of MRSA in equine nasopharyngeal and guttural pouch wash samples. Gram-positive, catalase-positive cocci isolated from nasopharyngeal and guttural pouch washes from 186 equine samples submitted to the New Bolton Center Clinical Microbiology Laboratory primarily for detection of Streptococcus equi were speciated and tested for penicillin binding protein by the slide agglutination test.

Eight methicillin-resistant Staphylococcal samples were isolated (4.3%), one of which was MRSA. The remaining 7 were identified as Staphylococcus pseudointermedius.

This study showed that MRSA prevalence in equine throat samples is low and appears to be similar to equine nasal carriage prevalence in low risk community populations. Equid Herpesvirus type 1 (EHV-1) is a highly contagious agent for horses, able to cause outbreaks of primary respiratory disease, retinopathy, myelopathy and/or abortion. Horizontal transmission is directly through nasopharyngeal droplet transmission, or, indirectly, through fomite transmission. Once expelled into the environment viral maintenance of infectivity will depend on a variety of factors but most importantly, will rely on viral envelope integrity. Surface tension forces, temperature fluctuations and UV-light exposure have shown with significant effects on envelope integrity in other herpesviridae.

We hypothesized that viral survival will be different if placed on various surfaces or materials, and if placed in different environmental conditions. An EHV-1 suspension was placed on surfaces or materials: plastic, fabric, leather and stall bedding materials shavings and straw. Materials were placed in different environments: constant 4°C, 'barn' and 'outdoor' environment'. Samples of each material and environment were collected at time points 0, 3, 12, 24, 36 and 48 hours. In the laboratory supernatants of materials immersed in 2 ml of virus culture medium, spun were collected and stored frozen. Collectively, samples were thawed and viral titration was done using a plaque assay. Statistical analysis used generalized linear models with random effects mixed models controlling for repeated measures. Statistical significance was assumed when p < 0.05.

Results showed significant differences upon first contact (t = 0) of the viral suspension with different materials, most noticeable with shavings and leather. Most materials and in environmental conditions other than 4°C showed a rapid decrease in viral survival, especially during the first 3 hours.

While results show significant reduction on some surfaces and materials over others, it is important to realize that viral maintenance of infectivity was still significant under simulated 'barn conditions' following the 3 hour time point. These results emphasize the importance of following diligent hygiene, individualized attire, isolation protocols, and the prudent use of biosecurity protocols when mitigating an EHV-1 outbreak. Bacterial culture and polymerase chain reaction (PCR) of nasopharyngeal washes (NPW) and guttural pouch lavage (GPL) samples have been used for both diagnostic testing and the detection of carrier animals for Streptococcus equi (S. equi), but have had low sensitivity. Previous in vitro studies were performed to determine the optimal laboratory technique and the detection limit for S. equi cells in 0.9% NaCl. This field study compared the Copan TM flocked swab culture and the Copan TM flocked swab PCR methods with the results of a direct-PCR (gold standard) in 193 NPW and GPL clinical samples from sick (134) and not sick (59) (convalescing and subclinical) horses to verify the previous in vitro results. We hypothesized that: the direct-PCR would detect S. equi in NPW and GPL samples more reliably than flocked swab culture; that the flocked swab-PCR method would detect S. equi in NPW and GPL samples with equivalent frequency as direct-PCR; and that the flocked swab culture would detect S. equi in NPW and GPL samples more reliably than traditional culture. 35 samples were positive for S. equi via the gold standard (direct PCR). The direct PCR will be submitted to the Clinical and Laboratory Standards Institute for consideration as a national standard operating procedure for the collection and processing of NPW and GPL samples for S. equi from horses. Neonatal Maladjustment Syndrome (NMS) is one of the most common diseases of the neonatal foal. Also called "dummy foal syndrome," clinical signs range from disinterest in the mare and inability to nurse to obtundation, recumbency, and seizures. The pathophysiology of NMS is not fully understood, and has often been attributed to hypoxic damage to the brain induced by perinatal asphyxia. However, foals diagnosed with NMS do not always have a history of such events, or hypoxic histopathologic changes in the brain at necropsy. An alternate hypothesis for the pathogenesis of NMS is the persistence of high concentrations of pregnanes. Pregnanes, allosteric modulators of the GA-BAA receptor, are normally elevated at birth and decrease rapidly within 48 h. Foals with NMS have previously been shown to have persistently elevated plasma pregnane concentrations.

The aim of this prospective study was to induce clinical signs consistent with NMS by the infusion of the pregnenolone derivative allopregnanolone, and to report the resulting clinical signs and electroencephalogram (EEG) recordings. Three healthy neonatal foals were recruited for this study. One foal was infused with saline, one was infused with 5% ethanol, and one foal was infused with allopregnanolone in 5% ethanol solution. EEG recording, serial physical examinations and neurobehavioral scores were performed before, during and after the infusions. Foals were assigned neurobehavioral scores of 0-18 using a previously described scoring system.

Physical examination and neurobehavioral scores of foals administered saline and 5% ethanol solution were within normal limits at all time points. The foal infused with allopregnanolone experienced transient bradycardia (48 bpm) and decrease in gastrointestinal borborygmi. Neurobehavioral score increased to 12 post-infusion of allopregnanolone, and was characterized by moderate obtundation, lack of awareness of the dam, absent suckle reflex, and lack of tone in the ears. EEG recording of all foals showed normal brain waveforms and normal slow wave sleep while the foals were recumbent prior to infusions. Foals administered saline and 5% ethanol continued to have normal EEG recordings, whereas the foal administered allopregnanolone had brain wave patterns similar to slow wave sleep while standing. Abnormal EEG recordings correlated with bradycardia and increased neurobehavioral score, but persisted longer than clinical signs. This study concluded that infusion of allopregnanolone successfully induced clinical signs compatible with NMS. Idiopathic headshaking has been recognized for over a hundred years and is a spontaneously occurring nociceptive and potentially crippling disorder of mature horses. Typical headshaking behavior includes sudden, violent flicks of the head, excessive snorting, striking at the nose and trying to rub the nose on the thoracic limb or ground. Although the etiopathogenesis of headshaking remains elusive, trigeminal neuralgia is regarded as the likely explanation of observed clinical signs.

The aim of this study was to compare sensory nerve conduction threshold and velocity of the trigeminal nerve using the infraorbital nerve in control and headshaking horses.

Control (n = 6) and headshaking (n = 6) horses were subject to general anesthesia and trigeminal nerve conduction studies using a Nicolet Viking evoked potential system. A pair of stimulating electrodes was placed at the gingival mucosa of the maxillary canine. Four pairs of recording electrodes were placed along the tract of the infraorbital nerve (point 1), maxillary nerve (point 2), spinal somatosensory (point 3 at C1), and cortical somatosensory evoked potentials (point 4 at fronto-parietal cerebral cortex). A reference electrode was placed between the stimulating and point 1 recording electrodes. Stimuli (0.1 ms) were applied at 2.5, 5, 10, 15, 20 mA.

The threshold of sensory nerve action potential occurred at low stimuli (2.5 and 5 mA) in horses with headshaking; and at higher stimuli (10 mA in 3 horses, 15 mA in 1 horse, and 25 mA in 2 horses) in control horses. Conduction velocity was not significantly different between groups.

Headshaking horses have a low threshold for inducing sensory action potentials upon minimal stimulation compared to control horses supporting involvement of the trigeminal nerve in the pathogenesis of affected horses. Further work validating equine headshaking as an animal model of human trigeminal neuralgia is warranted. The severity of ataxia in horses is graded on a scale from 0 (normal) to 5 (recumbent, unable to rise). The correct assessment of ataxia in horses is critical to diagnostics, treatment, and evaluation of response to treatment. However, this grading scale is somewhat subjective.

The objective for this study is to simulate ataxia in horses through drug administration, and use detailed quantifiable gait analysis to create a more objective grading scale for ataxia in horses.

For this study we used 8 adult Arabian horses, assessed before and after administration of 2 doses of xylazine. Physiological data were collected at baseline and 5, 10, 20, 30, 45, and 60 minutes after xylazine administration. Gait analysis took place between 5-10 min after xylazine and included: kinematic data collected on the Sato equine high-speed treadmill (Flat and Decline -10%), accelerometer data collected simultaneously, and kinetic data collected while horses were standing on a Kistler Instruments force plate.

Xylazine administration resulted in reduced heart rate, head height, lip tone, and ataxia. Data analysis shows an increased stride duration and stride length, decreased stride frequency, and increased side-to-side movement of the hind-quarters following xylazine administration. Data suggests that increased postural sway can be determined in horses after xylazine administration, through quantification of center of pressure measurements.

We can induce ataxia in horses using xylazine and we have determined numerous quantifiable gait parameters that are affected by sedation. Future work includes development of a scale incorporating these parameters and validation in clinically ataxic animals. Treatment of idiopathic headshaking in horses is complicated by incomplete understanding of underlying pathophysiology and partially effective treatments. If an inflammatory etiology exists, corticosteroids may be beneficial. This prospective, blinded clinical trial was designed to determine if an anti-inflammatory dose of dexamethasone relieves the signs of headshaking in a field setting. Cases were recruited from the general population and diagnosed by attending veterinarians. Pulsed dosing was oral dexamethasone (40 mg PO QD x5d, q3 weeks for 4 months) or placebo (inert syrup). Owners were blinded and asked to score the headshaking from 0-4 (4 = most severe) 3 days per week. The direction of change from median pre-to post-treatment headshaking scores (HS) was compared between groups by Fischer's exact test.

Twenty horses entered the study with twelve completing the trial. There was no significant difference between the numbers of horses in each group that had a decreased HS after any pulsed dosage (P = 0.38). The median change in HS after the first pulse was -0.25 (range: -1.5 -1) in placebos and -0.75 (range: -4 -0) in treated horses (P = 0.38). The median change in HS was the same after the third pulse. Although a consistently greater improvement in headshaking score was seen in the experimental group, this finding failed to reach statistical significance. The study was limited by small sample size, poor owner compliance and by the underlying seasonality of signs. No clear benefit was detected of dexamethasone therapy for idiopathic headshaking. Polocrosse is an equestrian sport similar to polo. Despite anecdotal reports of muscle pain and stiffness being common after competition there is no published data on the prevalence of, or risk factors for, exertional rhabdomyolysis (ER) in polocrosse horses. The purpose of this study was to report the prevalence of, and to investigate potential risk factors for ER in polocrosse horses.

A cross-sectional study of horses competing at the Australian National Polocrosse Championships in Warwick, Queensland, Australia in April 2012 was conducted. Owners were informed of the objectives of the study and asked to volunteer their horses on a team by team basis. To reduce selection bias at least 4 out of the 7 horses from each team were required to participate for the team to be enrolled. Serum creatine kinase (CK) and aspartate amino transferase (AST) were measured 1-4 h before, and 4-6 h after, a game. Data related to possible risk factors for ER was collected using a face-to-face questionnaire. Acute ER was defined as a threefold or greater increase in CK from the pre-exercise level. The association between potential explanatory variables and ER was examined using bivariate logistic regression models and multivariate analysis. Agreement between owner reported muscle soreness and ER was tested using Cohen's Kappa test. Significance was set at p 0.05. Blood samples were available for 144 horses. Questionnaires were available from 110 horses, with complete data sets from 93 horses. Pre-exercise CK levels greater than double the upper limit of the normal reference range were present in 11/144 (7.6%) of horses with the AST exceeding the upper limit of the normal reference range in 10/11 of these horses. In the remaining horses, an ER event occurred in 27/133 (20.3%). Overall 38/144 (26.4%) of the horses had evidence of muscle pathology either pre-or postexercise. The majority of ER events were mild but a post-exercise CK of > 2,000 U/L and > 5,000 U/L was observed in 14/144 (9.7%) and 5/144 (3.5%) of horses, respectively. Of the variables screened at the bivariate level, previous history of ER, rider gender, the day of competition, temperament, time of the match, number of chukkas played, and state of origin influenced the odds for ER. These were included in the final logistic regression model. A previous history of tying up increased the odds of ER by 15 fold (OR 14.98, 95% CI 3.40 -90.80, P < 0.01). Horses playing after midday were nine times more likely to develop ER (OR 9.22 95% CI 2.11 -55.63; P = 0.01) compared with horses that played before midday. Agreement between owner reported muscle soreness and ER as defined in the study was poor (j = 0.323).

In conclusion, the prevalence of ER in polocrosse is higher than has been reported in other equestrian sports and warrants further investigation. The poor correlation between owner reported disease and muscle enzyme changes suggests that future studies investigating ER should use objective measurements as owner reporting is likely to be unreliable and may result in spurious results.

E-60 LAMELLAR BIOENERGETICS STUDIED USING TISSUE MICRODIALYSIS. CE Medina-Torres 1 , CC Pollitt 1 , DW Richardson 2 , C Underwood 1 , S Collins 1 , AWvan Eps 1 . 1 University of Queensland, Gatton, QLD, Australia, 2 University of Pennsylvania, Kennett Square, PA.

Limited knowledge regarding the physiology of the equine hoof lamellae is partially responsible for our incomplete understanding of the pathophysiology of laminitis. The concealed nature of the lamellae between hoof wall and third phalanx makes in vivo study of this tissue difficult. With derangement of lamellar energy metabolism (bioenergetic failure) suggested as a contributor to different forms of laminitis, knowledge of lamellar bioenergetics is key to elucidating the pathophysiology of laminitis. The aims of this study were: 1) to develop a tissue microdialysis (lD) method suitable for sampling the lamellar extracellular fluid (ECF) for prolonged periods, 2) to determine the metabolic profile of lamellar tissue by measuring basic energy metabolites in the lamellar ECF in normal horses, and 3) to compare this energy profile to that of the skin dermis.

Microdialysis probes (0.5 mm diameter, 10 mm long, 100 kDa membrane) were inserted into the dorsal lamellae of one forelimb as well as the skin dermis over the tail base in 7 normal horses. Probes were continuously perfused at a rate of 1 ll/min and microdialysate was sampled simultaneously from the lamellar and skin probes at 2 hour intervals for a 24-hour study period. Samples were analysed for glucose, lactate, pyruvate, urea and glycerol concentration using a clinical microdialysis fluid analyser. Standard indices of energy metabolism including lactate:glucose (L:G) and lactate:pyruvate (L:P) ratios were calculated. Data were analysed non-parametrically, with significance set at p < 0.05.

Fluid was not significantly different between sites. The energy metabolite concentrations were similar to previous reports in equine muscle and in tissues of other species such as humans, pigs and mice.

The measured energy metabolites were consistent and stable for 24 hours, suggesting that this technique is suitable for evaluating disturbances in energy metabolism in lamellar tissue over time. The high lamellar L:G (more than twice that of the skin dermis, and other previously studied tissues) indicates a uniquely high rate of glycolysis; however the L:P is consistent with other tissues that have a high oxidative capacity. The measurement of drug concentrations in equine digital lamellar tissue is key to evaluating pharmaceutical means of laminitis prevention. The aim of this study was to develop a minimally invasive technique for sampling lamellar interstitial fluid using ultrafiltration.

A technique for inserting ultrafiltration probes (3 loops of 8 cm long 30 kDa semipermeable membrane) in lamellar tissue was developed in vitro using 15 cadaver limbs. Subsequently, ultrafiltration probes were placed in the lamellar tissue of 6 normal horses. The horses' comfort levels were judged using a pain scoring system and pedometers attached to their antebrachii. Ultrafiltrate was collected continuously for a 4 (n = 4) or 14 day (n = 2) study period. The fluid was sampled every 12 hours and the rate of collection calculated. Biochemical analyses were performed on ultrafiltrate and plasma collected at 12 to 24 hours post implantation (day 1) and at 84 to 96 hours post implantation (day 4) to establish the concentrations of albumin, alkaline phosphatase (ALP), aspartate aminotransferase (AST), calcium, cholesterol, gamma glutamyl transferase (GGT), lactate dehydrogenase (LDH), creatinine kinase (CK), phosphate, total protein (TP), globulin, total bilirubin, chloride, carbon dioxide, creatinine, glucose, magnesium, triglycerides, urea, sodium and potassium. Lamellar tissue surrounding the probe was submitted for histology to assess probe location and local tissue reaction. Data were analysed non-parametrically with significance set at p < 0.05.

Ultrafiltration probes were placed successfully in all 6 horses using an introducer inserted at the toe. The horses showed minimal signs of discomfort with no significant changes in pain score or pedometer readings during the study period. Ultrafiltrate was collected at 55 [30-63] lL/hr (median [interquartile range]). Fluid production decreased significantly with time from day 3 onwards. There was no significant change in the constituents of the ultrafiltrate between day 1 and day 4. The concentrations of albumin, ALP, AST, GGT, LDH, CK, globulin, TP, total bilirubin, calcium, cholesterol and phosphate were all significantly lower in ultrafiltrate compared to plasma (p = 0.03). Histological sections confirmed the location of the ultrafiltration probe membrane at the dermo-epidermal junction parallel to the long axis of the primary epidermal lamellae in all 6 horses. Epithelialization was present around the probe in all horses, but was more extensive in horses in the 14 day study group.

This study demonstrates that ultrafiltration can be used to sample equine digital lamellar interstitial fluid. The reduction in ultrafiltrate collection with time was attributed to epithelial encapsulation of the probe. The biochemical analytes in the ultrafiltrate were consistent with the constituents of extracellular fluid, excluding molecules that are protein bound or have molecular weights greater than 30 kDa.The concentrations of the analytes did not vary over 4 days, indicating the probes have potential for direct measurement of lamellar metabolites and pharmaceutical substances over this period. Working with primary muscle cell cultures presents several difficulties: they are heterogeneous (containing mixed populations of fibroblasts and myoblasts) and their derivation is time consuming and unreliable; furthermore, myoblasts often senesce after a limited number of divisions. The development of a clonal, conditionally-immortalised equine myoblast cell line would overcome these disadvantages and provide a valuable in-vitro tool for the evaluation of horse muscle physiology and disease. We hypothesized that transfecting primary equine skeletal muscle cultures with a plasmid (NIT-TAg) expressing the temperature-sensitive SV40 T Antigen driven by a Tet-off responsive promoter would enable the generation of a clonally-derived, myogenic cell line that would reliably differentiate into myotubes under permissive conditions. In addition, we aimed to characterise the optimum conditions for differentiation of these immortal cells into multinucleated myotubes.

A primary, equine skeletal muscle cell-culture was transfected with an eGFP-expressing plasmid via Lipofection â and Nucleofection â to compare transfection technique efficiency. Nucleofection was then used to transfect primary cell cultures with the NIT-TAg plasmid. Cells with stably-integrated transgenes were selected by incubating at 33°C in G418 (400ug/ml). Transfected cells were clonally selected and immunolabelled for desmin to identify myogenic lines that were subsequently differentiated by incubating with doxycycline (1ug/ml) at 37°C in reduced serum media. Expression of SV40 TAg and of other proteins associated with myotube differentiation, was evaluated by immunocytochemistry. In addition, the effect of cell number, substrate and differentiation media, on differentiation was also evaluated.

Lipofection resulted in very low transfection efficiency (<1%), whilst Nucleofection resulted in transfection of up to 46% of cells, in a plasmid DNA concentration-dependent manner. Unlike un-transfected control cells, a myogenic, desmin-positive clone expressed the SV40 TAg in selected environmental conditions, with its maximum expression at 33°C and in media without doxycycline. Immortalised cells from this clone could be differentiated into multi-nucleated myotubes in permissive conditions and optimum conditions for this differentiation were established.

Nucleofection of primary skeletal muscle cultures with the NIT-TAg plasmid resulted in successful stable transfection of myogenic cells that expressed the immortalising transgene. Cell proliferation continued in permissive conditions (33°C), whilst cells retained the ability to differentiate into myotubes when incubated at 37°C with doxycycline. This conditionally-immortal equine skeletal muscle cell line will provide a valuable model for future in vitro work. Physical exercise induces remarkable changes in homeostasis and can also influence immune response. The relationship between acute phase proteins and inflammation related to exercise has been studied in human and horses. Therefore, the aim of this study was to determine serum protein profile during recovery period in healthy trained horses submitted to prolonged exercise.

For this purpose, five trained sound Arabian horses were submitted to an 80 km endurance ride with three intermediate vet cheks. Blood samples were collected before and six, 12, 24 and 48 hours after endurance race. Total protein was determined by Biuret method and serum proteinogram obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The electrophoretic fractionation was performed by unidimensional electrophoresis system on 4%-10% gradient acrylamide gels (Ethical Committee CEUA Protocol 001624/11).

Total protein concentration did not change during recovery period, with maximum increase of 13% six hours after endurance ride. Concentration of ceruloplasmin, haptoglobin and IgA decreased 74%, 44% and 35% six hours after exercise, respectively. At this time transferrin reached a peak concentration and increased 58%, then decreasing continuously to basal levels. Haptoglobin remained decreased until 24 hours and returned to pre-ride levels 48 hours after exercise. Ceruloplasmin and IgA increased by 62% and 13% twelve hours after endurance race, reaching maximum increases of 157% and 24%, respectively, after 48 hours.

An 80 km endurance ride induces changes in serum protein profile indicative of acute phase response that persisted longer than 48 hours and acute phase protein with greater increase was ceruloplasmin.

Hemophilia A is an X-linked disorder caused by a deficiency in factor VIII (FVIII) and is the most common inherited coagulation disorder in horses. Although foals have been diagnosed based on deficiency in FVIII activity, the causative gene mutations have not been identified. The objectives of this project were to determine the sequence for the FVIII gene in the horse, identify the causative mutation in a colt with hemophilia A, determine if the mutation was a spontaneous event or due to maternal inheritance, and further the understanding of the genetics of inherited coagulopathies in veterinary species PCR reactions performed on genomic DNA from a normal horse, the affected colt, and the colt's dam amplified the coding regions of the equine FVIII gene, FVIII promoter, FVIII binding site on the von Willebrand factor (vWF) gene, and genes encoding FVIII transport proteins lectin mannose-binding 1 (LMAN-1) and multiple coagulation factor deficiency 2 (MCFD2). Sequence data was compared for identification of mutations and maternal inheritance. Hepatic mRNA was isolated from the affected colt and a normal horse, reverse transcription PCR was used to create first strand cDNA. PCR reactions were performed on the cDNA across the FVIII coding sequence. Western immunoblot was on hepatic lysate from the normal horse and affected colt.

Genomic sequence for the exons and splice sites of FVIII gene, FVIII promoter, and coding regions of LMAN-1 and MCFD2 were identical in all animals. A single nucleotide polymorphism (A>C) was detected in exon 22 of the vWF FVIII binding site in the colt, leading to a change from lysine to asparagine. The mutation was also identified in samples from 3 normal horses, suggesting there was no significant effect on the encoded binding site. PCR on hepatic cDNA revealed normal product for the foal and unaffected horse from exons 2-26 of the FVIII gene. A product was not obtained across exons 1-2 in the foal. Genomic sequencing of intron 1 identified several single nucleotide polymorphisms and a three base pair deletion in the foal. The mare was heterozygous for the deletion, which was not present in the DNA from seven normal horses. cDNA sequence confirmed that portions of intron1 were transcribed in the foal across the 5' splice site. Western immunoblot revealed normal sized FVIII protein in the normal horse, but a product was not obtained for the foal.

The mutation was isolated to the non-coding region of FVIII (specifically intron 1). Genomic sequencing of intron 1 suggested maternal inheritance. Transcription of the intron across the 5' splice site suggested activation of an aberrant splice site in intron 1. It is yet to be determined if this defect is responsible for hemophilia A in other horses. Physical exercise induces remarkable changes in homeostasis and the relationship between acute phase proteins and inflammation related to exercise has been studied in human and horses. Therefore, the aim of this study was to determine changes in serum protein profile in horses immediate after prolonged exercise.

For this purpose, blood samples were obtained from 11 Arabian horses that successfully finished an endurance ride competition; four horses completed 160 km, four horses 120 km and three horses completed 80 km. Samples were collected before and immediate after competition. Total protein was determined by Biuret method and serum proteinogram obtained by unidimensional SDS-PAGE gel electrophoresis.

Total protein concentration increased 5%, 14% and 10%, respectively, after 160, 120 and 80 km endurance ride. Serum concentrations of IgA decreased 5%, 33% and 30% after 160, 120 and 80 km endurance ride, respectively and a1acid glycoprotein decreased 15%, 53% and 38%. IgA is essential to gastrointestinal tract defense against bacteria and a1-acid glycoprotein can bind to endogenous and exogenous substances, including LPS. Increases in LPS during prolonged exercise have been reported, which could explain serum concentration reduction in both acute phase proteins. Transferrin is one of the most precocious acute phase protein and increased 21% in horses that completed 160 km. Increases in oxidative metabolism after long distance exercise have also been reported and may be a cause of the 48% decrease in haptoglobin serum concentration in this horses.

Serum proteinogram allowed detection of premature changes and help to understand the systemic alterations resulting from prolonged physical effort. Ceftiofur crystalline free acid (CCFA) is labeled for the treatment of equine lower respiratory tract infections caused by susceptible strains of Streptococcus equi subspecies zooepidemicus. Current labeling for the use of CCFA in horses states that 2 IM doses must be administered 4 days apart to provide approximately 10 days of therapeutic coverage, as defined by maintaining plasma concentrations of desfuroylceftiofur acetamide (DCA; the acetamide derivative of ceftiofur and desfuroylceftiofur metabolites) above 0.2 lg/mL. There is currently no information available to help practitioners determine what would be an appropriate dosing interval for long term administration of CCFA. The objectives of the present study were to determine if weekly administration of CCFA is sufficient to maintain therapeutic plasma concentrations of DCA and to compare the pharmacokinetics of DCA after SC administration to those obtained after IM administration.

In the chronic IM administration study, seven healthy mares were administered a total of 5 doses of CCFA (days 0, 4, 11, 18, and 25) . For the SC versus IM administration study, 2 doses of CCFA were administered by the SC route to six healthy mares on days 0 and 4. Six other mares received the same dosage regimen by the IM route. Concentrations of DCA in plasma were measured using HPLC with UV detection. Visible injection site swellings were measured. Pharmacokinetic parameters and injection site swellings were compared between the 2 routes of administration using Student's t test.

For both studies, plasma concentrations of DCA remained above the therapeutic target of 0.2 lg/mL at every time point sampled. Pharmacokinetic variables obtained after SC administration of CCFA did not differ significantly from those obtained after IM administration. However, the SC route of administration resulted in significantly larger injection site swellings compared to those obtained after IM administration.

After the initial regimen of 2 doses 4 days apart, weekly administration of CCFA is sufficient to maintain therapeutic concentrations in plasma if long term administration in necessary. Despite adequate plasma concentrations and pharmacokinetic profile, the clinical utility of the SC route of administration might be limited by the significantly larger external swellings compared to those obtained after IM administration. Non-steroidal anti-inflammatory drugs (NSAIDs) are commonly prescribed for the management of lameness in horses. The beneficial effects of NSAIDs are achieved by inhibition of the cyclooxygenase (COX) enzyme, which catalyses the formation of pro-inflammatory prostanoids. Meloxicam (MEL), a COX-2 preferential agent, has been advocated for this purpose and previous studies by our group have demonstrated that the drug may be safer than less selective COX inhibitors. MEL has been demonstrated to be an effective anti-inflammatory in vitro and in vivo, however comparisons with drugs in more common usage are lacking. Our aim was to objectively assess the efficacy of meloxicam in horses following lipopolysaccharide (LPS) induced synovitis by comparison to a positive control (phenylbutazone, PBZ).

A pilot study was conducted in 4 horses to characterise an in vivo method for induction of reversible lameness by direct comparison of two previously reported methods: intrasynovial injection of autologous blood or LPS. Data from this study demonstrated that LPS was the more suitable method, yielding an obvious but transient lameness, as well as increased synovial fluid nucleated cell counts and synovial fluid protein concentrations.

Synovitis was induced in a further 8 horses by injection of 0.5 ng LPS into the middle carpal joint one hour prior to the oral administration of PBZ or MEL in a cross-over study. Response to treatment was assessed by use of a composite orthopaedic pain scale, computerised in-shoe pressure measurement system, lameness grading, stride length, joint flexion, carpal circumference, synovial fluid cytology analysis and synovial serum amyloid A quantification. Following a 'washout' or recovery of greater than 14 days, horses received the second drug after induction of synovitis in the contralateral limb. Treatment order and limb were randomly allocated. No significant difference was observed between agents in any of the parameters evaluated and both drugs substantively attenuated the lameness observed following LPS administration. Conventional antiseizure therapy for horses is outdated. Levetiracetam (LEV: Keppra â ) is a new, safe, and efficacious antiepi-leptic drug (AED) with a unique mechanism of action, but pharmacokinetic studies in horses are lacking.

Eight horses received 20 mg/kg of intravenous LEV ( IV LEV), 30 mg/kg of oral, immediate release LEV (LEV IR ), and 30 mg/kg of oral, extended release (LEV ER ) in a 3-way randomized crossover design, with > 7-days washout period. Both oral formulations were crushed to be administered. Serum samples were collected over 48 hours, and LEV concentrations determined by immunoassay.

Maximum mean AE SD serum concentrations of LEV IR and LEV ER were 50.7 AE 10.6 and 53.6 AE 15.9 lg/mL, respectively. For IV LEV, LEV IR and LEV ER, half-life and area under the curve were 6.53 AE 1.29, 6.99 AE 1.97, 7.58 AE 2 hours and 17.1 AE 3.8, 24.4 AE 4.2 and 25.3 AE 6.4 mg*min/mL, respectively. Total body clearance of IV LEV and clearance/bioavailability of LEV IR and LEV ER were 1.23 AE 0.32, 1.28 AE 0.23 and 1.28 AE 0.34 mL/min/kg, respectively. Bioavailability of LEV IR and LEV ER was 96 AE 10 and 98 AE 13%, respectively.

Oral administration of 30 mg/kg of LEV IR or LEV ER in healthy adult horses is likely to achieve therapeutic concentrations for at least 18 hours. In other species, large variations in body size produce alterations in drug clearance and elimination half-life, necessitating dose adjustments beyond weight proportionality. Despite the substantial disparity in body size between Miniature Horses and light-breed horses, there are no studies investigating appropriate dosing of any veterinary drug in Miniature Horses. This study examined whether Miniature Horses should receive a different dose rate of flunixin meglumine than that used in typical lightbreed horses. We hypothesized that the total body clearance of flunixin would be faster in Miniature Horses as compared to Quarter Horses. A standard dose of flunixin meglumine was administered intravenously to eight horses of each breed, and plasma flunixin concentrations were determined over the following 24 hours. Data for each horse were analyzed by a three-compartment model, and pharmacokinetic parameters were compared between breed groups. The total body clearance of flunixin was 0.97 AE 0.30 mLÁmin À1 Ákg À1 in Miniature Horses and 1.04 AE 0.27 mLÁmin À1 Ákg À1 in Quarter Horses. Similarly, the elimination half-life of flunixin in Miniature Horses was 3.4 AE 1.1 hr and in Quarter Horses was 3.0 AE 1.0 hr. There were no significant differences between Miniature Horses and Quarter Horses in the majority of pharmacokinetic parameters for flunixin (P > 0.05). Therefore, flunixin meglumine may be administered to miniature horses at the same dose rate as is used in light-breed horses. Whereas adjusting dose to body weight is critical when therapeutics are used in miniature horses, further dose adjustments are not indicated when flunixin meglumine is administered intravenously to this breed. A suspicion of low efficacy with an ivermectin oral paste in two breeding and sport horses was solved using an other product with the same active ingredients and the same formulation, administered at the same dose, The parasitic status improved dramatically after the second treatment. The amount of oral paste delivered increment by increment revealed a large variation going of a normal dose to less 10% of the expected dose. The reason of this variation was the presence of large air bubbles in some syringes.

We took X-Rays pictures (Gierth 200 HF; 40 kV; 0,06s) of all the preparations of oral paste for horses, contained in syringes, available on the French market. Syringes (3 by brand) were newly acquired from trusted distributors and stored during 3 days at room temperature (18-21°C, horizontally). The brand name of each speciality, the lot number and the expiry date were recorded.

X-Ray pictures of commercial brand of equine oral paste raise questions. Indeed, some syringes contain significant air bubbles (Divamectin, Noromectin, Panacur paste, Telmin), whereas others had several smaller air bubbles (Eqvalan duo, Furexel combi, Strongid, Tenivalan).Consequences of such variation are a low dosage of the active ingredient in "small" horses and ponies (less than 300 kg), increasing the risk of low efficacy and potential chemo-resistance This raises the question of market authorisation of generic products whose are being approved based on bioequivalence vs the princeps product, especially ivermectin-based paste. Differences in manufacturing processes or lack of manufacturing data might explain such differences. More consistency in manufacturing processes and relevant Quality Control should be promoted to solve such issue. The objective of this study was to compare descriptive findings on radiographic and computed tomography (CT) of the lung in foals and to evaluate the relationship between lung density and measurements of PaO 2 .

Thoracic CT immediately followed by lateral thoracic radiographs were performed on 7 sedated foals (6.9 AE 3.8 days; range 3-12 days) in sternal recumbency. Descriptive evaluation of CT and radiographic images was performed by 3 reviewers. Results were compared by consensus agreement and a standardized scoring system to objectively grade presence and severity of radiographic patterns. Quantitative analysis of CT images was used to compute lung density based on measurements of attenuation (houndsfield units; HU). Arterial blood gases were obtained immediately prior to acquisition of CT and radiographic images.

On CT all foals demonstrated evidence of atelectasis ventrally. This was most pronounced caudal to the heart and corresponded to increases in lung density. Atelectasis was not identified on radiographs. Mild interstitial changes were identified caudodorsally in 3 foals. Imaging severity scores were higher for the CT images compared to radiographs. Mean lung density was -479 AE 47.3. PaO 2 immediately prior to CT was 68.7 mmHg AE 5.5 and following CT was 67.6 mmHg AE 8.9. Density and PaO 2 were not related (p = 0.577).

Differences in sensitivity between CT images and radiographs were identified and may have implications in the evaluation of pulmonary disease in neonatal foals. Additional research is needed to evaluate differences in sensitivity between CT and radiographic images of the lung in clinical cases. Clenbuterol causes an increase in tracheal mucociliary clearance rate (TMCR) and can cause repartitioning at higher doses. The clinical response to clenbuterol has been shown to diminish after 10 to 14 days. It was hypothesized that twice daily dosing of clenbuterol for a 3 week period would lead to tachyphylaxis in regards to TMCR and repartitioning effects.

Eight adult horses were enrolled in a cross-over design, randomized, blinded prospective study. Horses received either clenbuterol (0.8 lg/kg PO q12 h) or an inert placebo for 21 days. Every 3 days, TMCR was measured using a charcoal clearance technique, and%body fat obtained sonographically (2.47 + (5.47x rump fat in cm)). There was a 7 week washout period, with each horse repeating the protocol in the opposite arm. Data were analyzed using mixed effect regression analysis.

A significant difference in TMCR was seen between treated and control groups, with the treatment group being 0.73 cm/min (p = 0.04) faster on day 6 and 0.88 cm/min (p = 0.01) faster on day 9, but returning to values not significantly different from baseline on days 12, 15, and 18. A decrease in% body fat from baseline was noted only in the clenbuterol treatment group beginning on day 6 and continuing through day 21 (p < 0.05).

In conclusion, clenbuterol at 0.8 lg/kg resulted in an increase in TMCR with tachyphylaxis evident by day 12 of the study. Evidence of repartitioning was seen in horses administered the clinically utilized low dose of 0.8 lg/kg of clenbuterol with no evidence of tachyphylaxis by 21 days of treatment Bronchospasm results in airway obstruction in horses with heaves. Atropine is the most potent bronchodilator drug currently available in horses, but is associated with side effects limiting its use. Similarly to atropine, N-Butylscopolamonium bromide (NBB) is an anticholinergic agent with bronchodilatory properties. This study compared the bronchodilating and sideeffects of atropine and NBB in horses with heaves.

Eight horses with heaves were administered atropine and NBB, using a cross-over design. Heart rate, pupil dilation, rectal palpation, lung mechanics (pulmonary resistance, elastance), and arterial blood gases were measured, before, and 10 and 30 minutes after drug administration.

One horse treated with atropine developed colic signs. Pupil dilatation was only observed with atropine. Tachycardia developed in all horses, but was more marked with atropine. Lung function improved with both drugs but elastance values had returned to baseline at 30 min with NBB. There was no difference between drugs on arterial blood gas analysis.

The study shows that NBB has bronchodilatory properties similar to those of atropine, but is of shorter duration and associated with lesser systemic side-effects. Therefore NBB should be preferred over atropine in experimental studies. The design of the study does not permit to conclude on the use of NBB in heaves in clinical practice. CPAP is a form of non-invasive respiratory support commonly advocated for the management of respiratory insufficiency in human neonates. The technique preserves spontaneous respiration, does not require intubation and may result in less lung injury than mechanical ventilation. As CPAP is less invasive than mechanical ventilation, as well as more affordable with respect to equipment and expertise, the technique may be an attractive option in veterinary settings. To date, however, this approach has received little attention in the veterinary literature and there is no experimental or clinical data on which to base optimal implementation in neonatal foals or calves.

We have used CPAP empirically in a number of foals with spontaneously occurring respiratory disease. Treated patients demonstrated increased arterial oxygen concentrations without adverse effects reported with the technique. However, objective evaluation of the response to treatment, relative to other therapeutic interventions, is often difficult in clinical patients. Two projects were therefore undertaken to more rigorously assess the value of this intervention in neonatal calves and foals with pharmacologically induced respiratory depression.

Ten calves delivered by Caesarean section were administered diazepam, xylazine and fentanyl 20 to 24 hours post-partum, and randomised to receive either nasal oxygen insufflation (n = 5) or CPAP (n = 5) as a first treatment, followed by a period breathing ambient air and then administration of the reciprocal treatment. Oxygen insufflation was associated with increased arterial oxygen and carbon dioxide partial pressures, whereas hypercapnia was reduced following CPAP. Six healthy foals (2 to 28 days post-partum) similarly received oxygen insufflation (5L/min) or CPAP with supplementary oxygen (5L/min) as a first treatment, with treatments subsequently reversed after a period breathing ambient air. After a 48 hour 'recovery', foals again received both treatments in reverse order. In this study, arterial and inspired oxygen partial pressures (PaO 2 and FiO 2 ) were higher following supplementary O 2 insufflation alone than CPAP with supplementary O 2 . Increased CO 2 pressures were observed with both interventions. Although findings in calves suggest that CPAP may improve ventilation, our preliminary results suggest that optimisation of CPAP delivery requires regulation of FiO 2 and mechanisms to reduce rebreathing. Metabolic acidosis occurs commonly in calves with diarrhea. The quantitative effect of strong electrolytes, pCO 2 , and plasma protein concentrations in determining plasma pH and can be demonstrated with the physicochemical approach. The objectives of this study were to investigate and to characterize the mechanism of acid-base disorders in a large population of calves with naturally occurring diarrhea using the physicochemical approach.

The study population consisted of two data sets (DS1; DS2). DS1 included results from calves 28 days of age from a previous study that were admitted to the Atlantic Veterinary College Teaching Hospital (AVC) with a primary complaint of diarrhea (Lofstedt et al, J Vet Intern Med 1999; 13; 81-88) . DS2 was a collection of data from medical records of calves 28 days of age, admitted to the AVC between 2000 and 2010, with final diagnosis of neonatal diarrhea. Laboratory data collected included venous blood gases, plasma electrolytes Na + , K + and Cl -, all measured using ion selective electrodes and total protein (TP) (refractometer). Strong ion difference was calculated as SID = (Na + + K + ) -(Cl -); total charge of the plasma protein as A tot = 0.343 x (TP (g/ L); strong ion gap as SIG = A --AG; where Ais net negative charge of plasma proteins and A -= A tot / (1 + 10 (7.08-pH) ), and AG is anion gap as AG = (Na + +K + ) -(Cl -+HCO 3 -). A forward stepwise logistic regression was performed to determine the quantitative association between measured pH and physicochemical variables (SID, A tot and SIG).

Acidemia resulted primarily from unmeasured anions (SIG; r 2 value DS1 = 67% and DS2 = 35%; P < 0.001) followed by strong ions (SID; r 2 value DS1 = 11% and DS2 = 29%; P < 0.001). [A tot ] and p v CO 2 had minor but significant contributions with r 2 value of DS1 = 5% and DS2 = 7% (P < 0.001), and r 2 value DS1 = 9% and DS2 = 20% (P < 0.001)), respectively.

The explanatory power of the model was 92% and 91% for DS1 and DS2, respectively.

The results of this study indicate that strong unmeasured anions were the most important contributors to the acidemia in calves with naturally acquired diarrhea, likely due to increased plasma concentrations of D-and L-lactate, and emphasize the importance of the calculation of SIG when applying the physicochemical approach to evaluate acid-base abnormalities. Data were prospectively collected from 20 calves < 30 days old, admitted for diarrhea to the Atlantic Veterinary College Teaching Hospital during 2012. An admission venous blood sample was collected for blood gas analyses using an IRMA trupoint blood gas analyzer. Plasma biochemical profile and [L-lac] were measured on aliquot of sample using a Cobas C6000 analyzer, and a Lactate Pro analyzer, respectively. The total negative charge of the plasma proteins (A -Tp ) and albumin (A -Alb ) were calculated as A -Tp = 0.343 x (TP g/L)/(1 + 10 (7.08-pH) ), and A -Alb = 0.622 x (Alb g/L)/(1 + 10 (7.08-pH) ). The five different methods to quantify [UA -] (mmol/L) were: (1) anion gap as: AG= (Na + +K + )-(Cl -+HCO 3 -); (2) Constable's strong ion gap as: SIG Tp =A -Tp -AG, or as (3) . For calculation of [UA -] other than L-lactate, [L-lact] was substracted from AG, SIG Tp , SIG alb. The correlation between AG, SIG Tp , SIG alb , SIG K , and SIG org for estimating [UA -] was determined using Pearson's correlation test. The relationship between the five methods and [L-lac] was determined using linear regression. Subsequently, [L-lac -] was removed from SIG K , SIG org and they were recalculated. ROC analysis was used to identify the [L-lac] cutoff that optimized diagnostic sensitivity and specificity for predicting mortality. Predictive value of [L-lac] for mortality was determined using a logistic regression.

All five methods had an excellent correlation for estimation of [UA -] other than [L-lac] (r 2 ranging from 0.955 to 0.987; P < 0.0001). AG, SIG Tp , SIG alb , SIG K and SIG org correlated significantly with [L-lac] (r 2 = 0.58,0,62, 0.62, 0.57, 0.61; P < 0.001). The ROC analysis identified a [L-lac] ! of 2.5 mmol/L had a sensitivity of 89% and specificity of 72% for predicting mortality. Calves with elevated [L-lac] had significantly increased odds of mortality (OR=1.3; 95% CI=1.006-1.658) Calculated AG provided an accurate estimate of plasma [UA -] and [L-lac] in neonatal calves with diarrhea, likely because there were minor alterations in the plasma protein and phosphate concentrations. The moderate correlation between AG, SIG Tp , SIG alb , SIG K and SIG org and [L-lac] indicates the presence of other unmeasured anions, likely D-Lactate. Similar to other species, increased [L-lac] was a significant predictor of mortality in diarrheic calves. Demeanour has been associated with severity of metabolic acidosis in calves with diarrhea. The objectives of this study were to evaluate the correlation between degree of metabolic acidosis, determined using the Henderson-Hasselbach (H-H) and physicochemical approaches, and alterations in the demeanor of calves, specifically their attitude, posture and suckle reflex.

The analyses were based on two data sets (DS1; DS2). DS1 was derived from calves from a previous study 28 days of age, admitted to the Atlantic Veterinary College Teaching Hospital (AVC) with a primary complaint of diarrhea (Lofstedt et al, J Vet Intern Med 1999; 13; 81-88) . DS2 was extracted from medical records of calves 28 days of age with a diagnosis of diarrhea, admitted to the AVC between 2000 and 2010. Venous blood gas results and concentration of plasma Na + , K + , Cl -, and total protein (TP) were recorded. H-H variables were [HCO 3 -], base excess (BE (ecf) ) and anion gap (AG) that was calculated as AG (Na + +K + )-(Cl -+HCO 3 -). Physicochemical variables for each calf included measured strong ion difference (SID = (Na + +K + ) -(Cl -)); total charge of the plasma protein (A tot = 0.343 x (TP (g/ L)), strong ion gap (SIG = A --AG); with Abeing the net negative charge of plasma proteins (A -= A tot / (1 + 10 (7.08-pH) )). The association between acid-base variables and posture (scored as standing (St), sternal (SR) or lateral recumbence (LR)), attitude (scored as bright (Br), depressed (De) and comatose (Co)) and suckle reflex (scored as strong (Sg), weak (We) and absent (Ab)) of calves were evaluated using a one-way ANOVA test with Bonferroni correction for pairwise comparison.

Calculated values of SIG, and AG were significantly different between calves with normal, moderate and severe alterations in posture, attitude and suckle reflex (Table) . There was no significant association between demeanour and [HCO 3 -], BE (ecf) , p v CO 2 , SID and A tot (P > 0.05). Mean values (+/-SD) of AG and SIG for calves with different stages of demeanor.

*Different letters within a row indicated an statistically significant difference (P < 0.001).

The results indicate that clinical signs of posture, attitude and suckle reflex are influenced primarily by the severity of the increase of plasma concentration of unmeasured anions likely Dand L-lactate. These results emphasize the usefulness of calculations of the SIG and AG when applying physicochemical approach to determine acid-base disorders in diarrheic calves. Otitis media/interna is a frequent disease in calves but can remain subclinical, making antemortem diagnosis challenging on the farm.

The objectives of this study were to evaluate the sensitivity and specificity of the ultrasonography; to evaluate the repeatability of the technique between two ultrasonographers; and to compare its sensitivity to the sensitivity of the neurological examination to detect otitis media in calves.

Forty calves 19-50 days of age were selected from a veal calf farm, on the basis of results of ultrasound examination performed on the farm (examiner A), in order to obtain a similar number of otitis positive and negative bullae. Diagnosis from a neurological examination (NE) and from ultrasonography (US), performed a second time by examiner A (A') and by another examiner (B), was recorded for both tympanic bullae, as either negative, suspicious or positive. The calves were euthanized and submitted for necropsy, and histopathologic diagnosis was recorded. Sensitivity and specificity of the neurological examination and of ultrasound technique for A, A' and B were estimated using histologic result as the gold standard, after adjustment for sampling design. Kappa analysis was performed to evaluate the agreement between examiners and within the same examiner (A-A'). Sensitivities were compared using one-tailed Ζ test for two proportions.

Based upon histopathology, 45 bullae were affected with otitis media and 35 bullae were normal. Agreement between A-A', A-B and A'-B yielded к values of 0.46, 0.51 and 0.49, respectively. Sensitivity and specificity of the ultrasound imaging technique are acceptable for diagnosis of otitis media in calves and sensitivity of examiner B is superior to that of neurological examination. Moderate agreement suggests that the technique is repeatable. The purpose of this study was to genomically characterize a representative isolate of Staphylococcus chromogenes from a subclinical case of bovine mastitis. The second objective was to use this reference genome as a template to screen for potential virulence genes within other S. chromogenes isolates. Whole genome sequencing of a representative S. chromogenes isolate was performed using 454 sequencing. Analysis of the draft sequence fragments and identification of repetitive sequences was done using computer software available from the Sanger Institute. Polymerase chain reaction (PCR) and Sanger sequencing were used to link adjacent contigs into larger scaffolds. The genes within the reference genome were annotated. Polymerase chain reaction primers were designed to detect potential virulence factors using the reference genome. Banked S. chromogenes isolates (n = 23) were screened for potential virulence genes using traditional PCR methods.

To date, this research has focused on compiling a completed genomic sequence of S. chromogenes. A preliminary effort to identify virulence genes has been completed using NCBI analysis. This method has identified several genes of interest including a gene related to enolase, a cold shock protein, femA, and a zinc metalloproteinase. These genes were present within all screened isolates (23/23) suggesting that differences in pathogenicity between strains of S. chormogenes are likely not attributable to differences in the presence or absence of these particular genes.

Further examination of the genome and identification of differences in the presence or absence of other putative virulence factor genes between strain-types that cause different degrees of mammary inflammation will help improve our understanding of S. chromogenes mastitis. Otitis media/interna in calves is a disease with high morbidity and significant economic impacts. The benefit of cerebrospinal fluid (CSF) examination to assist in the diagnosis of this disease remains unknown.

We hypothesized that CSF analysis would show an increased mean total nucleated cell count (TNCC) in calves with confirmed otitis media/interna versus healthy calves, and that mean TNCC would differ between lumbar and cervical punctures within each group.

Forty calves were included in this study and originally selected based on neurological and ultrasound findings for suspicion or absence of signs of otitis media/interna. Immediately following euthanasia, a lumbar and a cervical CSF puncture were performed on each calf. A blinded TNCC and a cell differential (2x400 lL cytospins) were determined for each collection site. Diseased (n = 29) or healthy (n = 11) status was diagnosed following necropsy using a blinded histopathology grade assignment, which served as the gold standard technique.

Using a mixed linear model, the mean TNCC was significantly higher in calves with otitis media compared to healthy calves (p = 0.04). There was no significant difference in the mean RBC count between these two groups (p = 0.15). A paired T test showed no significant differences for TNCC and RBC counts between lumbar and cervical punctures within the otitis media/ interna or healthy groups (p = 0.93 and p = 0.15, p = 0.21 and p = 0.39, respectively).

These results confirm that CSF analysis may be a useful ancillary test in the investigation of otitis media/interna in calves. CSF collection at either the lumbar or cervical site show similar results. Pregnancy toxemia is a common metabolic disease that affects small ruminants in the last month of gestation, and has previously been associated with high mortality rates in dairy goats 1 . Prognostic factors have not been previously elucidated. The objective of this retrospective study was to describe the signalment, history, clinical and clinicopathologic findings, treatment, and survival rates of does with pregnancy toxemia and to determine prognostic indicators associated with doe and fetal survival.

Medical records of 56 does diagnosed with pregnancy toxemia over a 10-year period were reviewed. Information on previously listed parameters and outcome of both does and kids was evaluated. Statistical analysis included.

Boer goats were overrepresented (52/56, 92.8%, P???; waiting to get the data comparing Boer vs. other breed admission to OSU). Median age at presentation was 2.9 years (mean, 2.7 years). Parity was recorded in 17/56; of these, 58.8% were primiparous. Median duration of signs prior to presentation was 2.75 days (mean, 4.3 days). Time to expected kidding ranged from 1-30 days. Median body condition score was 2.5/5 (mean, 2.7). Complete blood count/biochemistry profile at admission was available for 46.4% of does (26/56). All does were neutrophilic. Hyperglycemia (80%, 32/40), hypoalbuminemia (77.7%), and hypocalcemia (74%) were common; hypoglycemia was present at admission in only 5% (2/40). Treatments included oral propylene glycol (11/56), dextrose-containing intravenous fluids (53/56), insulin (28/56), induction of labor (19/56), and caesarean section (7/56). Dystocia was reported in 58% and postpartum uterine pathology (RFM, metritis) in 36% of does that survived to kidding (6/56 did not survive to kidding by my count??). Thirty-nine (69.6%) does were discharged from the hospital following diagnosis and treatment. Number of fetuses/kids was recorded as twins (20%), triplets (67.3%), or quadruplets (12.7%). High BUN, low potassium, low bicarbonate, and low pH at admission were associated with higher dam mortality (Dr. Taylor: can we put a number on the bicarb level below which there was little/no survival?); does with increased BUN were 8.25 times more likely to die (what about OR for bicarb/pH?). Insulin administration (P < 0.008) and caesarean section were associated with lower doe survival rates. Does that survived to kidding were more likely to die if fetal mortality was high and more likely to survive to discharge if fetal survival was also high (P < 0.007) (there has to be a better way to say this….). Hyperglycemia, normocalcemia, and normophosphatemia in does at admission were correlated with higher survival rates in kids while low bicarbonate was associated with increased kid mortality. Twins had a higher survival rate than quadruplets. Overall mortality was higher in kids delivered via C-section.

We conclude that in this population of does, BCS at admission is a poor predictor of PT, hyperglycemia is common, and high BUN and low bicarbonate are poor prognostic indicators for dam survival. We also conclude that does with triplets or quadruplets are more likely to develop PT (assuming we can make this comparison based off info from the ABGA), and that kid survival rates are higher for twins and kids delivered vaginally. Vitamin E concentrations in alpaca serum vary greatly between diagnostic laboratories. In humans and cattle, vitamin E concentrations have been altered due to sample handling, cholesterol levels, and hemolysis of the samples. The purpose of this study was to determine clinically relevant variation between vitamin E concentrations with different serum handling techniques. The effects of hemolysis and cholesterol levels were also evaluated.

Blood was collected from 2 apparently healthy male alpacas. Whole blood samples were processed under conditions of exposure to fluorescent room light, contact with the rubber tube stopper, contact duration, and temperature before serum separated. The effects of hemolysis on vitamin E analysis was evaluated by the addition of known quantities of red blood cells to prepared serum samples which were then subjected to 3 freeze-thaw cycles. The serum was then evaluated for vitamin E and hemolysis index. Cholesterol concentrations were measured for all serum samples.

Vitamin E concentrations variations due to tube position were found to be statistically significant, however, no other processing condition was significant. A negative correlation was seen between hemolysis and vitamin E concentrations as expected from other species models. Although no correlation was seen between cholesterol and vitamin E due to the single collection time, preliminary data from ongoing research has proven otherwise. In conclusion, decreasing hemolysis during sample collection and upright storage prior to analysis will obtain the most accurate vitamin E evaluation. Elevated levels of cholesterol should also be taken into consideration when evaluating vitamin E concentrations in individual animals. Arsenic toxicosis is uncommon in cattle and successful treatment is rarely reported. This analysis reviews all cases of acute arsenic toxicosis reported in the literature and describes cases from Purdue University that reflect a favorable outcome. Clinical presentation of the disease, treatments and variables associated with survival are described.

One hundred and fifty six animals from 16 outbreaks were included in the study. The median age at diagnosis was 18 months. In 34% of cases, arsenic was ingested and in 65%, arsenic toxicosis occurred through transcutaneous absorption.

The most common clinical signs were sudden death (68%), diarrhea (33%), ataxia (29%), dehydration (22%) and respiratory distress (4%). The most common clinicopathologic abnormalities included azotemia (100%), hematuria (100%), increased liver enzyme activity (86%) and increased hematocrit (60%). One percent survived and the survival time for nonsurvivors ranged from 20 hours to 21 days. None of the clinical signs or clinicopathologic data was associated with survival.

Treatment was attempted in 24% of cases and was not associated with survival (P = 0.055); however, administration of an antidote and administration of fluids were associated with a better outcome (P = 0.036 and P = 0.009, respectively). In the animals presented to Purdue University, treatment with intravenous fluids and sodium thiosulfate resulted in decreased blood arsenic in all animals (P = 0.009) and a survival rate of 50%.

Acute arsenic toxicosis has a poor prognosis but this analysis suggests that survival of severe arsenic toxicosis is possible if aggressive fluid therapy and antidotes are administered. Regional intravenous perfusion (RIVP) of local anesthetics into the distal limb is often utilized to facilitate examination or treatment of severe bovine digital disorders. There is some evidence reported from human and equine patients of dissemination of infection subsequent to RIVP. Many cattle undergoing RIVP have infectious causes of lameness and many are not on antimicrobial therapy prior to RIVP. Hence there may be risk of mobilizing bacteria into the pedal circulation and subsequent localization distant to the original site of infection. The occurrence of pedal bacteremia following RIVP of 2% lidocaine was thus investigated in this study.

We hypothesized that cattle with deep digital sepsis (DDS) would become bacteremic in the pedal circulation following RIVP with 2% lidocaine, and that the most commonly obtained bacterial isolates would be Gram-negative anaerobes. To test this hypothesis, blood cultures were aseptically obtained from the digital circulation in ten distal limbs from adult cattle diagnosed with DDS that had not been subjected to antimicrobial therapy within the previous 48 hours, and from the digital circulation of ten healthy adult control cattle with no evidence of lameness or pedal lesions. Aseptic blood samples were acquired from the dorsal common digital vein of the affected limb in clinical cases of DDS immediately following tourniquet placement. Aseptic RIVP with 20 mL of 2% lidocaine was performed followed by examination and/or debridement of the digital lesions. A second aseptic blood sample was obtained 34-58 minutes following initial tourniquet placement, immediately prior to tourniquet release. Blood samples were acquired from the distal limbs of control animals following the same protocol. Both aerobic and anaerobic cultures were performed on all blood samples obtained. Duration of tourniquet application was recorded and clinical cases and control cases were time-matched with respect to length of tourniquet time.

One clinical case of DDS (1/10, 10%) was blood culture positive prior to RIVP. This cow and four additional clinical cases (5/10, 50%) were blood culture positive following RIVP and examination/debridement. All blood samples obtained from control animals were culture negative (0/10). Of the eight bacterial isolates obtained, five were Gram-positive facultative anaerobes. A two-sided Fisher's exact test was performed to compare prevalence of pedal bacteremia in DDS cases compared to controls. Cases of DDS were significantly (p = 0.016) more likely to suffer pedal bacteremia than were controls.

Bacteremia may be present in the pedal circulation following common treatment modalities for deep digital sepsis in cattle. Systemic antibiotics may be warranted prior to or in combination with regional perfusion of the digit in these patients. The purpose of this study was to describe the progression of lung consolidation using portable ultrasonography after experimental infection with Mannheimia haemolytica. Holstein bull calves, 4 months of age, were randomly assigned to either Infected (n = 6) or Control (n = 5) groups. At time = 0 h (baseline), each animal was assessed for lethargy, fever, nasal/ocular discharge, cough, and droopy ears. The lung at intercostal spaces 1 -10 of both hemithoraces was evaluated using a linear rectal transducer (6.2 MHz; depth 9 cm). Animals were then experimentally challenged with 25 ml of 10^9 cfu/mL of M. haemolytica at log phase (Infected) or sham challenged with 25 ml of sterile saline (Control) endoscopically delivered into the distal trachea. Animals were reassessed by clinical and ultrasonographic (US) exams at 2 h, 6 h, 12 h, 24 h, 48 h, 72 h, 96 h, and 120 h post-challenge. Animals were euthanized after the t = 120 h assessment and post-mortem examinations (PM) were performed.

Sonograms were normal in 4/5 Control and 6/6 Infected animals prior to challenge. New consolidation was noted 2 h postchallenge in 1/5 Control and 5/6 Infected animals (P = 0.08). Median time to severe consolidation (consolidated area > 3 cm x 1 cm) was 6 h (IQR 6-12 h) and time to maximum lesion within an intercostal space was 24 h (IQR 12-48 h). The maximum size of lesion was 18 cm (median; IQR 12-18 cm). Moderate agreement existed between US and PM diagnosis of consolidation (Kappa 0.6; CI 0.4-1). Results suggest lung consolidation occurs soon after infection, progresses rapidly, and can be diagnosed ante-mortem by ultrasonography on farm. The purpose of this study was to evaluate determination of haptoglobin in exhaled breath condensate (EBC) as a noninvasive tool in early diagnosis of bovine respiratory disease (BRD). A commercially available portable human device (Rtube â , Respiratory Research Inc, TX) was adapted for use in calves.

Holstein bull calves, 4-5 months of age, were randomly assigned to either Infected (n = 6) or Control (n = 5) groups. The animals were experimentally challenged with 25 ml of either sterile saline (Control) or 10^9 cfu/mL Mannheimia haemolytica (Infected) that was instilled into the distal trachea endoscopically. EBC was collected at time points t = -72 h, -48 h, -24 h, 0 h, 6 h, 12 h, 24 h, 36 h, 48 h, 60 h, 72 h, 84 h, 96 h, 108 h, and 120 h post-challenge. EBC samples were taken applying the device to the nose of the calves for 5 minutes. At each sampling period, blood for serum haptoglobin was also collected.

The collection device for obtaining EBC samples was well tolerated. The amount of exhaled breath condensate obtained over 5 minutes of collection varied from 0.1-1.0 ml and was dependent on the individual breathing pattern of the animal. EBC samples were evaluated using a commercial ELISA (Immunology Consultants Laboratory, OR) having a lower limit of detection of 0.1 ng/ml; however no haptoglobin was identified in the samples analyzed.

Analysis of haptoglobin in EBC of calves was not shown to be an indicator of lung inflammation, although determination of other biomarkers in EBC warrants continued research. Pro-inflammatory cytokines such as IL-1ß and TNF-a appear to play important roles in autoinflammation associated with amyotrophic lateral sclerosis (ALS), but underlying cellular mechanisms related to this process remain to be fully defined. Canine Degenerative myelopathy (DM) is a spontaneous large animal model of familial ALS of man and represents a valuable tool through which to evaluate the role of these pro-inflammatory cytokines in the neurodegenerative process. Here we quantify the expression of IL-1ß and TNF-a in conjunction with hsp70 (another potent stimulator of the innate immune response) in cerebrospinal fluid (CSF) and spinal cord sections from control dogs and dogs with histopathologically confirmed DM. Concentrations of hsp70, IL-1ß and TNF-a in CSF were determined using high sensitivity ELISA, and protein expression in various regions of the spinal cord was determined by immunohistochemical staining (IHC) and quantified using Aperio Scanscope (Aperio Technologies, Vista, CA). Concentrations of all three proteins were below the limit of assay detection in CSF for both normal (n = 10) and DM affected dogs (n = 10). Tissue analysis of IHC staining revealed enhanced hsp70 expression in ependymal cells of dogs with DM compared to controls (p = 0.003). IL-1ß and TNF-a staining was reduced in white matter of DM affected dogs, likely artifactual due to negative staining created by the pathology associated with DM. Based on these findings, hsp70 could serve as an important biomarker or trigger for sustained inflammation in DM and ALS and further research in this area is warranted. Canine degenerative myelopathy (DM) initially manifests as spastic upper motor neuron paraparesis with general proprioceptive ataxia (stage I) that progresses to paraplegia (stage II). Owners often elect euthanasia when their dog becomes nonam-bulatory paraparesis. When euthanasia is delayed, weakness spreads to the thoracic limbs (stage III) and lower motor neuron signs emerge as flaccid quadriplegia, widespread muscle atrophy and dysphagia (stage IV). No studies have been reported yet to describe the natural disease progression pattern and rate in various breeds. The objective of this study is to estimate the median duration of disease progression from stage I through IV.

Medical records of 151 dogs with histologic diagnosis of DM and the result of SOD1 E40K missense mutation were retrospectively reviewed. Dogs with complete information for signalment and clinical history including the apparent time of the onset of disease and disease stage advancement until euthanasia were included in the analysis. Other subjective characteristics recorded included breed, presence of urinary or fecal incontinence and dysphonia. The median times to progression for each stage plus the associated 95% confidence intervals were calculated. The associations between breed, age at onset, presence of urinary or fecal incontinence and the apparent duration of clinical stage advancement were evaluated using univariate Cox regression models. Out of 151 DM-affected dogs, 106 meet the inclusion criteria. Breed, age at onset, and presence of urinary or fecal incontinence was not significantly associated with the apparent duration of progression of disease. Eight dogs developed dysphonia in stage II. This study showed rate of progression dogs with DM. Urinary/fecal incontinence and dysphonia were further described in the clinical spectrum. Therapeutic interventions should strive to significantly prolong the duration of progression beyond the estimates reported in this study. Canine degenerative myelopathy (DM) is a progressive, fatal neurodegenerative condition that affects the spinal cord and brainstem of older dogs that has been proposed as a spontaneous large animal model for the human condition amyotrophic lateral sclerosis (ALS) which is similarly progressive and fatal. To date no effective therapeutic intervention has been identified capable of treating the condition. Stem cell therapy with mesenchymal stem cells has been proposed as a means of replacing defective neurons in ALS and consequently could prove a useful therapy for dogs with DM. This study was performed with the aim of assessing the safety of direct implantation of mesenchymal stem cells into the spinal cord of dogs affected with DM as a prelude to further clinical trials.

Two dogs in the advanced stages of DM were enrolled in this study: one 12 year old male-neuter German-shepherd cross, and one 9 year old female-neuter boxer. Clinical and neurological examinations were performed and both dogs graded according to the 15 point modified BBB scale proposed by Olby. Prior to surgery dogs were started on cyclosporine at 6 mg/kg BID. Dorsal laminectomies were performed to expose the T11, T13 and L2 spinal cord segments along with durotomies at each level. Allogenic mesenchymal stem cells, previously harvested from fat grafts of donor dogs were used for injection. A total of 10 6 cells in 10 x 10 ll aliquots was injected on midline into the spinal cord using a Hamilton syringe at a depth of approximately 4-6 mm over the 3 sites. Following surgery treatment with cyclo-sporine was continued for a further 14 days before discontinuation. Hemograms and biochemistry panels were performed on day 1, 2, 3, 5 and 7 following surgery along with repeat level of pain scoring and neurological and clinical examinations. Following discharge at 7 days post surgery, repeat examinations were performed at 2 weeks and then monthly to assess neurological function.

Both dogs were paraparetic and nonambulatory with good voluntary motor function in both hindlimbs prior to surgery. No abnormalities in clinical examinations or hemograms were noted post-operatively. In one dog slight deterioration was noted following stem cell injection with a decrease in motor function in one limb. In the other dog no change in neurological function was noted. In the first dog preoperative neurological status returned to normal within 2 weeks of surgery. Over the following 6 months no significant clinical abnormalities, alloydnia or other negative side effects were noted. No significant improvement was noted in either dog.

This study showed that direct injection of mesenchymal stem cells can be performed into the spinal cord of dogs with DM with relative safety. Further studies are needed to determine whether these are the most appropriate stem cells to implant into dogs with DM, and to determine the most appropriate site for delivery.

Carolina State University, Raleigh, NC.

Brain herniation is a relatively common finding on magnetic resonance imaging (MRI) secondary to intracranial pathologies, yet remains poorly described. We retrospectively identified dogs and cats with evidence of brain herniation on MRI and described clinical and imaging findings. Measurements were performed on mid-sagittal T2-weighted images and compared to a cohort of dogs and cats with normal brains. One hundred and twenty-one cases were identified including 30 cats and 91 dogs. Fifty-four had combined caudal transtentorial and foramen magnum herniations, 35 had caudal transtentorial herniation alone, and 32 had foramen magnum herniation alone, 15 of which were secondary to caudal occipital malformation with or without concurrent intracranial pathology. The location of primary pathology was cranial fossa (87), caudal fossa (10), and multifocal or diffuse (24). Signs attributable to brainstem compression were identified in 35 cases and suspected in 34. Fiftyseven cases were diagnosed with confirmed neoplasia, 20 with suspected neoplasia, 11 with confirmed inflammatory disease, 13 with suspected neoplasia or inflammatory disease, and 20 with other conditions. Survival ranged from 1 day to greater than 1 year with euthanasia or death occurring at the time of diagnosis in 36 cases. Morphometric data showed caudal displacement of the rostral cerebellum and cerebellar vermis with caudal transtentorial and foramen magnum herniation, respectively, compared to normals. We conclude brain herniation results from a wide range of pathologies located most commonly in the cranial fossa. Results confirm brain herniation on MRI may not always be accompanied by signs directly attributable to herniation. Intramedullary neoplasms of the canine spinal cord are infrequently reported. The objective of the study was to describe incidence, phenotypes, radiographic features when available, and clinical behavior of canine intramedullary spinal cord tumors. The study consists of a retrospective case series of 53 dogs with histologically confirmed intramedullary spinal cord tumors.

Intramedullary tumors comprised 16% (53/331) of all histologically confirmed tumors of the spinal cord over the study period. Primary tumors were diagnosed in 66% (35/53) of cases, with neuroepithelial-origin tumors comprising 51% (18/35) of all primary neoplasms. Intraspinal metastases of transitional cell carcinoma and hemangiosarcoma accounted for 66% (6/18 each) of all secondary tumors. Primary tumors were significantly more likely to affect younger dogs. Dogs with intramedullary metastases were most commonly presented for primary myelopathic signs (8/18, 44%). The majority of all tumors (52.8%) occurred in the T3-L3 spinal cord segments. All dogs having a cervical neurolocalization had primary tumors. Metastatic lesions had a shorter duration of signs before presentation but there was no difference in survival time between dogs with primary vs. secondary tumors. Radiographic studies were variable but indicate that MRI is the imaging modality of choice for intramedullary neoplasia.

Intramedullary spinal cord is an uncommon location for spinal cord tumors. Within this group primary tumors are more common and tend to occur in a younger population of dogs and in the cervical spine. Intramedullary metastases occur in older dogs, are rarely asymptomatic and neurologic dysfunction is a common chief complaint. Primary intramedullary tumors may have a protracted clinical course compared to intramedullary metastatic disease. Defining specific abnormalities in molecular signaling pathways in canine glioma is essential to both the understanding of basic gliomagenesis in this species, and to help define appropriate candidates for targeted therapeutic strategies. We utilized western blotting with archival canine tumor tissue and canine glioma cell lines to identify appropriate canine tissue reactive antibodies and to subsequently define protein expression and activation status of key regulatory pathways previously implicated in gliomagenesis and progression in human tumors.

Pathways investigated (specific proteins in parenthesis) included the p53 pathway (p53/MDM2/p21); the p16INK4a/Rb pathway (Rb); the PI3Kinase/AKT pathway (AKT/phospho-AKT/PTEN/MTOR/phospho-mTOR); the Ras/Raf/MAPKinase pathway (MAPK/phospho-MAPK). Protein was extracted from archival glioma tissues or from glioma cell lines (J3T-Bg, G06A, SDT-3G) +/treated with CPT-11 or temozolamide, and assayed by standard western blotting. Protein levels were assessed relative to normal cortex.

All antibodies used recognized proteins of the appropriate molecular weight. Chemotherapeutic challenge of glioma cell lines typically resulted in elevation of p53, p21 and MDM2 together with dephosphorylation of AKT and retinoblastoma. In tumor specimens, elevation of MDM2 and p21 was present predominantly in GBM. Retinoblastoma was elevated in high grade gliomas, whereas phospho-AKT was elevated more generally. PTEN protein was present in all tumors and normal cortex. Phospho-MAPK was elevated predominantly in low grade astrocytomas.

These preliminary data suggest that canine gliomas may have some similar molecular pathway alterations to human tumors, however further definition of the pathways in greater detail is warranted to inform appropriate choice of targeted therapies and interpretation of future therapeutic trials. An X-linked muscular dystrophy with truncated dystrophin has been described by Jones in the Japanese Spitz breed. The aim of this study was to identify the causative mutation and thereby develop a specific test to identify affected cases early in the course of disease and also identify carrier females.

DNA and RNA were routinely extracted from the muscle of an affected male Japanese Spitz. Using a combination of Duchenne muscular dystrophy (DMD) gene expression analysis and genome walking experiments, an X chromosome inversion that interrupts the DMD and retinitis pigmentosa GTPase regulator (RPGR) genes was identified. DNA was then isolated from saliva (Oragene â ANIMAL DNA collection kits) of a number of Japanese Spitz individuals, including unaffected individuals of both sexes (n = 14), and an obligate female (n = 1). Polymerase chain reaction amplification, using primers specific for the DMD/RPGR inversion breakpoints, identified the mutant X chromosome in all affected individuals and in the obligate carrier.

It is concluded that the identified inversion is the causative mutation for X-linked muscular dystrophy in the Japanese Spitz breed. We now have a simple, fast and accurate test for genotyping this locus. Identification of carrier animals and selected breeding should help to eliminate the mutation from the breed. The purpose of this study was to investigate retrospectively the efficacy, safety and long-term effects of combination therapy (prednisone plus cytosine arabinoside [P&CA]) compared with sole prednisone therapy in dogs with presumptive antemortem diagnosis of meningoencephalomyelitis of unknown etiology (MUE).

Medical records of dogs diagnosed with presumptive MUE treated with prednisone therapy alone or with P&CA between October 2003 and November 2012 were reviewed. Selection criteria required that all dogs had neurologic examination, documentation of clinical progression, results of blood work, cerebrospinal fluid (CSF) analysis and imaging (MRI or CT) consistent with MUE. Laboratory abnormalities, side effects, clinical and CSF responses to treatment were evaluated. Histopathological diagnosis was performed when feasible.

Twenty seven cases were included in the study. There were 16 females (15 entire, 1 neutered) and 11 entire males. Ages ranged from 9 month to 10 years. Overrepresented breeds included Bichon Maltese (n = 4, 15%) and French Bulldog (n = 3, 11%). Of the 16 cases in the P&CA group 11 dogs had evidence of focal neurological signs, 3 multifocal and 2 ocular. Of the 11 cases in the prednisone group 8 dogs had focal neurological signs, 2 multifocal and 1 ocular. In the P&CA group 69% had a good response to treatment, 25% had partial response and 6% had a poor response. Prednisone was reduced in dosage or discontinued in 44% and 13% of cases, respectively. In the prednisone group 45.4% had a good response to treatment, 36.4% a partial response and 18.2% a poor response. Within this group prednisone was reduced in dosage or discontinued in 30% and 10%, respectively. Age, gender and neuroanatomical localization had no effect (P > 0.25 in all cases) on survival time. Animals presenting seizures had shorter survival times (P = 0.05) than those not seizuring. Treatment was the only independent factor significantly affecting survival time. Animals treated with P&CA had longer survival times (P = 0.018) than those treated only with prednisone. Median survival time for animals treated with prednisone was 51 weeks. The median survival time could not be determined for animals treated with P&CA because more than 50% of the animals were alive when the study period concluded but mean survival time was 156 weeks. No signs of myelosuppression associated with CA were observed. Necropsy allowed confirmation of granulomatous meningoencephalomyelitis in 3 dogs within the prednisone group. In 9 cases (6 from the prednisone group and 3 from the P&CA) necropsy was not performed.

These data suggest that combination treatment of P&CA is a safe therapy and increases survival time compared to treatment with prednisone alone in presumptive MUE cases. The aim of the study was to prove the hypothesis that chemokine CCL19 is involved in the pathogenesis of mononuclear cell migration into the subarachnoidal space of dogs with inflammatory and non-inflammatory CNS diseases and may serve as a valuable biomarker for the progression of the diseases.

This retrospective study analyzed a total of 141 dogs. Experiments were performed on CSF and serum samples of dogs affected with steroid-responsive meningitis-arteritis (SRMA) during the acute phase (n = 25) as well as during treatment (n = 26). Dogs with SRMA were compared to dogs with presumed meningoencephalomyelitis of unkown origin (MUO) receiving no therapy (n = 16) and with patients receiving prednisolone therapy (n = 11). A control group consisted of dogs with Idiopathic Epilepsy (IE) (n = 21) and of healthy dogs (n = 6). Additionally, dogs with intervertebral disc disease (IVDD) of varying severity (n = 36) were analyzed. Concentrations of CCL19 were determined by sandwich ELISA. Migration assays were performed on seven selected CSF samples using a disposable 96-well chemotaxis chamber (SRMA n = 3; MUO n = 2; IVDD n = 2).

CCL19 was detectable in CSF samples of all dogs. Dogs with untreated SRMA/MUO displayed pronounced elevations compared to negative controls and patients receiving treatment. CSF cell count of untreated SRMA/MUO patients was significantly positively correlated with CCL19 CSF concentration. IVDD patients also had elevated CSF CCL19 concentration compared to negative controls, but values were considerably lower than in inflammatory CNS diseases. Selected CSF samples displayed chemotactic activity for mononuclear cells in the migration assay.

The hypothesis was confirmed that CCL19 is involved in the pathogenesis of SRMA/MUO, possibly perpetuating an autoimmune reaction. The marked chemotactic activity for mononuclear cells supports a conclusion that this chemokine could be associated with the migration of mononuclear cells into the subarachnoidal space of dogs. The elevation of CSF CCL19 in IVDD let suggest that it also plays a role in the secondary wave in spinal cord injuries (SCI). CSF CCL19 concentration may serve as a prognostic indicator in SCI and be a precious biomarker for developing new treatment schemes and verifying the success of treatment. The aim of the study was to confirm that canine distemper virus (CDV) infection of ex vivo isolated monocytes leads to a substantial generation of reactive oxygen species (ROS) which contributes to the pathogenesis of demyelination in the acute phase of the CDV infection. During the acute phase of the CDV infection peripheral blood monocytes migrate to the central nervous system. Microglia and recruited monocytes/macrophages are capable to synthesize ROS that are cytotoxic particularly to oligodendrocytes which might be one underlying mechanism leading to demyelination.

Monocytes were isolated with density gradient centrifugation by using 10 ml of whole blood from 10 healthy Beagles. Monocytes were infected with either a CDV-vaccination strain (Onderstepoort, Ond) or a virulent CDV-strain (R252). Non-infected monocytes were used as a negative control (Ctr). The expression of CD14 and the ROS generation were measured directly and 3 hours post infection (p.i.) by flow cytometry.

Both CDV-strains were capable to infect the isolated monocytes but the percentages and the intensities of infection were lower with the Ond-(mean value, mv = 32.5%) compared to the R252 strain (mv = 61.2%). The expression of CD14 was elevated by approximately 20% in the infected monocytes (Ond mv = 71.6%, R252 mv = 68.9%) compared to the Ctr (mv = 50.3%). Also the expression intensity of CD14 was up-regulated in CDVinfected monocytes compared to the Ctr (3 hours p.i., Ond: 4.9-, R252: 4.7fold). Interestingly, CDV-infected monocytes synthesized less ROS than the non-infected Ctr. Merely two dogs with mildly decreased leukcocyte cell counts showed an increased ROS generation in response to CDV infection. Dogs with the highest ROS generation also showed the highest CDV infection and the lowest total leukocyte and monocyte cell counts.

CDV had the potential to infect a high proportion of the isolated monocytes and depending on their immunological state, the dogs were able to generate ROS to a differing degree. In most of the dogs the ROS generation was decreased in CDVinfected monocytes. The differences in the reaction profile of the monocytes in healthy dogs can be due to genetic influences on the immune system and might answer the question, why only some dogs develop encephalitis following systemic CDV infection. The hypothesis was proven that the CDV infection of monocytes stimulates their ROS generation. The degree of the ROS production is however smaller in CDV-infected monocytes compared to their potential becoming evident in the non-infected monocytes. Canine joint-associated cervical spondylomyelopathy (JACSM) is a multifactorial disease that results in various degrees of myelopathic signs. The purpose of this study was to review the signalment, neurological examination, magnetic resonance imaging findings and outcome in dogs treated medically or surgically for JACSM.

Medical records were retrospectively reviewed from 27 cases diagnosed with JACSM between 2000 and 2012. The median age was 2 years. There were 15 Great Danes, 3 Mastiffs, 3 Newfoundlands, and 6 other large breed dogs. The neurological signs were graded from 0 (normal) to 5 (tetraplegic). Medically treated dogs (n = 7) had a median neurological grade of 2, and surgically treated dogs (n = 20) had a median neurological grade of 3. Magnetic resonance imaging showed lateral compression in 22 dogs and dorsal compression in 5 dogs. There was no correlation between severity of compression on MRI and neurologic grade. The median survival time (MST) of medically treated dogs was 53 months, and of surgically treated dogs was 46 months. The neurological signs in medically treated dogs progressed and 5 dogs were euthanized due to JACSM. Thirteen (65%) of surgically treated dogs were neurologically worse two days after surgery (P < 0.001). Fifteen of 18 dogs (83%) were improved at 4-8 weeks postoperatively and had a good to excellent long-term outcome.

In conclusion surgical treatment results in neurologic improvement and carries a good long-term prognosis in dogs with JACSM. However, medical management may be an option in mildly affected dogs. Morphometric studies of the cervical spine have been performed in large breed dogs with disc-associated cervical spondy-lomyelopathy (CSM), and have greatly assisted in the understanding of the disease pathogenesis. Such studies are lacking for giant breed dogs with the osseous form of CSM. The purpose of this study was to prospectively characterize and compare the morphometric features of the cervical spine of GDs with and without clinical signs of CSM using MRI.

Thirty client-owned GDs were prospectively enrolled, 15 clinically normal and 15 CSM-affected GDs. Inclusion criteria included a normal neurologic examination for the normal GDs, and clinical signs compatible with CSM and diagnostic confirmation using MRI for the CSM group. All dogs underwent MRI of the cervical spine (from C2-3 to T1-2 intervertebral spaces) with a 3.0 T MRI. Pulse sequences and planes acquired included: T1W (sagittal and transverse pre-contrast, sagittal post-contrast), T2W (sagittal, transverse, dorsal), and T2W sagittal images posttraction. Cranial and caudal articular facets areas were measured on T1W transverse images. Total facet area was calculated. On transverse T2W images, vertebral canal and spinal cord height, width, and area were obtained at the caudal aspect of the cranial vertebral body, at the cranial aspect of the caudal vertebral body and at the center of the intervertebral space from C2-3 to T1-2 intervertebral spaces. Right and left middle foraminal heights were also measured. The same investigator performed all measurements. Intraobserver agreement was calculated by repeating all measurements 3 times in 4 dogs. All measurements were compared for each intervertebral space between groups by use of a random-effects linear regression model.

Mean age at the time of enrollment was 2.3 years (range, 1-6.4) and 3.9 years (1-7.2) for the normal and CSM-GDs, respectively. Due to space constraints, only a portion of the results will be reported. Significant differences were found for the cranial articular facet area at all intervertebral spaces, for the caudal articular facet area at C6-7, and for the total facet area at all intervertebral spaces except T1-2, with CSM-GDs having larger facet areas. Regarding vertebral canal area, the measurements centered at the disc space and at the cranial aspect of the caudal vertebral body were significantly smaller in CSM-GDs from C2-3 through C6-7. The spinal cord area was significantly smaller on the CSM-GDs at C5-6 and C6-7 for all 3 levels measured. Middle foraminal height was significantly smaller in CSM-GDs from C3-4 through C7-T1. Intraobserver agreement was high for all measurements, with a median variation of 1.91% (0.36 to 10%).

This study provided the first MRI morphometric data of clinically normal GDs. The results indicate that GDs with CSM have vertebral canal stenosis throughout their cervical vertebral column at the disc space and cranial aspect of the vertebrae. Marked foraminal stenosis was also observed in GDs with CSM compared to normal GDs. An investigation of the anatomic features of the cervical spine of normal dogs is paramount for the understanding of the pathogenesis of cervical spondylomyelopathy (CSM) in giant breed dogs. Such investigation has not been performed. The purpose of this study was to prospectively characterize and compare the morphologic features of the cervical vertebral column of Great Danes (GDs) with and without clinical signs of CSM using magnetic resonance imaging (MRI).

Thirty client-owned GDs were prospectively enrolled, 15 clinically normal and 15 CSM-affected GDs. Inclusion criteria included a normal neurological examination for the normal GDs, and clinical signs compatible with CSM and diagnostic confirmation using MRI for the CSM group. All dogs underwent MRI of the cervical vertebral column (from C2-C3 to T1-T2 intervertebral spaces) with a 3.0 T MRI. Pulse sequences and planes acquired included: T1-weighted (T1W) (sagittal and transverse pre-contrast, sagittal post-contrast), T2-weighted (T2W) (sagittal, transverse, dorsal), and T2W sagittal images post-traction. Morphologic analysis included evaluation of the presence and degree of subarachnoid (SA) space and spinal cord (SC) compression, cause and direction of the compression, SC signal changes, articular joint characteristics, foraminal stenosis, intervertebral disc degeneration, SC contrast enhancement, differences at the compressive sites after traction, and visibility of ventral vertebral sinuses. The worst site of compression was recorded. A Fisher's exact test was used to analyze all results.

Mean age at the time of enrollment was 2.3 years (range, 1-6.4) and 3.9 years (1-7.2) for the normal and CSM-GDs, respectively. Due to space constraints, only a portion of the results will be presented. Within the control GDs, 9 sites of SA space and 2 sites of SC compression were noted in 6 dogs (40%). The 2 sites of SC compression were in the same control dog. Seventeen sites of SA space and 44 sites of SC compression were recorded in the CSM-GDs. All 15 CSM-GDs had at least 1 site of SC compression and all had more than 1 affected site when both SA space and SC compression were considered. The direction of compression was lateral caused by osseous changes in 72.7% and 96.7% of the affected sites for the control and CSM-GDs, respectively. The worst site of SC compression was at C6-7 in 8 out of the 15 CSM-GDs. Spinal cord parenchyma hyperintensities were noted at 14 sites in 9 CSM-GDs but none were seen in the normal group. Foraminal stenosis was present in 11 normal (73.3%) and 15 CSM-GDs (100%), but the number of sites and severity was significantly higher in the CSM-GDs (p < 0.001). Vertebral sinuses were visible in 78% and 47.6% of the normal and CSM-GDs intervertebral spaces imaged, respectively.

Clinically normal GDs can have MRI abnormalities without the presence of neurologic signs. The incidence of SC compression in control GDs was low (6.6%) but the incidence of foraminal stenosis was high (73.3%). The objective of this study was to evaluate the immediate postoperative recovery and the short-and intermediate-term follow-up of dogs with disc-associated-wobbler-syndrome (DAWS) treated with cervical-disc-arthroplasty (CDA) using the Adamo-Spinal-Disc. Eighteen dogs with over 2 months' history of DAWS diagnosed by MRI were evaluated. Thirteen dogs were treated for a single lesion, 4 dogs for a double lesion and 1 dog for a triple lesion.

The prosthesis and technique were similar to that described in the previous study [Adamo et al, Proc ACVIM, 2012] . For some cases, a redesigned thinner disc was employed in which the internal convex surface was replaced with PEEK (Polyether-Ether-Ketone). All dogs were evaluated neurologically shortly after surgery and with immediate and serial postoperative radiographs.

All dogs had uncomplicated recovery with good degree of distraction in the immediate postoperative radiographs. In the majority of dogs, the amount of distraction and the mobility at the treated sites decreased over time compared to the immediate post-operative status; this was less pronounced in the dogs treated with the thinner size implant. The loss of distraction (except for one dog) and the decreased mobility did not appear to be clinically significant. Median clinical follow-up was 18 months (1 -40 months). All but one dog showed improvement in neurological status during the observation period.

Cervical-disc-arthroplasty using the Adamo-Spinal-Disc appears to be a well-tolerated surgical technique, and it might be a valuable method to treat DAWS in dogs. The preliminary results of this study are promising. Long-term follow-up studies are underway. Several methods of surgical stabilization of the atlantoaxial joint (AA) are described in the veterinary literature. Threaded acrylic pins placed ventrally together with polymethylmethacrylate (PMMA) is a well-established technique that has been widely used in clinical cases. However, Kishigami tension bands, first described in 1984, are a less technically demanding procedure with potentially fewer complications. The purpose of this study was to biomechanically compare these two techniques in ventral-to-dorsal bending in both mature and immature dogs.

Seventeen normal canine cadavers weighing <15 kg were collected and radiographed to determine skeletal maturity. The cervical spines were dissected leaving bony and ligamentous structures intact. Eight mature spines and 9 immature spines were randomly divided into two groups. A Kishigami tension band was applied over the dorsal arch of the atlas and attached to the spinous process of the axis using orthopedic wire looped through two holes in the spinous process. The atlantoaxial joint was reduced as the wire was tightened and curled caudally prior to cutting.

In the second group, bilateral ventral acrylic pins were placed through the atlantoaxial facet angling cranially and dorsally, in the ventral pedicle of the atlas perpendicular to the spinal canal, and in the body of the axis in a ventromedial to dorsolateral direction. The pins were cut and PMMA was applied over the exposed implants. The specimens were then potted in custom steel pots and biomechanically analyzed in ventral-to-dorsal fourpoint bending using a Bionix MTS servohydraulic test system capable of a 5000N load at a speed of 5 mm/min. Loaddisplacement curves representing the degree of stiffness were compared between the groups. Stabilization using ventral pins and PMMA had a significantly greater stiffness than a Kishigami tension band when bending in ventral to dorsal bending. Within the stabilized vertebral segment, there was no significant difference between the stiffness of immature vs. mature bone. Further analysis in torsion and analysis in abnormal dogs will be helpful in establishing the clinical significance of these findings. Sepsis and acute lung injury are associated with substantial and long-term cognitive impairment. Activation of microglia is a potential cause of this cognitive change. Syndecan-4, a heparan sulfate proteoglycan (HSPG), is selectively increased in the lungs of mice treated with E. coli lipopolysaccharide (LPS) and is a marker of classically activated (M1) macrophages. The objective of this study was to measure syndecan-4 gene expression in the brains of mice following systemic administration of LPS. We hypothesized that syndecan-4 limits the extent of inflammation in the cerebral cortex caused by inflammation secondary to systemic LPS. C57Bl/6 mice were administered intratracheal (IT) LPS at 1 mg/kg and 3 mg/kg, and brain tissue was harvested at 6 h. Samples were analyzed using quantitative real-time PCR (Q-PCR) (n = 6 mice/time). The amount of mRNA recovered was reported as a fold increase over 0 hour controls, and values were reported as mean AESEM. Our studies show an increase in TNFa (91 AE SEM fold) and syndecan-4 (3.1 AE SEM fold) expression at 6 hours in cerebral cortex following intratracheal LPS at a dose of 3 mg/kg. No significant changes in syndecan-4 or TNF-a mRNA were seen in young (8 to 12 week old) mice treated with 1 mg/kg IT LPS; however, old (10 to 12 month old) mice treated with 1 mg/kg IT LPS showed significant increased TNF-a (19.1 AE SEM fold) and syndecan-4 (3.8 AE SEM fold) gene expression in the cerebral cortex. In conclusion, we found that syndecan-4 gene expression is increased in cerebral cortex following IT LPS administration, which may indicate an antiinflammatory role of syndecan-4, and that the inflammatory response in the CNS following IT LPS is more pronounced in older as compared with younger mice. Future studies to clarify the role of microglia in the CNS inflammatory response to systemic administration of LPS and the potential for syndecan-4 to limit this inflammation will be pursued in syndecan-4 null mice and their littermate wild type controls. There are three isoenzymes of creatine kinase (CK). CK-BB is found predominately in the brain, CK-MB is found in cardiac muscles, and CK-MM is found in skeletal muscles. Studies indicate that changes in total CK activity and isoenzyme distribution may be useful in determining the extent of neurological damage. Currently, conflicting reports exist regarding the stability of CK and its isoenzymes. The purpose of this study is to determine the optimal storage protocol for maximal detection of total CK and its isoenzymes in canine serum and cerebrospinal fluid (CSF).

Serum and atlantoaxial CSF were collected from 5 dogs deemed healthy by normal physical exam, neurological exam, CBC, serum chemistry, urinalysis, and CSF analysis. Samples were stored at 4°C, -20°C, and -80°C and analyzed at set time points for up to 6 months. Total CK activity was measured spectrophotometrically. Isoenzyme distributions were determined using the QuickGel â CK Vis Isoenzyme technique and densitometric scanning.

Total CK activity in serum remained stable at -20 ○ C and -80 ○ C for 6 months. Optimal storage for total CK activity in CSF was at -80 ○ C for 6 weeks. Interestingly, total CK activity in CSF was least stable at -20 ○ C. In serum, CK-MM and CK-BB followed similar degradation patterns at all temperatures for 4 weeks. In CSF, storage at -80 ○ C was optimal for both CK-MM and CK-BB, with greater than 70% retention for 8 weeks. CK-MB was negligible in serum and CSF. Optimal storage protocol is necessary for use of CK as a potentially useful biomarker in canines. Lactate dehydrogenase (LD) exists as five isoenzymes (LD-1 through LD-5) that are expressed throughout the body and can be detected in both the serum and cerebrospinal fluid (CSF). Studies in human medicine have demonstrated that changes in total LD activity and atypical isoenzyme patterns can indicate disease processes, including neurological abnormalities. However, the diagnostic significance of LD has been relatively unexplored in veterinary medicine. In order to ascertain the clinical usefulness of LD in veterinary medicine, this study sought to describe the normal levels and distribution of LD and its isoenzymes in the serum and CSF of healthy dogs.

Serum and atlantoaxial CSF were collected from 40 dogs deemed healthy by a normal physical exam, neurological exam, CBC, serum chemistry, urinalysis, and CSF analysis. Total LD activity was measured spectrophotometrically immediately after collection. Isoenzyme distributions were also determined within 5 hours of collection using the QuickGel â LD Isoenzyme technique and a densitometric scanner.

Serum levels of LD are higher than those found in the CSF. The median serum total LD in healthy canines is 74 U/L (CV 1.497642) while the median CSF total LD is 10 U/L (CV 1.290622). LD-5 is the predominant isoenzyme in canine serum, contributing nearly half of the total enzyme activity. Conversely, in canine CSF, LD-1 is the predominant isoenzyme, followed by LD-2 and LD-3. These findings will set the foundation for future studies of canine LD as a potentially clinically useful biomarker. Primary hematomyelia describes a localized region of hemorrhage within the spinal cord parenchyma with no identifiable underlying etiology. It is rare in humans and has not been well described in the veterinary literature.

This report describes clinical and imaging signs in three dogs: 1) 8 month old Labrador retriever presented for tetraparesis, worsening over one week; 2) 6 year old male Boxer presented for progressive paraparesis of eight weeks' duration; 3) 18 month old Siberian Husky cross with~48 hour history of progressive pelvic limb weakness and urinary incontinence. In each case, neurologic examination clearly signaled lesion localization: spinal cord segments C1-C5 in Case 1; T3-L3 in Case 2, and L4-S1 for Case 3.

MRI characteristics of the lesions were variable between cases, perhaps reflecting variation in the age of the hemorrhage, including both hyper and isointensities in the spinal cord on T2weighted imaging. A rim of contrasting intensity was observed in Cases 2 and 3. Lack of contrast enhancement of the intraparenchymal mass lesion and mild hydromyelia were the only consistent MRI characteristics. There were no specific MRI findings to differentiate primary hematomyelia from other intramedullary lesions with a mass effect. Blood tests revealed no evidence of clotting disorders in any of these dogs, and no evidence of systemic hemorrhage was observed.

Presumptive diagnosis was made through history and MRI findings, final diagnosis was confirmed by histological analysis of surgical biopsies and absence of underlying identified etiology. Each of the three patients made a good neurologic recovery after surgical removal of the hematoma. Traction responsive compressions in Cervical Spondylomyelopathy (CSM) are frequently treated with distraction fusion techniques, but long term distraction is seldom maintained.

The purpose of this study was evaluate with Xray, CT and MRI, a combined ventral and dorsal stabilization in regard to its ability of maintaining the applied distraction.

8 patients diagnosed with traction responsive ventral compression were treated. Clinical signs included ataxia (2), tetrapresis (4), tetraplegia (2).

Ventral stabilization was accomplished with a cylindrical titanium cage filled with cancellous bone inserted in the disc space, and in seven patients with a 6 screws cervical titanium locking plate, whereas in one with 6 screws and methacrilate. Dorsal stabilization was achieved by positioning one titanium screw in each vertebral facet joint through a left and right minimally invasive lateral approach, and a generous cancellous bone graft.

No violation of the spinal canal or of the foramen was observed at postoperative Ct. Patients were rechecked with Xray, CT and MRI at 2 to 18 months: shorter follow up was 4 month, the longer 18 months. The degree of decompression evaluated with MRI was considered excellent in the patients evaluated (7/ 8). In all the patients distraction was maintained at long term follow up. 6/8 patients regained a normal deambulation and 2/8 improved but a slight ataxia with hypometric gait in the thoracic limbs and hypermetric gait on the pelvic limbs persisted.

Combined dorsal and ventral stabilization can provide long term distraction in large breed dogs with traction responsive CSM. The purpose of this study was to determine the percentage of cats that achieved seizure control with serum phenobarbital (S-PB) concentrations of 15-45 lg/ml and to apply the 2011 International League Against Epilepsy (ILEA) classification to cats. Seizure control was defined as greater than or equal to a 50% reduction in seizures following phenobarbital administration.

The study design was multi-center retrospective, and included all cats with seizures except those with metabolic or toxic etiology. Cats were classified as having structural epilepsy, unknown epilepsy and probable unknown epilepsy. No cats were diagnosed with idiopathic epilepsy as genetic testing was not performed. Thirty-one cats met the inclusion criteria. Seizure control was achieved in 29/31 cats while on phenobarbital. Of these cats, 93% (27/29) had a S-PB concentration between 15 and 45 lg/ml. One cat from the unknown epileptic group had a S-PB concentration of 8 lg /ml and one cat with structural epilepsy had a S-PB concentration of 47 lg /ml. The remaining two cats did not achieve seizure control at S-PB concentration of 15-45 lg/ml.

This study demonstrates that the majority of cats (93%) will have seizure control with S-PB concentration of 15-45 lg/ml. In addition, the ILAE classification system was easily applied to veterinary patients and should be considered as a standard classification system for veterinary neurology. The most widely used method for determining the response to treatment in patients with cervical spondylomyelopathy (CSM) is based off of the owner and clinician's perception of the gait. This form of evaluation is highly subjective and can suffer from observer bias. The purpose of this study was to utilize selected force plate kinetic and digital video motion capture kinematic parameters before and after medical or surgical treatments, with the intent to develop a more reliable system for detecting, quantifying, and differentiating gait abnormalities in CSM.

Nine Doberman Pinchers with MRI-confirmed CSM were prospectively studied. Neurologic examinations were performed in all dogs both prior to treatment and 8 weeks later. Treatment consisted of medical management (n = 6) or surgery (n = 3). Force plate analysis and 3-D motion capture were performed at initial presentation and 8 weeks after starting treatment. Force plate analysis was performed in all four limbs of all dogs using a stationary force plate and computer system (Kistler Model 9687A). At least 4 runs of ipsilateral limbs were collected. Kinetic parameters evaluated peak vertical impulse (PVI), peak propulsive force (PPF), and peak mediolateral force (PMLF), and peak vertical force (PVF). For the kinematic study, dogs were fitted with a lycra bodysuit and 32 reflective markers representing specific anatomic landmarks from all four limbs, head and trunk were applied. 3-D motion capture was performed with 15 infrared cameras in a designated capture space using the Vicon8i motion capture system. Kinematic parameters included number of pelvic limb strides, stifle flexion and extension, maximum and minimum thoracic limb distance, and thoracic limb stride duration. A random effects linear regression model was used to analyze the data.

The results of the kinematics data indicated that while not found to be significant at 5% (P = 0.08; P = 0.110, respectively), maximum and minimum thoracic limb distance was found to be larger after treatment, with the maximum increasing from 190.1 mm to 203.1 mm (difference of 12.95 mm) and the minimum increasing from 130.2 mm to 140.6 mm (difference of 10.41 mm). These parameters were the only ones closest to significance at 5%. Force plate analysis revealed that, when comparing the means from each limb of all dogs, PVF was significantly different after treatment (P = 0.049). After performing a random effects linear regression utilizing the mean values from all limbs combined of all dogs before and after treatment, the difference of the PVF was significantly larger (P = 0.027). All other parameters were deemed to be not statistically significant.

In conclusion, while the maximum and minimum thoracic limb distance was not found to be significantly different between groups, the increase in distances suggests that the thoracic limbs may provide a way of measuring a patient with CSM's response to treatment. Additionally, PVF may also provide an objective means of assessing patient's clinical improvement. Cervical spondylomyelopathy (CSM) is one of the most common spinal diseases of large breed dogs. The most commonly affected breed is the Doberman Pinscher, with a breed prevalence of 5.5%. In spite of the high incidence of CSM in Dobermans, no study has identified a heritable etiology. Two large studies, one from England with more than 200 Dobermans, and another from New Zealand with 170 Dobermans failed to demonstrate an inheritable trait. The authors believe that the high incidence of CSM in Dobermans suggests a heritable basis and set out to investigate this hypothesis in Doberman Pinscher dogs from North America.

Medical records of the Veterinary Medical Center of the Ohio State University were searched for the terms "wobblers, CSM, and CVI". Owners of the identified dogs were contacted with a request to provide pedigrees. All owners of Dobermans with CSM diagnosed by one of the authors (RdC) at the Ontario Veterinary College were also contacted. Additionally, the Dobequest database (a Doberman health and wellness registry) was also searched and owners were contacted and asked to submit the medical records for review. All identified dogs (from all sources) with pedigree having at least 3 generations were classified into confirmed cases (those with signs suggestive of CSM and diagnostic confirmation by myelography, computed tomography or magnetic resonance imaging), or suspected (those with signs suggestive of CSM based on clinical examination with or without radiographs but without a definitive diagnosis). For purposes of pedigree analysis, cases were believed to be unaffected if they were at least 10 years of age or older and had no history of neurologic disease consistent with CSM.

A total of 60 Dobermans and their families were used in this study (38 confirmed cases and 22 suspected). Forty-five of the suspected or confirmed cases included additional information regarding the disease status (normal, suspected or confirmed) of their ancestors enabling the construct of multiple large family groups to study pattern of inheritance. These 45 confirmed and suspected cases included 20 females and 25 males, indicating no gender preference and suggesting an autosomal mode of inheritance. Five families included clinical and historical information on at least 3 confirmed or suspected affected dogs. An autosomal recessive mode of inheritance was ruled out based on the presence of an affected dog produced from the mating of an affected to an unaffected dog suggesting that an autosomal dominant mode is most likely.

We conclude that CSM in the Doberman pinscher is an inheritable disease that appears to be inherited as an autosomal dominant trait. Intracerebral hemorrhage (ICH) is a lethal stroke subtype. Advances in neuroimaging techniques, such as computed tomography and magnetic resonance imaging (MRI), enable prompt diagnosis of hemorrhage on brain parenchyma, which is necessary for the treatment of acute cerebrovascular accidents. Even though MRI has become a useful imaging tool to detect acute hemorrhage of human, standard diagnostic protocols for canine ICH are deficient in experimental and clinical neurology. Therefore the purpose of this study was to evaluate the diagnostic value of MRI in a canine model of ICH.

In 2 dogs, ICH was induced by injecting arterial autologous blood into the brain parenchyma. In another 2 dogs, 625 U of bacterial collagenase from Clostridium histolyticum was delivered into the parietal lobe over 5 min with a micro infusion pump. Imaging was performed on 3 T and 7 T MR systems at 1 day after induction of ICH. In blood injection model, hemorrhagic lesions within brain parenchyma were characterized by hypointense on T2-weighted images. However the signal intensity of corresponding areas on T1-weighted images was different in two dogs of blood injection model; In one dog, hypointensity was noted, but the other dog had hyperintensity. In collagenase injection model, all hemorrhagic lesions were characterized by hypointense on gradient-echo T2-weighted images and T1weighted MPRAGE images. Abnormal lesions of ICH were more evident at 7 T compared to 3 T. This study suggests that MRI may be a reliable diagnostic tool during the acute stage of canine ICH. Human studies have demonstrated that changes in total LD activity and atypical isoenzyme patterns can indicate neurological disease processes. However, conflicting reports regarding the stability of LD made it necessary to determine storage parameters before further study of the enzyme could be pursued. The purpose of this study was to maximize detection of LD and its isoenzymes in canine serum and CSF through proper storage.

Serum and atlantoaxial CSF were collected from 5 dogs deemed healthy by normal physical exam, neurological exam, CBC, serum chemistry, urinalysis, and CSF analysis. Samples were stored at 22°C, 4°C, or -20°C and analyzed at set time points for 2 months. Total LD activity was measured spectrophotometrically. Isoenzyme distributions were determined using the QuickGel â LD Isoenzyme technique and densitometric scanning.

Serum and CSF stored at -20°C retained greater total LD activity than at 4°C or 22°C. Total serum LD activity remained stable at -20°C for 4 weeks. CSF LD was more labile, retaining 86% of total activity 24 hours post-collection, but remained constant thereafter for 4 weeks at -20°C. Detection of LD isoenzymes in serum and CSF was optimized when stored at -20°C. While serum isoenzymes retained most of their activity for 4 weeks, CSF isoenzymes degraded rapidly, particularly LD-2, and thus should be evaluated within 72 hours. These findings will be important when immediate processing is unavailable. Great Danes (GDs) suffer predominantly from osseous-associated cervical spondylomyelopathy (CSM). Magnetic resonance imaging (MRI) is considered the gold standard in the diagnosis of CSM. However, computed tomography (CT) provides excellent bony detail. The purpose of this study was to prospectively evaluate and compare the morphology of the cervical vertebral column of CSM-affected GDs by use of non-contrast CT and MRI.

Fifteen client-owned GDs were prospectively enrolled. Inclusion criteria included the presence of clinical signs compatible with CSM and diagnostic confirmation using CT and MRI. All dogs underwent non-contrast CT (8-slice helical scanner) of the cervical spine under sedation (IV hydromorphone and dexmedetomidine) in sternal recumbency. Tranverse images aligned parallel to the endplates (slice thickness, 3 mm) were acquired. Soft tissue and bone windows were used for evaluation. MRI was obtained under general anesthesia with a 3.0 T MRI with the dogs in dorsal recumbency. Pulse sequences and planes acquired included: T1W (sagittal and transverse pre-contrast, sagittal postcontrast), T2W (sagittal, transverse, dorsal), and T2W sagittal images post-traction. The C2-C3 through T1-T2 intervertebral spaces were imaged with both modalities. Morphologic analysis included evaluation of the presence and degree of spinal cord (SC) compression (categorized as presumptive SC compression for CT images), worst site of compression, cause and direction of the compression, articular facet characteristics, and presence of foraminal stenosis. A Fisher's exact test was used to analyze all results.

Mean age was 3.9 years (1-7.2), and mean duration of signs was 1.9 years (0 months-5 years) at the time of enrollment. Due to space constraints, only a portion of the results will be reported. One hundred and five intervertebral spaces were evaluated (seven spaces per dog). Sixty-nine and 61 spaces were considered presumptively compressive or compressive based on CT and MRI, respectively. Eleven sites that were presumptively compressive on CT were non-compressive on MRI, and 3 sites that were considered compressive on MRI had not been categorized as such on CT. Agreement in the identification of the worst site of compression between CT and MRI was seen in 10 out of 15 dogs (66.6%). No significant differences were obtained regarding direction or cause of compression, degree of articular facet proliferation, and presence of foraminal stenosis between CT and MRI. The evaluation of the regularity of articular facets was significantly different (p = 0.008), with CT being more sensitive for facet sclerosis and irregularity.

Both non-contrast CT and MRI are useful in the evaluation of osseous-associated CSM. MRI offered direct visualization of the spinal cord and facilitated identification of compressive lesions. Canine osteosarcoma (OS) remains therapeutically challenging due to metastases development. We previously reported that zoledronate (ZOL) inhibits OS cell migration and disrupts cytoskeletal elements through the inhibition of Rho-GTPase prenylation. Despite these promising data, ZOL's antimetastatic activity, as a single-agent, was only observed when pre-treating cells in-vitro prior to mouse injection in murine OS models. In addition to cell migration and polarization, Rho GTPases promote anoikis resistance through activation of PI3K/AKT survival pathway. In lieu of single-agent studies, we hypothesize that ZOL-mediated perturbation of Rho-GTPase activity might sensitize OS cells to apoptotic triggers by virtue of PI3K/AKT attenuation; thereby amplifying the therapeutic activities of chemotherapy.

The effects of ZOL on K7M2 murine OS cells were investigated by characterizing 1) Cdc42 and Rac-1 Rho GTPases, p-p85 PI3K/p85 PI3K and p-AKT/AKT expressions by Western blot analysis; 2) apoptosis by flow cytometry; 3) combination index assessment by MTT assays; and 4) in vivo metastases development in a murine OS model. ZOL 1) reduced Cdc42 and Rac-1 membrane expression and p85 PI3K and AKT phosphorylation; 2) induced minimal apoptosis as a single agent at biologically achievable doses; 3) synergized with doxorubicin and cisplatin to inhibit cell proliferation assessed by combination index calculations; and 4) increased survival time when combined with doxorubicin in a murine model of experimental metastasis.

These study results suggest that ZOL can attenuate the PI3K/ AKT pathway in OS cells; allowing for ZOL's potential synergism with chemotherapy which subsequently augments inhibition of cell proliferation in vitro and anti-metastatic activities in vivo. These findings provide a biologic rationale for combining ZOL with chemotherapy in the treatment of OS metastases.

( Phagocytic function and oxidative burst are decreased in dogs with neoplasia compared to healthy dogs. In people, this is linked to more morbidity and poorer prognosis. This study compares neutrophil phagocytosis and oxidative burst in dogs with lymphoma before and during treatment to normal dogs. We hypothesized that dogs with lymphoma would have decreased phagocytic function and respiratory burst prior to receiving chemotherapy and normal function after chemotherapy.

Blood was collected from normal dogs (n = 10) at day 0 and 42, and dogs with lymphoma (n = 8) at day 0 and in remission at day 42 of a standardized CHOP protocol. Whole blood was stimulated with FITC labled opsonized E. coli and phorbol 12myristate 13 acetate (PMA). Percentage and quantity of cells undergoing phagocytosis and oxidative burst were analyzed using flow cytometry. Data were analyzed using two way repeated measures ANOVA.

There was no significant difference in the number of cells phagocytizing bacteria between normal dogs or dogs with lymphoma (p = 0.338). Dogs with lymphoma had lower respiratory burst activity when stimulated with E.Coli than normal dogs at day 0 (P = 0.019) and day 42 (P < 0.001). There was no significant difference in respiratory burst between day 0 and 42 in the lymphoma group or in the response to PMA in normal and lymphoma dogs.

This data suggests that dogs with lymphoma have normal phagocytic function but decreased respiratory burst, and that respiratory burst function is not regained in remission. This implies that dogs with lymphoma have innate immune dysfunction compared to healthy dogs.

( We examined the effects of CK2 downregulation on feline cancer cell lines. CK2 is a serine/threonine protein kinase complex that is essential for cell growth and survival and is also a potent suppressor of apoptosis. Increased expression has been found in most human cancers, making it an attractive therapeutic target. We hypothesized that CK2 downregulation would result in impaired growth and survival in feline cancer cells.

We verified the target sequence of feline CK2a and CK2a' by DNA sequence analysis of polymerase chain reaction products from peripheral blood cells from two healthy cats and from the feline SCCF-1 and K12 laryngeal squamous and mammary carcinoma cell lines. CK2 expression in feline oral squamous cell carcinomas (FOSCC) was assessed using immunofluorescence. SCCF-1 and K12 cell lines were treated with anti-CK2 oligonucleotides (OGNs) and CK2 expression and growth inhibition in treated cells were determined using immunoblotting and thymidine incorporation, respectively.

FOSCC samples showed overexpression of CK2 in tumour cells compared to adjacent normal tissue. OGNs directed against the CK2a and CK2a' subunits caused a time dependent decrease in CK2 protein expression and inhibited growth.

Our results show that CK2 expression can be inhibited in feline tumour cells by targeting CK2a and CK2a' subunits using OGNs and that reduction in CK2 is associated with growth inhibition. A nanocapsule delivery system has been shown to be safe in laboratory animals, with effective tumor targeting, paving the way to test safety and efficacy of this approach in pre-clinical studies of cats with spontaneously occurring tumors. Survivin is a member of the inhibitor of apoptosis (IAP) family, which inhibits apoptosis and cell proliferation in many types of human cancer cells, and is expected to be a novel agent for human cancer therapy. Canine histiocytic sarcoma (HS) requires highly effective medical therapy because of its aggressive local infiltration and very high metastatic rate. The aim of this study was to evaluate the influence of cell growth rate, chemosensitivity, and phagocytic activity by survivin inhibition in canine HS cell lines.

Canine survivin siRNA and YM155 (a small-molecule suppressor of human survivin) were used as survivin inhibitors. Seven HS cell lines, including 3 chemoresistant HS cell lines, and canine fibroblasts as the normal cell control were used in this study. Annexin V staining for apoptosis evaluation and MTT assay for cell growth rate and chemosensitivity were performed.

After transfection with canine survivin siRNA, induction of apoptosis, decrease in cell growth rate, and enhancement of chemosensitivity were observed in all HS cells. After administration of YM155, these changes were observed in 3 of 7 HS cell lines, and the other 4 HS cell lines showed only slight down-regulation by YM155. In addition, fibroblasts were influenced only to a negligible extent by survivin inhibition.

In conclusion, survivin was related to the inhibitory activities against canine HS. Thus, survivin-targeted therapy might constitute a potentially useful novel therapeutic approach for canine HS. Canine histiocytic sarcomas (HS) have aggressive local infiltration and very high metastatic rates. The cellular morphology of canine HS shows highly undifferentiated cells with a mixed pattern of round-cell and mesenchymal appearance. However, these tumors are often necrotic and inflammatory. These features can sometimes be misleading, making a definitive diagnosis difficult. Immunohistochemical staining using cell surface antigens (CD11b, CD11c, CD86, and MHC class II antigens) is extremely helpful for the final diagnosis of this tumor; however, this technique requires special antigens and frozen samples.

Real-time (RT)-PCR is a method that can quantitate mRNA levels. We established an RT-PCR method to analyze the abovementioned surface antigens. The aim of this study was to apply this technique to fine-needle aspirated (FNA) samples and evaluate its accuracy.

Eight dogs with suspected histiocytic sarcoma were included in the present study. FNA samples were collected from sarcoma masses. Normal tissues of healthy dogs from sites that corresponded to tumor origin sites in affected dogs were also analyzed for mRNA levels of each antigen. Tissues obtained from surgical biopsy or autopsy were used for histopathological diagnosis, including immnunohistochemical staining.

Biopsied samples could not lead to a definitive diagnosis, while RT-PCR showed more than 200 times mRNA levels of each antigen compared with normal tissues. In some dogs, the final diagnosis could not be confirmed by the histopathological method until postmortem examination.

These results indicated that cell surface antigen analysis by using RT-PCR has prospects of clinical application. The development of vaccines against cancer in human oncology is gaining increasing importance as a therapeutic approach which can complement standard chemotherapy and/or targeted therapies to achieve increased survival and improved quality of life for patients. In the specialty of veterinary oncology, there is an unmet need for additional therapeutic interventions such as immunotherapy, due to increased demand of owners seeking advanced options for cancer treatment for their pet. The use of high pressure, needleless devices as an enhancing tool for plasmid DNA delivery led to recent approval by USDA of Oncept TM , a therapeutic cancer vaccine directed against tyrosinase for the therapy of melanoma in dogs.

An alternative approach to improve plasmid DNA delivery is Electro-Gene-Transfer (EGT). In vivo electro-gene-transfer of plasmid DNA (DNA-EGT) is a safe methodology resulting in greater DNA cell uptake, enhanced protein expression and concomitant increase in longer term immune responses against the target antigen in a variety of species. The approach uses brief electrical pulses which create transient "pores" in the cell membranes that allow large molecules such as DNA or RNA to enter the cell cytoplasm. We have set up an optimal vaccination protocol in companion animals by means of a device, currently utilized and approved in Europe for Electrochemotherapy applications and adapted to Electro-Gene-Transfer.

We show that DNA-EGT can induce strong immune response against dog telomerase or dog HER2/neu targets especially when combined with Adenoviral (Ad) vectors. The vaccine resulted in significant therapeutic effects in dogs affected by B-cell malignant lymphosarcoma (LSA). The evaluation for other tumor types, such as mesothelioma, hemangiosarcoma, melanoma and renal cancer is in progress. No adverse effects have been observed in any canine or feline patient that could be attributed to immunotherapy.

We have established an optimized procedure for DNA-EGT. The technology is simple, safe and most importantly the vaccine prolongs the survival of canine patients when combined with Standard of Care. Thus, DNA-EGT holds promise as an adjunct therapy for the treatment of cancer in dogs and cats. Mammary tumours (MT) are among the most commonly encountered neoplasm in dogs. Forty one to fifty-three per cent of all mammary tumours in dogs are malignant, with carcinoma being the most common malignant type. Cyclooxygenase-2 (COX-2) expression in human tumours can be induced by growth factors, cytokines, oncogenes and other factors. Induction of COX-2 has been implicated in various cancers. Recently, systemic inflammatory response, as evidenced by elevated circulating concentrations of acute phase proteins, has been shown to be independently associated with poorer survival in human patients with advanced cancer. Elevated C-reactive protein identifies tumours capable of producing significant amounts of proinflammatory cytokines, in particular interleukin-6, and therefore with the potential for more rapid growth of tumour cells. The purpose of this work was to study the relationship between the systemic inflammatory response (C-reactive protein, CRP and Haptoglobin, Hp) and local inflammatory response (COX-2 expression) in different canine mammary tumors.

Serum concentrations of CRP and Hp were determined in serum samples from healthy dogs (n = 20) and dogs with mammary tumors (n = 41), prior to surgery. Forty one mammary tumours were submitted for histopathological and immunohistochemical study. Mammary tumour type was determined by histopathologic examination according WHO Guidelines. From all studied samples, 32 were malignant MT and 9 were benign MT. Simple or complex adenomas (n = 3), mixed benign tumours (n = 6), carcinomas (n = 24), fibrosarcoma (n = 1) and mixed malignant tumour (n = 7) were studied by immunohistochemistry for COX-2 expression. A gradation score according the percentage of cells expressing COX-2 and the intensity was applied.

All the adenomas and 5 out of 6 (83.3%) mixed benign tumours were COX-2 positive. Immunostaining of malignant tumours revealed that 4 of 7 (57.14%) mixed malignant tumors and 12 of 24 (50%) carcinomas were COX-2 positive. CRP levels ranged from 0.05-70.6 mg/L, with a mean value of 1.7 mg/L. Hp values ranged from 0.11-128.9 g/L, with mean value of 9.69 g/L. No significant differences were observed between benign and malignant MT according COX-2 expression and intensity. No significant correlation has been found between CRP and Hp levels and COX-2 expression or intensity in MT. According our results, there is no association between systemic and local inflammatory response in canine mammary tumors. Recent evidence in human cancer patients and rodent models suggests that the neutraceutical sulforaphane (typically found in raw cruciferous vegetables) may have utility in prevention of cisplatin induced renal toxicity, however there is little examination on its effects on cancer cells with or without conjunctive chemotherapeutics. To examine the potential safety and effects on canine cancer cells three canine cell lines were treated with a serial dilution of sulforaphane (0-50 uM) for 48 hours and 6 day growth curve assays were performed using MTT assays. Apoptosis was examined using annexin V staining, or caspase 3 immunoblotting. Further viability assays were performed to examine sulforaphane treatment in conjunction with either palladia (mast cells) or doxorubicin (osteosarcoma). Statistical analysis was performed using analysis of variance with Tukey's post hoc analysis or Student's T test to determine significance. Results suggest that all cell lines have reduced cell viability in both the 48 hour MTT proliferation and the growth curve analysis at between 2 and 3 uM of sulforaphane (p < 0.05) with apoptosis occurring in two of the three cell lines (mast cells and osteosarcoma cells) at concentrations between 10-20 uM. Sulforaphane did not promote cell survival nor did it promote further cell death in the face of other chemotherapeutic insults in mast or osteosarcoma cells. Further mechanisms for reduced cell viability are being examined, however sulforaphane does not confer a survival advantage for these cell lines making it a potential neutraceutical to investigate for chemotherapy-induced renal cytotoxicity. We previously described a population of lymphoid progenitor cells (LPCs) in canine lymphoma that persist in the xenotransplantation setting. Our hypothesis was that LPCs behave like tumor-initiating cells and that PSC-833, a selective inhibitor of the ATP binding cassette B1 transporter (ABCB1, a.k.a., p-glycoprotein/multidrug resistance protein-1) would sensitize them to doxorubicin and improve remission. Dogs with therapy-na€ ıve diffuse large B-cell lymphoma were eligible to participate. Twenty dogs were enrolled into a double-blinded, placebo controlled, multi-institutional study. The experimental and control groups received oral PSC-833 (7.5 mg/kg) or placebo twice daily for 5 days, respectively, followed by 5 doses of doxorubicin 21 days apart, with the first dose reduced to 21 mg/m 2 to mitigate potential side effects of ABCB1 inhibition. LPCs in lymph node biopsies and peripheral blood at diagnosis, at the end of neoadjuvant period, and 1-week after the first chemotherapy dose were quantified using flow cytometry. Interim data analyses show a trend to delayed progression in the PSC-833 group compared to placebo, and significantly delayed progression associated with stabilization or reduction in the percent of LPCs in lymph nodes at the end of the neoadjuvant therapy period. The relationship between ABCB1 + lymphoma cells and LPCs and the results from final analyses with censoring after 12-months of follow-up will be discussed. There is currently limited prognostic information available and no definitive method for distinguishing between a benign or malignant splenic mass prior to surgery. The purpose of this study was to determine whether age was significantly different between dogs with benign versus malignant splenic masses.

Medical records of dogs examined between August 2009 and August 2012 because of a splenic mass were reviewed. Information obtained included dog age, sex, breed and weight; whether hemoperitoneum was present; histologic diagnosis; and survival time.

18 (36%) had hemangiosarcoma (HSA), 2 (4%) had other types of malignant masses, and 30 (60%) had benign masses. Mean age of dogs with HSA was 10.4 years +/-2.1 years (range, 4 -15 years), mean age of dogs with benign masses was 10.4 years +/-2.9 years (range, 7 -13.5 years), and mean age of the 2 dogs with malignant masses other than HSA was 14.2 years (range, 13.5 -15 years). There was no significant difference (P = 0.98) in age between dogs with HSA and dogs with benign masses.

Results suggest that age is not useful in differentiating between HSA and benign lesions in dogs with splenic masses. Feline oral squamous cell carcinoma (FOSCC) is an important problem in veterinary oncology. Cats are typically presented when tumors are beyond surgical resection. Outcomes of radiotherapy and chemotherapy have been disappointing. Targeting signaling pathways is a contemporary strategy for impairing growth of cancer cells. FOSCC has been shown to express epidermal growth factor receptor (EGFR) and activation of EGFR by binding of epidermal growth factor triggers signaling cascades resulting in proliferation.

Two approaches have emerged for interrupting EGFR signaling in human head and neck squamous cell carcinoma that may be applicable to FOSCC. They include: 1) blockade of EGF binding to EGFR by a monoclonal antibody such as cetuximab, and 2) targeted small molecules that inhibit activation of signaling tyrosine kinases (TKs).

We employed immunohistochemistry, flow cytometry, western blot, and cell viability techniques to begin to evaluate these strategies for FOSCC therapy. Archived FOSCC biopsies and the FOSCC cell line SCCF1 (provided by Dr. T. Rosol) were studied. Effects of several tyrosine kinase inhibitors (TKIs) that target EGFR or other TKs associated with epithelial cancer were determined as was binding of cetuximab to FOSCC.

Immunohistochemistry confirmed cetuximab bound FOSCC biopsies. Although western blot confirmed expression of EGFR and the downstream transcription factor Ctip-2 by SCCF1 cells, cetuximab didn't bind SCCF1 cells when examined with flow cytometry. Gefitinib suppressed SCCF1 growth but erlotinib, lapatinib, or vandetinib, TKIs that target EGFR, did not. Dasatinib and sunitinib were markedly more potent inhibitors of SCCF1 proliferation. EGFR/TK targeting offers potential options for FOSCC therapy. Recent advances in molecular technologies, such as high throughput sequencing technology, have allowed for comprehensive evaluation of the intestinal bacterial population (microbiome). The objective of this study was to characterize the fecal microbiome of dogs with lymphoma by high throughput sequencing technology and to investigate the impact of chemotherapy.

Six dogs presented at the Ontario Veterinary College Oncology Service had a cytological or histological diagnosis of multicentric lymphoma and treated with a 25-week CHOP protocol. Fecal samples were collected at presentation and 9 weeks after beginning of treatment. High throughput sequencing of the 16S rRNA gene PCR products was performed. Sequences were classified into operational taxonomic units (OTUs) and bacterial structures compared by the Classical Jaccard measure of dissimilarity and the inverse Simpson's diversity index.

A total of 67,469 sequences were obtained, of which 56,693 were high quality reads and therefore used for final analysis. The number of OTUs per animal varied from 38 to 182. Treatment had no significant impact on diversity (P = 0.264). Bacterial communities before and after chemotherapy were not significantly different compared by the parsimony test (P = 0.932). The main phyla commonly on feces of dogs, Firmicutes, Bacteroidetes and Proteobacteria, did not change significantly after treatment (P = 0.963, P = 0.857 and P = 0.953, respectively). A phylogenetic tree clustering animals by similarities among bacterial communities confirmed that animals tended to cluster by individuals and not by time of sampling. This is the first report of the use of high throughput sequencing to address changes caused by chemotherapic drugs on the canine intestinal microbiome. No apparent changes on the fecal microbiome were evident after 9 weeks of chemotherapy in dogs with multicentric lymphoma. Gastrointestinal effects of chemotherapy might be more related to changes in immune status or other factors rather than alteration of the gut microbiome. Obesity is a risk factor for transitional cell carcinoma (TCC) development in dogs. Adiponectin is a hormone produced by adipocytes. It exerts its effects, at least in part, through adiponectin receptors (AdipoR1 and AdipoR2). In humans, AdipoR expression is altered in cancers that are associated with obesity. Depending on concentrations, adiponectin can have positive or negative effects on cancer cell proliferation. The role of adiponectin and AdipoR in canine TCC is not known. The objective of this study was to determine expression of AdipoR1 in TCC cell lines and in tissues from normal canine bladder and primary and metastatic TCC.

AdipoR1 was detected in canine tissues and TCC cell lines by immunohistochemistry and Western blot, respectively, using ab70362 (Abcam, Cambridge, MA). The percentage of immunoreactive cells and staining intensity (0.5-3 + ) were recorded. Tissues with immunoreactivity in greater than or equal to 10% of cells were considered positive for AdipoR1 expression.

AdipoR1 was detected in 12 of 12 (100%) normal canine bladder samples, 16 of 35 (46%) primary TCC tissues, 2 of 5 (40%) lymph node metastases, and 6 of 11 (55%) lung metastases. Percent positive cells ranged from 10-95% with a median of 2 + stain intensity. AdipoR1 was detected in 5 of 11 (45%) TCC cell lines.

AdipoR1 was expressed in 100% of normal bladder tissues. AdipoR1 was expressed less frequently in canine TCC cell lines and tissues. Further study is warranted to determine the potential protective role and other effects of AdipoR1 in canine TCC development.

( Metastasis is the major cause of death in dogs with osteosarcoma (OSA). Inhibiting OSA cell invasion and migration could suppress metastatic potential of this cancer. Hepatocyte growth factor (HGF) is the ligand for c-Met, a receptor tyrosine kinase, and stimulation of c-Met by HGF has been implicated in the genesis and malignant progression of human OSA. Recent studies have focused on the role of tyrosine kinase inhibitor (TKI) targeting the HGF/c-Met signaling axis, in which TKIs have been found to decrease invasiveness of human OSA cells in vitro.

Using scratch and matrigel invasion assays, this study evaluated the role of HGF in stimulating canine OSA cell invasion and migration and the potential for c-Met inhibition by the TKI crizotinib to mitigate this activity. HGF promoted significant OSA cell invasion and migration in a dose-dependent manner, with 10 ng/ml demonstrating the maximal effect using the D17, COS, Clone-4, and POS canine OSA cell lines. The effect was most pronounced in COS cells. Results of dose-response experiments using the Clone-4 cell line indicated a crizotinib concentration of 30 nM was effective at inhibiting cell invasion and migration. HGF could overcome the inhibitory effects of crizotinib in a dose-dependent manner helping to confirm crizotinib mediated on-target effects against c-Met.

Invasion and migration are malignant properties of OSA cells that appear to be mediated by HGF stimulation of the c-Met receptor and targeting c-Met signaling with a TKI such as crizotinib may represent a valid therapeutic approach to develop for canine OSA. Cancer is most often disease in older animals, however therapeutic efficacy is limited by current treatment as surgery, chemotherapy, or radiation. Photodynamic therapy (PDT) is one of new rising modalities for management of cancers because it is selective to target-tumor cells with less side-effect. This study aims to establish the safety of systemic administration with a newly developed photosensitizer without light activation in normal dogs.

10 adult normal beagles who received the photosensitizer intravenously were classified into two groups as high-dose (H group; 2 dogs, 10 mg/kg) and low-dose (L group; 8 dogs, 2 mg/kg), and the low-dose group was further divided into four subgroups by different dilution volume and infusion rate. All dogs were excluded from light for post 48 hours and monitored activity, vital signs, blood work, and thoracic radiographs on day 0, 2, 3, 7, and 10. They were euthanized and performed necropsy on day 10.

On radiographic examination, one of L group showed bronchointerstitial pattern changes on left caudal lung lobe. No other dog had clinical significant changes. On gross observation of necropsy, generalized hemorrhage of lung surface was appeared in each two of L group and H group. And they had inflammatory cell infiltration and loss of structures in the peribronchial and alveolar tissue on histopathologic findings.

This study suggests that administration of fotoditazin intravenously had no remarkable clinical adverse effect without exposure of light. However respiratory potential in dogs could be related total dose, dilution volume, or infusion rate. Feline hypersomatotropism (HS) appears to be a significant cause of feline diabetes mellitus. However, successful treatment of HS is currently challenging. Radiotherapy and hypophysectomy seem the only effective therapeutic modalities, yet come with significant disadvantages. Medical options would be desirable although somatostatin (sst) analogues and dopamine agonists have thus far proven largely ineffective. Pasireotide (SOM230), a novel multi-receptor ligand sst analogue with high binding affinity for sst receptor subtypes 1, 2, 3 and 5 has been shown to suppress growth hormone (GH) and insulin-like growth factor-1 (IGF-1) in rodents as well as humans suffering from HS. Additionally, direct and indirect anti-tumor activity has been observed in vitro including sst receptor-mediated apoptosis and anti-angiogenesis. The current study aimed to assess the potential of SOM230 as a treatment modality for naturally occurring feline HS.

Feline HS was diagnosed in eight diabetic cats by documenting serum IGF-1 concentration > 1000 ng/ml (radioimmunoassay) and presence of a pituitary enlargement (computed tomography). On day 1 and 5, serum IGF-1 concentration was established and glycemic control assessed using a 12-hour blood glucose (BG) curve, measuring BG every 2 hours. On day 2, 3 and 4, the cats were injected with 0.03 mg/kg SOM230 s.c. BID. The initial insulin dose was dictated by the choice of the attending clinician, although was reduced according to regular BG measurements during the treatment period to avoid hypoglycemia. Pre-and post-treatment IGF-1, average 12-hour BG and insulin dose were compared using a paired t-test (significance at P < 0.05).

All eight cats showed a significant decrease in serum IGF-1 (mean+/-SD day 1: 1884 + /-218 ng/ml; day 5: 1169 + /-395 ng/ ml, p = 0.001) and average 12-hour BG (day 1: 20 + /-5 mmol/l; day 5: 13 + /-4 mmol/l; p = 0.002). A significant insulin dose reduction was necessary in all cats (day 1: 10.8 + /-6 iu/injection; day 5: 3.1 + /-2 iu/injection; p = 0.015). No side effects were noticed during or after the 3 day treatment period, apart from hypoglycemia in one cat, which resolved after provision of food and reduction of insulin dose.

The current study indicates that SOM230 is able to rapidly decrease GH and IGF-1 concentrations in feline HS. This therefore suggests that sst receptors are present in most feline somatotrophinomas, which has previously been unclear given the disappointing results during somatostatins and sst analogue therapy attempts. A return of insulin sensitivity was seen, enabling improved glycemic control to be established with reduced doses of exogenous insulin in all cats. In light of these results, a clinical trial with a longer-acting formulation of SOM230 is currently being conducted to establish long-term effects and potential for diabetic remission.

EN-2 ECTOPIC SUBLINGUAL THYROID NEOPLASIA IN THE DOG: 25 CASES (1995 -2012 . MR Broome 1 , ME Peterson 2 , J Walker 1 . 1 Advanced Veterinary Medical Imaging, Tustin, CA, 2 Animal Endocrine Clinic, New York, NY.

Thyroid embryology defines the normal migration of thyroid tissue from its origin as an epithelial proliferation in the floor of the pharynx at the base of the tongue (ie, the foramen caecum) along the path of the thyroglossal tract. The embryologic path of the thyroid gland includes the tongue and hyoid apparatus. Failure to associate fully with the embryologic aortic sac leads to the incomplete descent of the thyroid and the presence of sublingual ectopic thyroid tissue. Ectopic thyroid tissue in this location can become occasionally become neoplastic. Two previous reports describe a total of 9 cases of sublingual thyroid carcinoma in dogs (JAVMA 1989; 195:1606; ASVS Veterinary Symposium 2011; 217) . No reports of benign thyroid neoplasia developing in this location have been identified. Medical records reviewed between 1995-2012 revealed 519 dogs with thyroid carcinoma, confirmed by thyroid scintigraphy. During that period, 25 dogs with ectopic sublingual thyroid neoplasia were identified (5% of all thyroid carcinoma cases). The dogs ranged in age from 4-15 years (median, 9 yrs), with 13 neutered males and 12 spayed females. Breeds included mix (6 dogs), Golden Retriever (4), American Staffordshire Terrier (2), and Labrador Retrievers (2). Eight dogs (32%) had high serum T4 levels, consistent with hyperthyroidism. Twelve dogs had normal total T4 levels, 4 dogs had low values, and 1 dog's T4 value was not available. Four dogs had pulmonary metastases confirmed by scintigraphy and 4 had concurrent cervical disease, and 1 dog had both concurrent cervical disease and pulmonary metastasis. Seventeen dogs had histopathologic confirmation and 7 dogs had cytologic confirmation of thyroid malignancy. Fourteen dogs were treated with surgery; of these, 9 had excision of the basihyoid bone. All tolerated surgical intervention, including hyoid bone resection. Gross surgical excision of thyroid disease was confirmed scintigraphically in 7 dogs, 5 of which required hyoid bone resection. Thirteen dogs were treated with high-dose radioiodine (dose range, 40-130 mCi; mean, 95 mCi). No lasting complications following radioiodine were identified. Radioiodine therapy resulted in a marked response, as measured by percent decreased radionuclide uptake (range 42-100%, mean, 80%) in 8 dogs with gross disease. Radioiodine was also successful in ablating normal thyroid tissue in 3 dogs without persistent gross disease. Nine dogs were treated with both surgery and radioiodine. Surgery was performed prior to radioiodine therapy in all 9 dogs. One dog with a thyroid tumor deemed initially surgically unresectable had a successful surgical excision following radioiodine therapy.

Conclusions: In dogs, ectopic sublingual thyroid carcinoma is not uncommon. Surgical excision of the basihyoid bone is often indicated for gross resection of laryngeal thyroid carcinoma and is well tolerated. Adjunctive high dose radioiodine therapy is indicated for cases of incomplete surgical excision of local disease or when metastatic disease is confirmed scintigraphically. Hyperthyroidism develops in cats secondary to 1 or more autonomously functional thyroid adenomas. The progressive thyrotoxicosis that ensues causes the chronic suppression of endogenous TSH release and ultimately the atrophy of normal thyroid tissue in these cats. This thyroid atrophy can lead to a period of transient hypothyroidism following curative radioiodine therapy. Once T4 values fall, circulating TSH levels increase, leading to reactivation of the previously suppressed and atrophied thyroid tissue in the large majority of these cats.

Between 30-40% of cats with hyperthyroidism have pre-existing chronic kidney disease (CKD). Iatrogenic hypothyroidism has been shown to contribute to worsening of azotemia and shortened life expectancy in cats with pre-existing CKD (Williams et al, J Vet Intern Med. 2010; 24:1086) . In hyperthyroid cats with concurrent azotemia, the transient hypothyroidism that follows radioiodine therapy may contribute to additional renal function decline and worsening of the cats' CKD stage.

The purpose of this study was to evaluate if prevention of this transient hypothyroidism would blunt the progression of azotemia commonly seen following the resolution of thyrotoxicosis in these cats with preexisting CKD. In this study, 195 hyperthyroid cats with concurrent CKD (IRIS stage 2 to 3) were treated with radioiodine (range, 1-10 mCi, median, 3 mCi). Of the 195 CKD cats, 85 cats were discharged on L-T4 (0.1 mg, PO q24 h), whereas the remaining 110 cats served as controls (no L-T4 supplementation). In both groups, total T4, BUN, and creatinine levels were recorded before treatment and then again at 1, 3 and 12 months following radioiodine therapy.

Following successful radioiodine therapy, both groups of cats with preexisting CKD demonstrated increases in serum BUN and creatinine levels that gradually progressed over the 12-month period (Table 1) . However, the percent rise in median creatinine concentrations in the 85 cats treated with L-T4 was significantly less than the rise in the 110 cats not supplemented with L-T4 (12.5% vs 33.3%; P < 0.05). These results suggest that L-T4 supplementation of radioiodine-treated cats with CKD may help limit progression of azotemia, presumably by avoiding the transient hypothyroidism that commonly develops after radioiodine therapy. Hyperthyroid cats are reported to have reduced blood ionised calcium concentrations (iCa) and elevated plasma parathyroid hormone (PTH) concentrations, however, the pathophysiological mechanism for these changes is unknown. Intestinal absorption of calcium is regulated by calcitriol, and reduced plasma calcitriol concentrations are reported in human Graves' disease patients. Therefore, if hyperthyroidism was associated with calcitriol deficiency in cats, this could be the pathophysiological mechanism for hypocalcemia and elevated plasma PTH concentrations. The aim of this prospective study was to investigate the association between iCa and plasma concentrations of PTH and calcitriol in hyperthyroid cats.

Newly diagnosed hyperthyroid cats (plasma total thyroxine (TT4)>55 nmol/l) from two London-based first opinion practices were prospectively recruited into the study in 2011-12. A group of non-azotemic euthyroid (TT4 < 40 nmol/l) geriatric (>9 years old) cats (control) were also recruited into the study for comparison of iCa and plasma calcitriol concentrations. The Mann-Whitney U test was used to compare baseline variables between hyperthyroid and control cats. Multivariable linear regression analysis was used to identify independent predictors of iCa in hyperthyroid cats. The first multivariable model included variables associated with iCa at the 10% level (P < 0.1) in the univariable analysis. The second model included TT4, PTH and calcitriol. Data are presented as median [25th, 75th percentile] and statistical significance defined as P < 0.05.

Hyperthyroid cats had significantly lower iCa than control cats (1.24 [1.21, 1.27] mmol/l, n = 45 vs. 1.26 [1.23, 1.29] mmol/l, n = 52; P = 0.003). Hyperthyroid cats also had significantly higher plasma calcitriol concentrations than control cats (115 [99, 144] pmol/l, n = 20 vs. 89 [61, 108] pmol/l, n = 10; P = 0.037). Linear regression analysis identified TT4 (P < 0.001), venous pH (P = 0.019), and plasma globulin concentration (P = 0.002) as being associated with iCa at the 10% level. Plasma concentrations of creatinine (P = 0.109), albumin (P = 0.125), PTH (P = 0.691) and calcitriol (P = 0.646) were not associated with iCa at the 10% level in the univariable analyses. In the first multivariable model, TT4 (P = 0.003) and venous pH (P = 0.026) were independently associated with ionised calcium concentration, and plasma globulin concentration tended towards an independent association with iCa (P = 0.084). In the second multivariable model, only TT4 was an independently associated with iCa (P = 0.003). Plasma concentrations of PTH (P = 0.442) and calcitriol (P = 0.492) were not independently associated with iCa after adjustment for TT4. Hyperthyroid cats have increased plasma calcitriol concentrations, therefore calcitriol deficiency does not appear to be the pathogenetic mechanism for altered calcium homeostasis in hyperthyroid cats. One interpretation of these data is that thyroid hormones cause hypocalcemia either directly or through another currently undetermined mechanism that is independent of control by PTH and calcitriol. The calcium sensing receptor (CaSR) is a G protein-coupled receptor expressed in the parathyroid gland, kidney, bone and intestine, where it regulates plasma ionised calcium concentrations (iCa). Over 200 mutations have been identified in the human CaSR, some of which cause gain or loss of function, resulting in inherited defects of calcium homeostasis. Single nucleotide polymorphisms (SNPs) in the CaSR have also been linked to the severity of secondary renal hyperparathyroidism (SRHP) and in the pathogenesis of urolithiasis involving calcium containing stones. The coding region of the feline CaSR (fCaSR) spans 6 exons, in which four SNPs have previously been identified in six young healthy cats; three of which were synonymous and one non-synonymous (R1044P). The aim of the current study was to identify further SNPs in the fCaSR in cats with normal and dysregulated plasma iCa, for use in future genetic studies.

Twenty renal azotemic (plasma creatinine concentration >2.0 mg/dl and USG <1.035) cats >9 years old, seen at two firstopinion practices since January 2008 and eating commercially available renal diets were identified. Genomic DNA was extracted from buffy coat-enriched packed cells, collected into potassium EDTA tubes and stored at -80°C. The coding sequence of fCaSR was available on Ensembl. Missing intronic sequence flanking exon 3 was identified by performing basic local alignment search tool (BLAST) using the available feline CaSR sequence against the feline genome via Ensembl. Sequences were confirmed by polymerase chain reaction (PCR) amplification and sequencing of exons, using previously published or newly designed fCaSR-specific primers in flanking intronic regions. PCR products were sequenced in 10 cats with high iCa (iCa>1.34 mmol/l) and 10 cats with low/normal iCa (iCa 1.34 mmol/l).

The high iCa (range 1.42-1.85 mmol/l) group included 5 domestic shorthair (DSH), 1 domestic longhair, 2 Burmese, 1 Persian and 1 Russian blue cross cat. The low/normal iCa (range 1.14-1.3 mmol/l) group included 7 DSH, 2 Burmese and one British shorthair cat. Sequence alignment against the fCaSR sequence revealed 8 novel SNPs in the coding region; synonymous SNPs at nucleotide positions 732(C>T), 981(C>T), 1863(G>C), 1992 (C>T), 2061(C>T) and 2109(C>T) and non-synonymous SNPs at nucleotide positions 2780(C>T) and 3088(G>A), which change amino acid residues 927 (proline to leucine) and 1030 (alanine to threonine) respectively. The SNP at position 981 was present exclusively in the Burmese cats (one homozygous and one heterozygous cat in each group). The SNPs at positions 1992, 2061, 2780 and 3088 were present exclusively in the Persian cat (all homozygous).

In total 12 SNPs have now been identified in the fCaSR. Further research is required to determine whether these are associated with ionized hypercalcaemia, SRHP or urolithiasis in cats of different breeds. This study was designed to evaluate whether stereotactic radiation therapy is an effective therapy for dogs with pituitary dependent hyperadrenocorticism (PDH).

Twelve dogs were diagnosed with PDH based on clinical signs, endocrine testing, and abdominal imaging. Dogs received three consecutive fractions of stereotactic radiation therapy directed at the pituitary gland. An ACTH stimulation test, endogenous ACTH, total T4, and TSH were obtained at 0, 1, 3, 6, 9, and 12 months post-treatment.

Three patients were euthanized prior to completion of the study: 2 due to neurological disease and 1 due to progression of clinical signs, each at 6 months post-treatment. Three dogs required thyroid supplementation due to hypothyroidism.

Pre-ACTH cortisol decreased significantly over time (p = 0.03). Post-ACTH cortisol did not change significantly (p = 0.63). Endogenous ACTH did not change significantly (p = 0.16). T4 and TSH did not change significantly over time (p = 0.40 and p = 0.25, respectively). Six owners reported improvement in their dogs' clinical signs throughout the course of the study; of these dogs, four had post-cortisol levels within normal limits at the time the last sample was collected.

Although stereotactic radiation therapy was effective in treating a subset of patients with PDH, this therapy does not appear to be uniformly effective in managing the majority of dogs with PDH, at least for the initial 12 months after stereotactic radiation therapy. Primary hyperaldosteronism in cats has historically been associated with adrenocortical tumors, but recently primary hyperaldosteronism associated with adrenocortical hyperplasia was reported. Diagnosis of primary hyperaldosteronism in cats without an overt adrenal tumor can be difficult due to the lack of definitive testing. Our objective was to determine if the oral administration of fludrocortisone acetate would suppress endogenous serum aldosterone in healthy cats. If so, a suppression test could potentially be used as a functional diagnostic test in cats suspected to have primary hyperaldosteronism.

Eight research cats, determined to be healthy based on normal physical examination, complete blood count, serum chemistry, urinalysis, and urine protein:creatinine ratio were divided into two groups of four. Fludrocortisone acetate (0.05 mg/kg q12 hours per os) and placebo (q12 hours per os) were administered to each group in a crossover design. Both groups were treated for four days with a four-day washout period between phases. Blood was collected for measurement of serum aldosterone during each phase at baseline (d = 0 or 1) and at 4-hr post pill on days 2, 3, and 4. Aldosterone was measured using a commercially available radioimmunoassay kit previously validated for use in cats. Within each group, aldosterone concentrations were compared between days using a repeated measures ANOVA on ranks. Between groups, concentrations were compared on each day using the rank sum test. Significance was set at the p < 0.05 level.

The median (range) baseline serum aldosterone concentration in cats receiving placebo and fludrocortisone was 142.5 (22-239) pg/mL and 162 (100-258) pg/mL, respectively (P > 0.05). Aldosterone concentrations were significantly lower on day 2 (median aldosterone concentration 57.5 [25-95] pg/mL) in fludrocortisonetreated cats compared to baseline. Aldosterone concentrations on day 3 and 4 were not significantly different from baseline. In response to fludrocortisone, serum aldosterone declined >40% from baseline in 8/8, 7/8, and 6/8 cats on days 2, 3, and 4, respectively. With placebo treatment, random decreases of aldosterone >40% from baseline occurred in 2/8, 2/8, and 0/8 cats on days 2, 3, and 4, respectively.

Administration of oral fludrocortisone at 0.05 mg/kg q 12 hours for 3 doses caused a significant suppression of serum aldosterone concentration in healthy cats. Suppression of aldosterone would be expected to be absent in cats with primary hyperaldosteronism. Further investigation in patients with known or suspected primary hyperaldosteronism is indicated to determine the clinical utility of this test as a tool for definitive diagnosis. The largest published case series of spontaneous feline hyperadrenocorticism (HAC) includes 10 cases. The aims of this study were to evaluate the outcome of diagnostic tests and treatments in a larger series of feline HAC. Thirty-two cats with spontaneous HAC were identified from ten referral hospitals: 27 cats with pituitary dependent (PDH) and 5 with adrenal dependent (ADH) disease. Results of ACTH stimulation tests, dexamethasone suppression tests (DST), imaging and treatment were recorded. ACTH stimulation tests in 23 cats with non-adrenal illness were used to determine test specificity. Sensitivity was defined as the proportion of positive test results in HAC cats, and specificity as the proportion of negative results in control cats.

Twenty of 23 controls had within reference range 60-minute post-ACTH cortisol concentration (specificity 87%). Seven of 18 HAC cats had abnormal post-ACTH concentration (sensitivity 39%), 27 out of 30 lacked cortisol suppression at 8 hours on the DST (sensitivity 90%). The sensitivity of abdominal ultrasonography for differentiation of ADH versus PDH was 90%. Pituitary masses were identified in all 9 cats that underwent cranial imaging (CT or MRI). Eleven cats were euthanized, died or lost to follow up. The most successful treatment in the 17 surviving cats with PDH was trilostane (n = 9) for which survival ranged from 0.5 to 21 months.

The DST is the initial test of choice for diagnosis of feline HAC. Abdominal ultrasonography and cranial imaging (CT or MRI) are accurate for differentiation of ADH from PDH. Trilostane is effective for treatment of feline PDH. Hyperosmolality occurs in many critical illnesses. Intracellular hyperosmolality is a compensatory response to serum hyperosmolality and a rapid drop in serum osmolality during treatment leads to tonic fluid shifts and subsequent cellular edema.

Spurious elevation of erythrocyte mean corpuscular volume (MCV) on automated cell analyzers is a well characterized lab error that occurs in hyperosmolar patients when erythrocytes acclimated to the hyperosmolar patient serum swell when placed in isotonic media for analysis. A difference between the automated and manual MCV (dMCV) greater than 2 fl has been shown to predict hyperosmolality in humans. The purpose of this study was to investigate dMCV as a marker for serum hyperosmolality in dogs and to examine the relationship between dMCV and three methods of determining serum osmolality: measured (OsM M ), calculated (OsM C ) and calculated effective (OsM CE ) osmolalities.

OsM C , OsM CE , and dMCV were calculated from routine blood values and OsM M was measured using freezing-point depression in 121 dogs admitted to the intensive care unit. For each osmolality method, dMCVs of normosmolar (OsM < 320 mOsM) and hyperosmolar (OsM ! 320 mOsM) dogs were compared using a Student's t-test. Receiver operator characteristic (ROC) curves were generated and area under the ROC curve (AUROC) calculated for each osmolality method.

The dMCV of hyperosmolar dogs was significantly larger than that of normosmolar dogs for all three osmolality methods. dMCV predicted hyperosmolality as determined by OsM M (AU-ROC = 0.7738) better than it predicted hyperosmolality as determined by OsM C (AUROC = 0.7063) and OsM CE (AUROC = 0.7337). A dMCV cut-off of 2.96 fL yielded the best sensitivity (76%) and specificity (71%) for hyperosmolality determined by OsM M .

These data support that dMCV ! 3 fl indicates serum hyperosmolality. Because dMCV predicts measured osmolality better than it predicts either calculated method, osmoles not included in these calculations may contribute to the tonic effects of serum hyperosmolality. Pheochromocytoma (PHEO) is a rare malignant catecholamine-secreting tumor of the adrenal medulla. Catecholamines and metanephrines in plasma and in 24-h urine are approved biomarkers for the detection of the disease in humans, however, the question which of the tests is best is controversial. We previously demonstrated that measurement of urinary catecholamine and metanephrine to creatinine ratios is helpful for the diagnosis of PHEO in dogs and that urinary normetanephrine to creatinine ratio may be the best test to discriminate between PHEO and hypercortisolism (HC). Knowledge on plasma catecholamines and metanephrines in dogs is scarce and no comparison between urinary and plasma parameters has been performed. The objective of the study was to measure urinary as well as plasma catecholamines and metanephrines in dogs with PHEO, HC and in healthy dogs and to determine the test with the least overlap between the group.

Six dogs with PHEO, 9 dogs with HC (6 with ATH, 3 with PDH) and 10 healthy dogs were included. Urine samples were collected into HCL containing tubes to ensure a pH 2, blood samples were collected on ice, centrifuged at 4°C and immediately snap frozen in liquid nitrogen. All samples were stored at -80°C. Urinary epinephrine (U-Epi), norepinephrine (U-Norepi), metanephrine (U-Meta) and normetanephrine (U-Normeta), and epinephrine (P-Epi), norepinephrine (P-Norepi), free and total metanephrine (PF-Meta and PT-Meta) and free and total normetanephrine (PF-Normeta and PT-Meta) were analysed by HPLC. Urinary catecholamines and metanephrines were expressed as ratios to urine creatinine concentrations. Data were analysed by non-parametric tests (P < 0,05). Similar to our previous findings U-Epi, U-Norepi, U-Meta and U-Normeta were significantly higher in dogs with PHEO and U-Norepi and U-Normeta were significantly higher in dogs with HC compared to healthy dogs. Comparison between dogs with HC and dogs with PHEO revealed significantly higher U-Meta and U-Normeta in the latter group. U-Normeta was the only parameter with no overlap. In dogs with PHEO P-Norepi, PF-Meta, PT-Meta, PF-Normeta, PT-Normeta were significantly higher and in dogs with HC P-Norepi, PF-Normeta and PT-Normeta were significantly higher than in healthy dogs. Comparison between dogs with HC and dogs with PHEO showed significant higher PF-Meta, PT-Meta, PF-Normeta, PT-Normeta in the PHEO group. Overlap was present with all 4 parameters, but was least with PF-Normeta and PT-Normeta. According to our results U-Normeta, PF-Normeta and PT-Normeta are valuable parameters for the diagnosis of PHEO, so far U-Normeta performed better than the plasma parameters.

( Acromegaly is increasingly recognized as a cause of insulinresistance in diabetic feline patients. This study was designed to describe the sonographic changes in the abdominal organs of acromegalic cats. Cats were included if they presented to North Carolina State University or Colorado State University from January 2002 to October 2012 with poorly controlled diabetes mellitus, IGF-1 concentrations > 100 nmol/L and had an abdominal ultrasound examination (AUS) performed with report available. A control group included age-matched cats that had an AUS performed for investigation of disease unlikely to affect liver, kidneys, pancreas or adrenal glands (e.g. lower urinary tract disease).

Twenty five cats were included in each group. IGF-1 concentrations in the acromegaly group ranged from >148 to 638 mmol/l. Median left and right kidney length were significantly greater in the acromegaly group compared to controls (acromegaly-left: 47.0 mm; control-left: 38.1 mm; p < 0.0001; acromegaly-right: 47.0 mm; control-right: 42.2 mm; p = 0.0003). Hepatomegaly and bilateral adrenomegaly were reported in 63% and 53% of acromegalic cats respectively, and in none of the controls. Median left and right adrenal width were significantly greater in the acromegaly group compared to controls (acromegaly-left: 5.4 mm; control-left: 3.5 mm; p < 0.0001; acromegalyright: 5.4 mm; control-right: 3.6 mm; p < 0.0001). Median pancreatic thickness was significantly greater in acromegalic patients compared to controls (13.5 mm vs. 6.1 mm; p = 0.0003). Pancreatic changes were described in 79% of the acromegalic cats and 9% of the controls. These findings indicate that compared to non-acromegalic cats, acromegalic patients have larger kidneys, liver, adrenals and pancreas. Hyperthyroidism is a common feline endocrinopathy. However, prevalence varies widely with geographical location. Anecdotal reports suggest this disorder is rare within the Irish feline population. The aim of this study was to document the prevalence of hyperthyroidism in geriatric cats in the greater Dublin area of Ireland and to assess risk factors for development of the disease.

Practitioners within the study area were requested to select cats presenting to their clinic aged 10 years or older, in which blood sampling was being performed for geriatric health screening or clinical investigation purposes, independent of suspected thyroid status. Serum samples were submitted to University Col-lege Dublin Diagnostic Endocrine Laboratory for total thyroxine (T4) measurement by a chemiluminescent method (Canine Total T4, Immulite 1000, Siemens). Cats were classified as hyperthyroid, equivocal or euthyroid based on a total T4 concentration of > 60 nmol/L, 30 -60 nmol/L or < 30 nmol/L, respectively. Repeat measurement of total T4 after 4-6 weeks, or free T4 by equilibrium dialysis was recommended in all equivocal cases. Animals receiving treatment for hyperthyroidism were excluded. In order to fulfil the study criteria a questionnaire completed by the client and veterinarian detailing historical and physical information was requested with each submission. Associations between different categorical variables were analysed by Chisquare or Fisher's exact test. Total T4 concentrations in hyperthyroid animals reported to have palpable or non-palpable goitre were compared using the Mann-Whitney U test. A P-value of < 0.05 was considered statistically significant.

A response rate of 77% from 45 targeted practices was achieved. Samples were submitted from 508 cats; 181 male, 238 female and 89 unreported. Thyroid hormone analysis identified 107 (21%) hyperthyroid, 53 (10%) equivocal and 348 (69%) euthyroid cases. Weight loss (P < 0.0001), polyphagia (P < 0.0006) and dyspnoea (P = 0.0370) were significantly associated with a diagnosis of hyperthyroidism. Vomiting or diarrhoea was not (P = 0.684 and P = 0.2758 respectively). Cats with goitre were more likely to be diagnosed as hyperthyroid (odds ratio (OR) = 2.958, 95% CI = 1.778 -4.920) compared to those without. Tachycardia (P = 0.0018), but not the presence of a cardiac murmur (P = 0.1892) had a significant association with hyperthyroidism. Increasing age was the only significant risk factor (P = 0.0015). A relationship between sex, breed, vaccination status, parasite control, environment or preferred food flavour was not established. There was no significant difference between the total T4 concentration of hyperthyroid cats with or without reportedly palpable goitre (P = 0.3316).

Hyperthyroidism is not uncommon in Irish cats. Historical and physical examination findings including weight loss, polyphagia, dyspnoea, goitre and tachycardia are significantly associated with hyperthyroidism. Surprisingly age was the only recognised risk factor for the development of the disease. The identified but unexplained inability of the study practitioners to palpate goitre in hyperthyroid animals over a range of total T4 concentrations may explain the previously perceived low prevalence of this condition in Ireland. Ehrlichiosis is a stressful and catabolic disease with possible adrenocorticol compromising due to immunological challenges. This study aimed to evaluate serum cortisol and DHEA-S of dogs naturally infected by Ehrlichia canis, either in acute or subclinical phase, with positive Dot-ELISA and nPCR, negative for Babesia canis and Leptospira spp. (n = 21, Group 1) and compare them to healthy dogs (n = 10, Group 2).

All dogs underwent ACTH stimulation test (5 lg/kg IV). Serum cortisol and DHEA-S were assessed by radioimmunoassay and ELISA (kit In Vitro â 55060) respectively, before and one hour after ACTH administration. Cortisol and DHEA-S values were analyzed with Tukey-Kramer test and delta cortisol with Student -t test (p < 0.05).

Significant difference in cortisol levels was noticed before and post-ACTH in Group 1 (28.78 ng/mL; 143.80 ng/mL; p < 0.0001) and 2 (38.52 ng/mL; 165.72 ng/mL; p < 0.0001). However, no significance in serum DHEA-S was noticed at any time point (Group 1 0.064 lg/mL; 0.071 lg/mL; p > 0.05; Group 2 0.021 lg/mL; 0.023 lg/mL; p > 0.05). No difference was detected in cortisol before and post-ACTH or in delta cortisol between both groups (p > 0.05). There was significant difference in DHEA-S between groups (p < 0.0001), three times higher in Group 1 in both moments.

The results show dogs naturally infected by E. canis have adequate cortisol secretion and adrenocortical competence, illustrated by delta cortisol values. Increased DHEA-S concentrations in dogs with E.canis show the stress involved in the disease, once this hormone tends to increase its concentration in these circumstances. Hypoadrenocorticism in both dogs and humans is characterised by corticosteroid deficiency requiring lifelong hormone therapy. In contrast to humans, the pathogenesis in dogs is not well characterised, although autoimmune mechanisms are suspected. Approximately 45% of humans with Addison's disease as part of autoimmune polyglandular syndrome type 1 (APS1) and APS2 with premature ovarian failure (POF) have autoantibodies to the P450 side-chain cleavage enzyme (P450scc). The current study aimed to establish the prevalence of P450scc autoantibodies in dogs with recently diagnosed hypoadrenocorticism.

The coding region of canine P450scc was amplified, cloned and S 35 radiolabelled recombinant protein expressed by in vitro transcription and translation; an autoradiogram showed the protein was of expected size. A radioimmunoprecipitation assay was validated using human sera of known P450scc autoantibody status. Sera from 32 dogs with no history of endocrine or immune-mediated disease were used to establish a threshold for positivity (mean counts per minute + 3 standard deviations). Sera from dogs with hypoadrenocorticism taken within 3 months of diagnosis were then tested to investigate the presence of autoantibodies.

Twenty of 48 cases (42%) were positive for P450scc autoantibodies. Breeds with three or more dogs represented were German shepherd dog, 3 positive, 2 negative; Labradoodle, 1 positive, 3 negative; crossbred, 4 positive, 0 negative; bearded collies, 2 positive, 1 negative; jack russell terrier, 1 positive, 2 negative; standard poodle, 0 positive, 3 negative; Staffordshire bull terrier, 1 positive, 2 negative. Positive dogs had a mean age of 4y1 m and comprised 9 females and 11 males and were not significantly different to the negative population of dogs with hypoadrenocorticism.

This cross-sectional study establishes a prevalence estimate for P450scc autoantibodies in dogs with recently diagnosed hypoadrenocorticism which is similar to that seen in human patients with APS1 and APS2 with POF. These findings help confirm an immune-mediated component to canine hypoadrenocorticism and identify P450scc as a potential autoantigen. Dysregulation of adipokines is well known to be associated with visceral obesity; however, the literature contains no information regarding circulating levels of adipokines in dogs with naturally occurring hyperadrenocortism (HAC). The objective of this study was to evaluate the relationship between circulating adipokines and cortisol concentrations in dogs with pituitary-dependent HAC (PDH). A total of 22 dogs were enrolled in this study; all the dogs were overweight according to their body condition score. Group A comprised seven healthy dogs and Group B comprised 15 client-owned dogs diagnosed with PDH. The Group B dogs were treated with trilostane, and assessed before and after the treatment.

The serum leptin and insulin concentrations in Group B were significantly higher than those of Group A, but there were no significant differences in the levels of adiponectin, resistin, tumor necrosis factor (TNF)-a, interleukin (IL)-1b, IL-6, IL-10, or IL-18. The serum cortisol concentrations positively correlated with leptin concentrations; the serum leptin concentrations posi-tively correlated with insulin concentrations. There were significant decreases in the levels of leptin, insulin, and cortisol after the treatment with trilostane; however, the mean leptin value in Group B after the treatment was still higher than that of Group A. This study revealed that the up-regulation of circulating leptin level in dogs with PDH might directly correlate with hypercortisolemia. In addition, these findings indicate that the dysregulation of adipokines is likely to be associated with complications in dogs with uncontrolled HAC. Chronic hypercortisolemia may primarily result in clinical signs and organs damages in consequence of glucocorticoid effects, and in kidneys it may cause the increase of glomerular filtration rate as well as of glomerular permeability, mostly causing renal proteinuria which is characterized by urinary proteins loss of high molecular weight proteins (HMWP > 60 kDa~albumin). However tubular damage may also be involved and the impairment of tubular reabsorption of filtered proteins is characterized by loss of low molecular weight proteins (LMWP). The aim of this study was to evaluate, sequentially, the urinary proteins patterns in dogs with pituitary dependent HAC, before and during the follow-up treatment with trilostane; the hypothesis is whether hypercortisolemia may be associated to the development of renal damage. Ten dogs (6 to 13 y-old) with HAC (diagnosis based on low-dose dexamethasone suppression or ACTH-stimulation tests and abdominal ultrasound) were studied and data were recorded before (T0) and during 60 (T1), 120 (T2) and 180 days (T3) of treatment. None of the HAC dogs had or developed arterial hypertension and/or azotemia. Exclusion criteria were any concomitant disease that could cause secondary renal disease or preor post-renal proteinuria. Control group was composed of 14 clinically normal adult dogs (punctual observation). Six HAC dogs had normal urinary protein-to creatinine ratio (Coomassie blue) (UPC < 0.5) before and during all the period of treatment and developed with good control. Four out of 10 HAC dogs showed increased UPC since before (ranging from 0.7 to 15.1) and along the treatment and had poor control. Qualitative evaluation of proteins (SDS-PAGE) revealed that all HAC dogs, before and during the course of the treatment, showed higher percentage of LMWP (T0 = 73.3 to 94.2% of total proteins; T1 = 61.2 to 100%; T2 = 59.3 to 100%; T3 = 60.0 to 100%) in comparison to clinically normal dogs that had equivalent percentage of HMWP (44.6%AE 12.5; meanAESEM) and LMWP (55.4%AE12.5) (unpaired t test; p < 0.05). Moreover, higher number of proteins bands of LMWP was also detected in all HAC dogs during follow-up, most variation of 5 to 8 bands, while control dogs had scare or 3 to 4 bands. Thus, the qualitative longitudinal evaluation of urinary proteins suggested that hypercortisolemia may be more associated to the development of tubular damage than glomerular, and that tubular alteration persisted during treatment of the period of this study, for dogs with good or poor control. Even for those four dogs with poor control that had increase of UPC, urinary LMWP predominated showing that qualitatively evaluation of proteins could provide more information regarding to the location of renal damage, and then adding more information for the better understanding of the effect of cortisol on kidneys. Hyperadrenocorticism (HAC) as a possible cause of a gallbladder (GB) mucocele has been recently described in dogs; however, there is currently a lack of evidence regarding the association between the two disorders. Reduction of circulating cholecystokinin (CCK) levels has been suggested to contribute to the development of gallbladder disorders in humans. The objective of this study was to examine the serum CCK concentrations in dogs with pituitary-dependent HAC (PDH).

A total of 28 dogs were enrolled in the study. Fourteen healthy dogs were divided into two groups: seven dogs without GB sludge (Group A) and seven dogs with GB sludge (Group B). The 14 client-owned dogs diagnosed with PDH; they were divided into two groups: eight dogs with GB sludge (Group C) and six dogs with a GB mucocele (Group D). The concentrations of serum CCK were serially determined during four hours following a high-fat meal. The concentrations of serum leptin, adiponectin, resistin, and insulin were also measured. The fasting serum CCK concentrations in Group C were significantly lower than those of Groups A and B; however, no significant differences in postprandial CCK concentrations were found between groups A, B, and C. Moreover, in Group D, the fasting serum CCK concentrations were significantly higher than those of Group C. In Group D, the levels of serum insulin but not leptin, adiponectin and resistin were significantly higher than those of Group C. This study found no association between PDH and a GB mucocele related to serum CCK concentrations in dogs. Glucose tolerance tests are used in humans to diagnose prediabetes and diabetes, and are based on glucose concentration at 2 h following glucose administration. One in 150-400 domestic cats develop diabetes analogous to human type 2 diabetes, but tests for prediabetes and diabetes are not well characterized or commonly utilized in clinical practice. Thus, cats are usually not diagnosed until clinical disease is evident. Data suggests that dosing glucose per kg confounds glucose tolerance test results in obese dogs, causing higher peak (2 min) glucose concentrations, and subsequently higher values for glucose area under the curve, and higher absolute glucose concentrations. As the volume of distribution of glucose does not increase linearly with increasing fat mass, when administering intravenous glucose at a dose based on bodyweight, overweight and obese cats are potentially overdosed in comparison with lean cats, which may lead to a false positive assessment of impaired glucose tolerance. The aim of this study was to investigate changes induced by obesity in measures of glucose tolerance: glucose concentrations, T ½ and time to return to baseline, to inform development of a standardized test for glucose intolerance in lean and obese cats.

Data from frequently sampled iv glucose tolerance tests and insulin sensitivity tests of healthy neutered cats (n = 16; 6 males;10 females) between 1 and 5 years of age were analysed retrospectively before and after cats were fed ad libitum for 9-12 mths to promote weight gain.

Bodyweight and body condition score (BCS) were positively correlated with peak and 2-min glucose concentrations, 2-h glucose concentrations, and T ½. Mean 2-h glucose differed significantly between lean and obese cats. In lean and obese cats, there was a direct causal effect of 2-min glucose on 2-h glucose. 2-min glucose increased 20.5 mg/dL (1.14 mmol L) for every extra unit of BCS above 5/9, and 26.6 mg/dL (1.48 mmol/L) for every additional kg weight gain above initial weight. For every 18 mg/dL (1 mmol/L) increase in 2-min glucose, 2-h glucose increased by 1.6 mg/dL (0.09 mmol/L) when data was adjusted for body condition score. T1/2 was not affected by peak glucose concentration. Dose of glucose (0.3 vs 0.5 g/kg bodyweight) was positively correlated with 2-min glucose. Equations were developed to adjust either glucose dose or 2-hour glucose concentration to compensate for dosing per kg in obese cats. For every BCS unit increase above 5/9, measured 2 h glucose should be adjusted down by 1.62 mg/dL (0.09 mmol/L), or glucose dose of 0.5 g/kg reduced by 0.05 g/kg/unit BCS increase.

In conclusion, dosing glucose IV on a per kg basis in obese cats results in a small increase in glucose concentration at 2 h, and could lead to some cats being incorrectly classified as having mild impaired glucose tolerance. This finding is important for establishing appropriate reference ranges for a simple glucose tolerance test to evaluate glucose tolerance in overweight and obese cats.

ADRENAL GLAND ULTRASONOGRAPHY IN DOGS WITH HYPOADRENOCORTICISM. R Lobetti 1,2 , E Lindquist 2,3 , J Frank 2,3 , D Casey 2 , K Marek 2 , T Timon 2 . 1 Bryanston Veterinary Hospital, Box 67092, Bryanston, South Africa, 2 SonoPath, NJ, 3 Sound Eklin New Jersey Mobile Associates, NJ.

Hypoadrenocorticism can be a life-threatening disease if not treated immediately. Although a tentative diagnosis can be made on clinical signs and laboratory findings, a definitive diagnosis can only be made on an ACTH stimulation test. Unfortunately typical clinical signs and laboratory findings are not evident in all cases and ACTH stimulation test results are usually not immediately available. As abdominal ultrasonography is widely used, it would be ideal as a diagnostic aid for hypoadrenocorticism. To date there are only 2 studies that have shown small adrenal glands in dogs with hypoadrenocorticism on ultrasound.

The purpose of this study was to identify a reliable set of adrenal ultrasonography parameters that could be used to identify dogs with hypoadrenocorticism.

The records of 81 privately owned dogs that had abdominal ultrasonography done as well as an ACTH stimulation test were retrospectively evaluated. The dogs were divided into three groups: Group 1 consisted of 37 dogs with clinical signs and/or a sonogram appearance of their adrenal glands suspicious of hypoadrenocorticism and confirmed on an ACTH stimulation test. Group 2 consisted of 19 dogs with clinical signs and/or a sonogram appearance of their adrenal glands suspicious of hypoadrenocorticism but ruled out by a normal ACTH stimulation test. Group 3 consisted of 25 dogs that had no clinical signs or biochemical evidence of hypoadrenocorticism, normal sonogram appearance of their adrenal glands, and a normal ACTH stimulation test. Descriptive statistics were used to describe the data and one-way analysis of variance with Bonferroni and Tukey-Kramer comparisons used to test for statistical differences between the groups. The level of significance was set at p < 0.05.

Results showed that the median right adrenal length in Group 1-3 was 1.75 cm, 1.8 cm, and 2.03 cm, respectively. Median left adrenal length in Group 1-3 was 1.77 cm, 2.08 cm, and 2.1 cm, respectively. There was no statistical difference between the right and left adrenal gland and within groups. Median right adrenal thickness in Group 1-3 was 0.34 cm, 0.37 cm, and 0.6 cm, respectively. Median left adrenal thickness in Group 1-3 was 0.31 cm, 0.4 cm, and 0.6 cm, respectively. In both right and left measurements, groups 1 and 2 were statistically different from group 3 but there was no statistical difference between groups 1 and 2.

The study concluded that the ultrasound finding of small, flattened, isoechoic adrenal glands should be an alert for possible hypoadrenocorticism, prompting additional confirmatory function testing and/or therapeutic intervention. Iatrogenic hypothyroidism following treatment of feline hyperthyroidism can have deleterious effects on renal function. Serum total thyroxine concentration (T4) is commonly used to evaluate therapy, but no study has examined the use of both serum free thyroxine by equilibrium dialysis (FT4ed) and thyroid-stimulating hormone (TSH) concentrations. The purpose of this study was to compare the ability of T4, FT4ed, and TSH concentrations to diagnose treatment-induced hypothyroidism in hyperthyroid cats receiving methimazole. We hypothesized that FT4ed would identify more cats with iatrogenic hypothyroidism as compared to T4.

A total of 65 samples from previously-diagnosed hyperthyroid cats receiving methimazole therapy and with T4 concentrations < 48 nmol/L were included. Samples had been submitted to the diagnostic laboratories at Auburn University (n = 22) and Michigan State University (n = 43). T4, FT4ed and TSH concentrations were measured via assays previously validated for use in cats. Correlation was tested via a Spearman Rank Order test. Significance was set at the p < 0.05 level.

The median (range) T4, FT4ed, and TSH concentrations were 20 (3-48) nmol/L, 20 (3-57) pmol/L, and 0.1 (.08-9.1) ng/ml, respectively. Overall, 23 cats (35%) had an elevated TSH concentration (>0.30 ng/ml). For cats with elevated TSH concentrations, median T4 and FT4ed concentrations were 11 (3-40) nmol/L and 12 (3-35) pmol/L, respectively. Cats with normal TSH concentrations (<0.30 ng/ml) had median T4 and FT4ed concentrations of 23.5 (5-48) nmol/L and 27.0 (6-57) pmol/L, respectively. The percentage of cats with an elevated TSH concentration in combination with a low T4 (<10 nmol/L), FT4ed (<10 pmol/L), or both T4 and FT4ed concentration were 17%, 12% and 11%, respectively. Eleven cats (17%) had an elevated TSH despite normal T4 and FT4ed concentrations. One cat (1.5%) had a normal TSH despite low T4 and FT4ed concentrations. Of 24 cats with T4 concentrations between 10-25 nmol/L (low end of the reference range 10 nmol/L), 7 (29%) had an elevated TSH. Of 26 cats with FT4ed concentrations between 10-25 pmol/L (low end of the reference range 10 pmol/L), 11 (42%) had an elevated TSH. A significant positive correlation was found between T4 and FT4ed concentrations (p = <0.001). A significant negative correlation was found between both T4 and TSH concentrations (p = < 0.001) and between FT4ed and TSH concentrations (p = <0.0001).

The data suggest that FT4ed does not identify more cats with iatrogenic hypothyroidism as compared to T4. As some cats had an elevated TSH concentration despite having a normal T4 or FT4ed, further investigation may be warranted. The PFCs have been used in a wide range of consumer, including residential, products (e.g., stain-resistant coatings for carpets and upholstery). Carbon-fluoride bonds are highly stable, making PFCs extremely resistant to biodegradation. Thus, PFCs have become globally distributed and are ubiquitously present in serum of wildlife and people. Despite this, comparatively little is known as to how people are primarily exposed, and what (if any) health risk is associated with chronic, low-level exposure. It is hypothesized that house dust may represent a significant exposure route because PFCs can slough or volatilize from products used indoors, subsequently adsorbing to and accumulating within house dust. The purpose of this study was to determine if PFC serum levels in domestic cats tended to increase in proportion to time spent indoors and whether analyte patterns reflected that of food sources [e.g., fish products with high perfluoro-octane sulfonate (PFOS) but low perfluorohexanesulfonate (PFHxS)] or with house dust (PFOS + PFHxS + perfluorooctanoate (PFOA) ─ with high PFHxS levels in the most contaminated dust).

In 2008, serum was obtained from feral and pet cats presenting to shelters and clinics in the Raleigh, NC area, including the NCSU VTH. PFC serum levels were measured using high-resolution time-of-flight mass spectroscopy. Data on housing status was available for 50 cats. From least to greatest indoor residential exposure, cats were grouped as follows: (a) feral cats (n = 5), (b) pet cats living outdoors 40-75% of the time (n = 8), (c) a resident shelter cat (n = 1), (d) pet cats living indoors ! 90% of time (n = 4), and (e) pet cats living exclusively indoors (n = 32).

Total PFC (ng/mL) serum (mean; min. & max.) concentrations were: (a) 12.9 (2.9, 24.2), (b) 17.8 (4.4, 33.1), (c) 31.7, (d) 37.0 (4.6, 63.9), and (e) 51.1 (0.5-376). In feral cats, PFC levels and analyte patterns were not unlike that reported for wild felidae, with PFOS predominating. On the other hand, for cats living largely (90-100%) indoors, PFOS+PFOA+PFHxS was typically detected, with PFHxS the predominate analyte in cats with the highest total PFC concentrations.

Results indicate that house dust is a major exposure route for PFCs in indoor cats likely due to prolonged contact with carpeting and upholstery and to dust ingestion via grooming. Dust may also be an important exposure source for humans, especially children, who likewise spend time playing on floors and engage in hand-mouth transfer of dust. Hence, cats may be ideal sentinels to better assess PFC exposure risk in children. Studies are in progress to assess whether the differences in PFC serum levels or patterns in these cats may have been associated with specific clinical abnormalities or disease syndromes. (This abstract do not reflect US EPA policy). The combination of prednisone and cyclosporine has become a common treatment for immune-mediated conditions. Both prednisone and cyclosporine have been shown to have an influence on insulin release and effects. Anecdotally, we have noted a higher incidence of hyperglycemia in dogs treated with this drug combination. The aim of this study was to determine if there is a difference in serum glucose concentrations in dogs being treated with immunosuppressive doses of prednisone and cyclosporine versus prednisone alone, or in combination with other immunosuppressive medications.

The Texas A&M University medical database was searched for dogs diagnosed with immune-mediated thrombocytopenia and/or immune-mediated hemolytic anemia from 2010-2012. The signalment, weight, immunosuppressant drugs used and doses were recorded. The highest serum glucose concentration after starting the medication(s) was also recorded. Cases were excluded if there was no follow-up. Cases were divided into two groupsthose receiving prednisone and cyclosporine and those receiving prednisone alone or in combination with another drug (azathioprine or mycophenolate). Data was checked for normality using a D'Agostino & Pearson omnibus test. A Mann-Whitney test and an unpaired t test was used to compare glucose concentrations between the groups and a Fisher's exact test was used to compare proportions of dogs with serum glucose concentrations above or below the reference interval.

In total, 40 cases fulfilled the inclusion criteria. There were 27 dogs that received prednisone and cyclosporine, and 13 dogs that received prednisone alone or in combination with another drug. Median serum glucose concentration was 146 and 112 mg/dl respectively, and this difference was statistically significant (P = 0.0005). Dogs that received prednisone and cyclosporine were 17.45 (95%CI: 1.9-154.4) times more likely to have a recheck serum glucose concentration above the reference interval (60-135 mg/dl) than dogs that received prednisone with or without another medication. Within the prednisone and cyclosporine group, dogs under 10 kg had a significantly increased mean serum glucose concentration compared to those greater than 10 kg (217.2 vs. 143.6 mg/dl; P = 0.0243). There were no associations between doses of drugs and serum glucose.

In conclusion, dogs (especially those under 10 kg) undergoing treatment with immunosuppressive doses of prednisone and cyclosporine had increased serum glucose concentrations compared to those that received prednisone alone or with another immunosuppressive medication. Further studies are warranted. Humans with fasting glucose above normal, but below diabetic, are classed as having impaired fasting glucose. Impaired glucose tolerance is diagnosed based on increased glucose concentration at 2 h after oral or iv glucose administration in a standardized test. Humans with impaired fasting glucose or impaired glucose tolerance below levels considered diabetic, are classed as prediabetic, and at high risk of developing type 2 diabetes. Human prediabetics outnumber diabetics 3-4:1. We have previously reported the upper cutpoint for casual blood glucose in cats, but tests for pre-diabetes and subclinical diabetes in cats are not well characterized, and therefore, cats are not typically diagnosed until clinical diabetes is evident.

The aims were to establish cutpoints for healthy neutered cats ! 8 years of age for fasting and 2 h glucose using a standardized test protocol with paw or ear samples and a portable glucose meter calibrated for feline blood. All cats were client-owned and healthy on the basis of client history, physical examination and a routine blood profile. Of the 82 cats tested (aged 8-18 years), 21 were Burmese and 61 non-Burmese (22 lean-BCS 3-5/9), 20 overweight-BCS 6-7/9; and 19 obese-BCS 8-9/9). Following ! 18 h fast, a catheter was inserted into the cephalic vein. After 3 h, fasting glucose was measured from the ear or paw using the Abbott AlphaTRAK. Glucose (0.5 g/kg bwt) was administered i.v. over 30s and glucose measured at 2 min and 2 h. Reference intervals were determined after Box-Cox transformation and exclusion of outliers. The cutpoints were defined as the upper limits of the 95% reference intervals. Based on a priori knowledge that overweight and obese cats have abnormal glucose tolerance, cats of BCS 7-9/9 were excluded from the fasting and 2 h reference interval calculations. Reference intervals for Burmese were pooled with non-Burmese because the percentage differences of the medians and interquartile ranges for the sub-groups were 50% and 100%, respectively. Based on the 95% reference interval, the fasting glucose cutpoint for cats with BCS 6/9 (n = 44) was 6.3 mmol/L (113 mg/ dL); the associated 90% confidence interval was 6.1-6.5 mmol/L (110-117 mg/dL). 2/82 cats were classed as having impaired fasting glucose (BCS 5 and 7/9). The cutpoint for 2 h glucose established using cats with BCS 6/9 was 10.0 mmol/L (180 mg/dL) (90% confidence interval 9.1-10.8 mmol/L (164-194 mg/dL). Six of 82 cats were classed as having impaired glucose tolerance (4 with BCS 8 or 9/9 including 1 Burmese, 2 with BCS 7/9). We recommend that 6.3 mmol/l (113 mg/dL) be used as the cutpoint between normal and impaired fasting glucose, and that 10.0 mmol/L (180 mg/dL) be used as the 2-h glucose cutpoint between normal and impaired glucose tolerance in a simplified intravenous glucose tolerance test using a glucose dose of 0.5 g/kg with blood glucose measured from ear or pad samples using a portable glucose meter calibrated for feline blood. Type 1 diabetes mellitus (T1DM) is a frequently diagnosed endocrinopathy in dogs with an increasing prevalence, challenging life-long management and frequent complications. Pancreatic islet transplantation is the only non-invasive, curative treatment for T1DM. A first step in the application of pancreatic islet transplantation in veterinary patients is to develop ethically acceptable protocol for islet isolation using readily available laboratory equipment. The goal of this study was to develop a canine pancreatic islet isolation method yielding sufficient islet mass of high purity from canine cadaveric donors.

Pancreata were procured from dogs euthanized for reasons unrelated to this study. Initial anatomic studies were performed to evaluate efficacy of pancreatic perfusion. Collagenase digestion was performed using a Ricordi chamber and temperaturecontrolled perfusion circuit. Islets were separated from the exocrine tissue using a discontinuous density gradient and a standard laboratory centrifuge. After isolation, islet yield was calculated and viability was assessed using dual-fluorescent staining techniques.

Islet isolation was completed in 6 dogs. Median (interquartile range) islet yield was 36,756 of islet equivalents (IEQ) per pancreas (28,527 IEQ). A high degree of islet purity [87.5% (10%)], and viability [87.4% (12.4%)] were achieved. The islet yield achieved using this technique would require approximately 1 pancreas per 5 kg body weight of the recipient dog. Purity and viability of the isolated islets were comparable to those achieved in human islet transplant programs and would be sufficient for clinical transplantation. Based on initial results, clinically relevant islet yield and quality can be obtained from canine cadavers using standard laboratory equipment. Total serum osmolality is comprised of effective osmoles, those solutes that exert a tonic effect and cause water to shift extracellularly, and ineffective osmoles, those solutes that do not exert a tonic effect. In health, the major contributors to osmolality include the monovalent ions, glucose, and urea. In certain disease states, the concentrations of various organic anions such as lactate, acetoacetate (AA), and b-hydroxybutryrate (BHB) rise and increase serum osmolality. However, it is unclear whether these solutes exert a tonic effect and contribute to serum tonicity. The purpose of this study was to determine whether lactate, AA, and BHB act as effective or ineffective osmoles in a canine red blood cell (RBC) model.

Tonic effects of various solutes on RBCs were determined using an automated cell counting device (Scepter2.0, Millipore) to detect changes in RBC diameter (dRBC). RBCs were washed and acclimated in 300mOsM sodium chloride (NaCl) for 10 minutes. RBCs were then added to control or test solutions and incubated for 30 minutes prior to dRBC measurement. First, 50 mM of known effective (glucose) and ineffective (urea) osmoles were added to separate 300mOsM NaCl solutions and dRBCs compared to a control (300mOsM NaCl). Next, the tonic effects of organic anions were examined: dRBC of chloride (Cl), BHB, and lactate were compared in 300mOsM solutions of sodium (Na) salts; dRBC of Cl, AA, and lactate were also compared in 300mOsM solutions of lithium (Li) salts. The Wilcoxon rank test was used to compare dRBC between groups (a<0.05; n = 15). dRBC in urea was larger than in control (p = 0.014); dRBC in glucose was smaller than control but this difference was not statistically significant. Thus, ineffective osmoles cause an increase in dRBC and effective osmoles do not. For the organic anions, dRBC was larger in NaBHB (p < 0.001) and smaller in Na-lactate (p < 0.001) when compared with NaCl control. dRBC was larger in LiAA (p < 0.001) and Li-lactate (p = 0.003) when compared with LiCl control. In this model, BHB and AA act as ineffective osmoles and do not exert a tonic effect. In contrast, the tonic effect of lactate appears to be dependent on the associated cation. Diabetic nephropathy has become the most important cause of end-stage renal disease in human. Chronic hyperglycemia is the main component in the development of complications along the course of the disease. Long term complications are common in humans, such as diabetic nephropathy (DN), characterized mainly by glomerular lesions and proteinuria of loss of high molecular weight proteins (HMWP > 60 kDa~albumin). Initial changes include glomerular hyperfiltration and renal hyperperfusion. Morphological changes also may occur including thickening of the glomerular basement membrane, glomerular hypertrophy, mesangial and tubulointerstitium expansion. However, tubular damage may also be involved resulting in loss of low molecular weight proteins (LMWP < 60 kDa). In dogs, there are a few studies that investigated the relationship between diabetes mellitus (DM) and renal injury. The aim of this study was to sequentially evaluate the urinary proteins, quantitatively through urinary protein-to creatinine ratio (UPC; Coomassie Blue) and urinary albumin-to creatinine ratio (UAC; ELISA Immunology Consultants Laboratory) and qualitatively by SDS-PAGE during the course of DM in dogs, and to identify the segments of the nephrons that could be involved. Ten normotensive dogs with DM (age from 8 to 13 yr-old) had good fasting glycemia (252.9 AE 22.6 mg/dL; mean SEM) and were followed-up for 6 months (T1 to T6). Exclusion criteria were any other concomitant diseases that could cause renal secondary damage or pre-or post-renal proteinuria. Control group was composed of 14 clinically normal adult dogs (punctual observation). All diabetic dogs had UPC < 0.25 throughout the course (ranging from 0.07 to 0.25) and UAC ranged from 0.0 to 0.000801 (microalbuminuria = UAC 0.03 to 0.3). Thus, all diabetic dogs had normal values of UPC and UAC, and SDS-PAGE revealed that those diabetic dogs, in comparison to clinically normal dogs, had equivalent percentage of LMWP proteins (kruskal-wallis; p > 0.05) (diabetic dogs: T1 = 55.4 AE 8.0%; T2 = 61.9 AE 8.8%; T3 = 59.7 AE 10.5%; T4 = 67.0 AE 8.1%; T5 = 64.3 AE 4.6%; T6 = 52.4 AE 10.8%) with variation of 1 to 4 bands, while the control group had 55.4 AE 12.5% of LMWP (ranging from 19.7 to 64.7%) and 3 to 4 bands. Thus, the sequential evaluation of urinary proteins suggested that the diabetic dogs did not develop tubular or glomerular damage during this study. Furthermore, the good glycemic control may have contributed to prevent DN in dogs. Moreover, further studies in larger number of cases and longer period of observation should be conducted for the better understanding of mechanisms underlying renal injury that may be involved in DM dogs. Central diabetes insipidus (CDI) is one of many differentials for polydipsia and polyuria in cats. Response to synthetic AVP, desmopressin, is used to support a diagnosis of CDI but does not rule out psychogenic polydipsia. Etiology of CDI may be attributed to pathology of the hypothalamus or posterior pituitary or remain unknown. Over 50 mutations in arginine vasopressin (AVP) have been reported in humans with CDI. The objective of this study was to determine if mutations in the feline AVP gene are present in cats with juvenile-onset CDI.

Three cats with a diagnosis of CDI were included based on onset of clinical signs by six months of age, absence of renal disease and, in two cases, positive response to desmopressin therapy. Controls were chosen based on normal urine concentration. The cDNA sequence for feline AVP was first determined using 3'-rapid amplification of cDNA ends and used to design primers for PCR from genomic DNA. The entire feline AVP gene including three exons and two introns was amplified from three CDI and 10 normal cats and sequenced bidirectionally.

One of the three CDI cats was found to have a significant disruption to the 5' half of the AVP gene, suggesting absence of gene expression. Another CDI cat has an 84-base deletion in intron 2 that is also present in one of the normal cats, thus the clinical significance is unclear. AVP mutations do occur in cats with CDI and may have diagnostic potential. Primary idiopathic hypertriglyceridemia of Miniature Schnauzer (MS) is a frequent disorder. The genetic and metabolic causes remain unknown. MS dogs with hyperlipidemia can develop pancreatitis, seizures, vacuolar hepatopathy and diabetes, therefore early diagnosis is important. This study characterized basal (fasting) and postprandial hyperlipidemia (after 4 hours) in 55 MS (27 male and 28 female) with a mean age of 5.66 AE 3.3 years. The prevalence of insulin resistance (through Homeostasis model assessment-HOMA score) in hyperlipidemic dogs was also evaluated. Overall, thirty-one dogs (56.4%) had hyperlipidemia from which 13 had hypertriglyceridemia (HTG). Eleven dogs (20%) had hypercholesterolemia (HCOL) and seven (12.7%) had both HTG and HCOL. Medium serum concentrations of triglycerides and cholesterol were 205.96 AE 313.83 mg/dL and 261.30 AE 127.18 mg/dL, respectively. A mild basal HTG (between 100-400 mg/dL) was found in 12 (21.8%) animals and 8 (14.5%) dogs presented a moderate to severe (TG > 400 mg/ dL) HTG. The postprandial evaluation revealed moderate to severe (TG > 442 mg/dL) HTG in 10 (18.2%) cases. All animals that presented high postprandial TG levels, also presented high basal TG concentrations, demonstrating a strong relationship between these two measurements (Pearson coefficient, r = 0.97). The basal hyperinsulinemia (24.7 AE 13.8 lU/mL) was identified in 45% of animals (n = 14/31) and an elevated HOMA score (6.57 AE 4.26) was found in 90.3% of cases (n = 28/31). The results from this study suggest that healthy MS have a high incidence of hyperlipidemia that is often associated with insulin resistance, shedding light on the clinical importance of this disease in this breed.

THERAPY OF CANINE HYPERLIPIDEMIA WITH BEZAFI-BRATE. VDe Marco 1 , KSM Noronha 1 , TC Casado 1 , ER Nakandakare 2 , JC Florio 2 . 1 University of Santo Amaro, 2 University of São Paulo, Brazil.

The primary and secondary hyperlipidemia are common in dogs and its treatment is necessary to prevent clinical complications such as pancreatitis, seizures, liver disease and diabetes. The therapy of mild hyperlipidemia comprising a fat restricted diet, but in more severe cases pharmacological treatment is necessary. Bezafibrate (BZF) is effective in the treatment of hypertriglyceridemia in humans, however there are no clinical studies in dogs. The objectives of this study were to assess the efficacy of BZF in reducing serum triglyceride (TG) and cholesterol (CHO) in hyperlipidemic dogs, identify a therapeutic protocol for this drug and assess possible side effects such as muscle pain, emesis, diarrhea and elevated CK and TGP levels. Only animals with moderate to severe hypertriglyceridemia (TG> 350 mg/dL) were treated with BZF every 24 hours for 30 days before introduction of any other therapy according to the protocol: ¼ tablet 200 mg for dogs weighting less than 12 kg, ½ tablet 200 mg for dogs weighing between 13 and 25 kg, 1 tablet 200 mg for dogs weighing over 25 kg. We studied 46 dogs (26 females and 20 males) with a mean age of 9 AE 3 years. Fifteen dogs (32.6%) had primary hyperlipidemia and 31 (67.4%) secondary hyperlipidemia, which included hyperadrenocorticism (41.3%), hypothyroidism (15.2%) and chronic corticoideterapia (10.8%). All 46 (100%) dogs had hypertriglyceridaemia and 33 (71.7%) had both hypertriglyceridaemia and hypercholesterolemia. After 30 days using BZF, normalization of serum TG (TG <150 mg/dL) was observed in 91.3% of cases (n = 42/46) and of CHO (CHO < 270 mg / dL) in 66 7% (n = 22/33) of cases. Means and standard deviations of serum TG and COL before (752 AE 663 mg/dL and 428 AE 217 mg/dL) and after therapy (110 AE 82 and 244 AE 71 mg /dL) were significantly lower (p < 0.005, paired Student t test). The bezafibrate dose most used with a 95% confidence interval was 5.3 to 6.1 mg/kg (range: 4-10 mg/kg). No side effects were observed and there was no statistical difference between the values of ALT and CK before and after therapy. It can be concluded that bezafibrate is a safe and effective drug for the canine hyperlipidemia therapy. Five cats with normal gastrointestinal tracts were euthanized for reasons unrelated to this study and samples from duodenum, mid-jejunum, ileum, cecum and colon were collected, processed for histology and labeled with immunohistochemical preparations for GIP, GLP-1 and GLP-2. In each cat labeled cells were enumerated by counting 15, randomly selected 400X fields from each sample and then averaged. The results are presented (Table 1) as mean (AESD) of that average for the 5 cats. Mean (AESD) per high power field (x400) of positive cells stained with anti-GIP, anti-GLP-1, or anti-GLP-2 in 5 cats with normal intestines.

In contrast to other mammals where K cells are not present in the ileum and aborally, GIP-expressing cells are abundant throughout the intestines in cats. GLP-1-expressing cells are most abundant in the ileum as in other mammals, but, in contrast, they are abundant throughout the intestines, including in the duodenum in cats. GLP-2-expressing cells do not follow the same distribution of GLP-1, suggesting more than one distinct population of L cells is present in cats. These differences suggest a different physiologic role of K and L cells in cats compared to other mammals and could be important in diseases such as diabetes and obesity. Peptide YY (PYY) and Glucagon-Like Peptide-1(7-36) amide (GLP-1) are anorexigenic enteroendocrine hormones secreted postprandially by intestinal L cells. Reports have shown that secretory patterns of both PYY and GLP-1 are altered in obese human subjects and rodent models; this supports a possible role for these hormones in regulation of insulin sensitivity. Little is known about the role of these hormones in feline obesity and diabetes.

The aim of this study was to evaluate the secretory patterns of these two hormones concurrently in healthy lean (LC) and obese cats (OC). Cats fed standardized commercially available diets were assessed to determine if body condition, sex or dietary macronutrients altered secretion.

The LC (BCS 1-5/9) group (5 males and 5 females) had a mean age of 8.7 AE 3.6, BCS 4.8 AE .4, and weight 4.4 AE .7 kg. The OC (BCS 6-9) group (6 males and 5 females) had a mean age of 6.64 AE 2.6, BCS 7.1 AE .8, and weight 6.3 AE 1.1 kg.

For ]. Blood sampling and complete physical examinations were performed at three time points; baseline (after 2 wks on high c/h diet) then at 2 and 4 wks on the high protein diet. Samples were taken after a 10 hour fast and one hour postprandially after being given 5 to 10 minutes to complete approximately 40 kcal of the appropriate soft diet. Commercially available rodent PYY and GLP-1 ELISA kits were validated for use in cats. No statistically significant changes in weight occurred in either group for the duration of the study. Plasma PYY and GLP-1 concentrations did not differ between the LC and OC groups. In LC, plasma PYY concentrations increased after a meal at baseline (Pre vs Post feeding: 73.9 AE 28.8 vs 116.4 AE 46.1 pmol/L) and 4 weeks (72.5 AE 33.9 vs 131.4 AE 63.3 pmol/L) on the high protein diet. Similarly, plasma GLP-1 concentrations increased after a meal in the LC at baseline (Pre vs Post feeding: 8.8 AE 2.2 vs 13.1 AE 2.6 pmol/L), 2 weeks (10.1 AE 2.4 vs 13.7 AE 2.7 pmol/L), and 4 weeks (10.3 AE 2.7 vs 15.5 AE 3.8 pmol/L) on the high protein diet. However, pre-and post-prandial plasma PYY and GLP-1 concentrations did not differ among OC on either the high c/h or high protein diets. Our results suggest that the expected meal-induced increase in plasma PYY and GLP-1 concentrations is attenuated in OC compared to LC. It is unknown whether a deficit in PYY and GLP-1 secretion and/or signaling plays a role in feline obesity. Although, pancreatic biopsy remains the gold standard for evaluating pancreatic pathology, it is not widely used as a diagnostic tool for acute or chronic pancreatitis in cats because of its invasiveness and because of historical concerns of inducing pancreatitis. The feline pancreatic lipase immunoreactivity (fPLI) test is the most specific and sensitive test for pancreatic inflammation currently available and is widely used for diagnosing pancreatitis. The aim of this study was to evaluate the effect of pancreatic biopsy on the serum fPLI concentrations.

Forty-four cats underwent laparotomy because of clinical signs of chronic small bowel disease. Because chronic enteritis, cholangitis, and/or pancreatitis were among the differential diagnoses, biopsies were taken of the small bowel in three or more locations, the pancreas, and the liver. Pancreatic biopsies were obtained using a 4 mm biopsy punch. The entire pancreas was inspected for areas of gross abnormality. If such an area was identified, the biopsy was collected in this area. If there were no areas of gross abnormalities, a random site on the edge of the pancreas was chosen. Hemorrhage was controlled with direct pressure in over 95% of the cases; a hemostatic powder or a single suture was used in the remaining cases. A blood sample was collected for measurement of serum fPLI concentration (as measured by Spec fPL â ) prior to surgery, 2-4 hours post-op (n = 10), and approximately 24 hours post-op (n = 44) for measurement of serum fPLI concentration. Serum fPLI concentrations were compared pre-op and post-op using a Wilcoxon matched-pairs signed rank test with a p-value of <0.05 being considered statistically significant.

Serum fPLI concentrations did not increase significantly at 2-4 hours post-op (medians: 2.2 vs 1.75 lg/L; p-value: 0.432). However, there was a significant increase in serum fPLI concentrations 24 hours after collecting the biopsy (medians: 2.2 vs. 2.9 lg/L; p-value: 0.043). When cats were divided into cats with histopathological evidence of pancreatic pathology of any kind on the collected biopsy and those that did not, only the cats without such evidence had a significant increase in serum fPLI concentrations (medians: pre-op: 1.7 vs. post-op: 2.9 lg/L; p = 0.0137). In the group of cats with pancreatic pathology, the fPLI interpretation changed from "within the reference interval" to "pancreatitis" in 1 cat (n = 18), while it changed in 3 cats without pancreatic pathology (n = 26). However, none of these cats showed clinical signs of acute pancreatitis.

Pancreatic biopsy using a 4 mm biopsy punch leads to significant increases in serum fPLI concentrations, but not to clinically overt pancreatitis. Feline pancreatitis is a diagnostic challenge and controversy exists as to the sensitivity and specificity of newer diagnostic tests. No information has been published on diagnostic utility of currently available lipase assays compared to the gold standard histopathology. While the commonly used test Spec fPL is generally regarded as most useful, it is the authors' clinical impression that the 1,2-o-dilauryl-rac-glycero-3-glutaric acid-(6′methylresorufin) ester (DGGR) lipase -a colorimetric assay used as the routine diagnostic at the authors' institution -is equally as useful. Thus the goal of this study was to compare both assays versus a standardized pancreatic histopathological assessment.

The entire pancreas of 40 cats was examined irrespective of the cause of death. Pancreata including surrounding mesentery and a portion of the duodenum were harvested within 3 hours of death. All pancreata were sectioned every 0.5 cm, stained with HE and examined microscopically by a board-certified pathologist in a standardized blinded fashion. Histologic sections were scored for lesions of acute (AP) and chronic pancreatitis (CP) with a scoring system based on previously published classification schemes. AP was defined as edema and/or necrosis and neutrophilic inflammation, CP was defined as fibrosis and mononuclear cell infiltration. Corresponding blood samples were obtained immediately prior to euthanasia. Spec fPL concentrations were measured through IDEXX Laboratories; DGGR-lipase activity was measured on a Cobas Integra 800 analyzer (Roche). Histopathologic assessment revealed mild AP (3), severe AP (2), mild CP (22), and severe CP (1). The following histopathologic findings were recorded without concurrent pancreatitis: edema (1), fibrosis (5), cystic degeneration (4), atrophy (2), neoplasia (4), nodular hyperplasia (8 mild, 1 moderate), and islet amyloidosis (5). Two pancreata were normal. Combining AP and CP, the overall sensitivity and specificity were 48 and 54% for Spec fPL, and 41 and 62% for DGGR-lipase. When considering mild CP as clinically irrelevant, the overall sensitivity and specificity for pancreatitis were 60 and 54% for Spec fPL, and 60 and 63% for DGGR-lipase. Sensitivity and specificity for all forms of AP were 50 and 54% for Spec fPL, and 50 and 62% for DGGR-lipase. Sensitivity and specificity for severe AP were 67 and 57% for Spec fPL, and 67 and 64% for DGGR-lipase. Sensitivity and specificity for all forms of CP were 48 and 54% for Spec fPL, and 39 and 62% for DGGR-lipase. Sensitivity and specificity for severe CP were 100 and 54% for Spec fPL, and 100 and 63% for DGGR-lipase. On the base of our results we conclude that test results of both lipase assays show only moderate sensitivity and specificity compared to histopathology, while the DGGR-lipase yields better specificity. Diagnosing acute pancreatitis (AP) in the dog can be challenging. The aim of this study was to determine the utility of measurement of Spec cPL â and amylase and lipase activities in peritoneal fluid as complimentary diagnostic tools in a population of dogs diagnosed with AP based on clinical signs, ultrasonographic findings, and serum Spec cPL concentrations.

Fourteen dogs with peritoneal fluid secondary to pancreatitis and nineteen dogs with peritoneal fluid secondary to nonpancreatic disease were prospectively enrolled into this study. Dogs were included into the pancreatitis group if they had at least two clinical signs consistent with pancreatitis (i.e., lethargy, inappetence, weakness, vomiting, abdominal pain, diarrhea), ultrasonographic findings considered consistent with acute pancreatitis based on images reviewed by a board-certified radiologist (i.e., pancreatic enlargement, changes in parenchymal echogenicity, irregular pancreatic margins, peripancreatic fluid, hyperechoic surrounding mesentery, corrugation of the surrounding small intestines), a serum Spec cPL concentration supportive of a diagnosis of pancreatitis (Spec cPL >400 lg/L), and peritoneal fluid. Dogs were included into the nonpancreatitis group if they had a final diagnosis of a nonpancreatic disease, a serum Spec cPL concentration that did not support a diagnosis of pancreatitis (Spec cPL < 200 lg/L), and peritoneal fluid.

Spec cPL concentration and, amylase and lipase activities in peritoneal fluid were measured and compared between the two groups. Sensitivity and specificity were calculated based on cutoff values set a-priori (serum Spec cPL; 400 lg/L) or based on a ROC curve with a goal of maximizing sensitivity, while reaching a specificity of > 80% (serum lipase and amylase activity; Spec cPL concentration and lipase and amylase activities in peritoneal fluid).

The sensitivity and specificity of peritoneal fluid Spec cPL concentration (cut-off value: 500 lg/L) were 100% and 94.7%, respectively. The sensitivities and specificities for peritoneal fluid amylase activity (cut-off value: 1000 U/L) and peritoneal fluid lipase activity (cut-off value: 500 U/L) were 71.4% and 84.2% for amylase activity, and 92.9% and 94.7% for lipase activity, respectively.

Peritoneal fluid Spec cPL concentration was sensitive and specific for diagnosing AP in dogs. Peritoneal fluid Spec cPL concentration should be considered as a complimentary diagnostic tool in dogs with peritoneal effusion suspected of having AP. Peritoneal fluid lipase activity, although not as sensitive as Spec cPL concentration, may also support a diagnosis of AP in dogs. There are strong breed predispositions with chronic pancreatitis in dogs. English Cocker Spaniels (ECS) are frequently affected and genetic predisposition is suspected in this breed. More recently, chronic pancreatitis in this breed has been associated with IgG4 + plasma cells infiltration, suggesting an immune-mediated mechanism. The purpose of this study was to investigate the association between chronic pancreatitis in ECS and the major histocompatibility complex class II loci (the dog leukocyte antigen loci DLA-DRB1, -DQA1, -DQB1).

DNA was extracted from 40 cases and 82 controls. The second exon of the canine DLA-DRB1, -DQA1 and -DQB1 was amplified by PCR and purified. All fragments were sequenced and the sequences obtained were identified by comparing them to a canine genotype databse.

Haplotypes distribution was found to be significantly different between cases and controls with higher frequency of the haplotype DLA-DQB1*00701 among cases, and the lower frequency, compared to controls, of the DLA-DQB1*02001 haplotype.

These results confirmed DLA haplotypes are a contributory factor in the development of chronic pancreatitis in ECS. These results parallel the results of previous study indicating that DLA-DQB1*00701 was found more frequently in ECS affected by anal sac adenocarcinoma and primary immune-mediated haemolytic anaemia. This may indicate a predisposition of ECS for a multi-systemic immune-mediated disease, similar to IgG4 + related disease described in human patients. Paired with the measurement of the pancreas-specific lipase (Spec fPL), abdominal ultrasonography (AUS) is felt to be a key diagnostic test in cats with suspected pancreatitis. However no information is available on how Spec fPL results compare with ultrasonographic findings compatible with pancreatitis. Therefore the purpose of this study was to evaluate the agreement of Spec fPL results and ultrasonographic pancreatic findings in cats with suspicion of pancreatitis examined at a veterinary teaching hospital.

Between 11/2008 and 08/2012 all cats whose diagnostic workup included a Spec fPL and an AUS examination were included. The time interval between Spec fPL determination and ultrasound examination was always < 24 hours. AUS examinations were performed by residents who were supervised by board-certified radiologists, or by board-certified radiologists. The following parameters were evaluated: ultrasonographic diagnosis of pancreatitis (yes/no), pancreatic size, pancreatic echogenicity, mesenteric echogenicity, pancreatic margins, the presence of cysts and/or masses, common bile duct/pancreatic duct diameter, and the presence or absence of free fluid. The agreements between Spec fPL and ultrasonographic findings were assessed using Cohen's kappa coefficient (SPSS Version 20). Values of kappa from 0.00-0.20 are considered a slight, 0.21-0.40 a fair, 0.41-0.60 a moderate, 0.61-0.80 a substantial, and 0.81-1.00 an almost perfect agreement.

Out of 161 included cases, 75 (46.6%) had an ultrasonographic diagnosis of pancreatitis. The following abnormalities were recorded (n): Hypoechoic pancreas (44), free fluid (43), pancreatic enlargement (35), mixed-echoic pancreas (33), hyperechoic mesentery (30), irregular pancreatic margins (26), dilation of common bile duct (14), dilation of pancreatic duct (13), hyperechoic pancreas (11), pancreatic mass (6), pancreatic cyst (5). An ultrasonographic diagnosis of pancreatitis and Spec fPL (cut-off 5.4 lg/l) had a kappa of .26 (SE .07). Maximum agreement (kappa .33; SE .07) was found with a Spec fPL cut-off > 16 lg/l. The following kappa values (Spec fPL cut-off 5.4 lg/l) were calculated for ultrasonographic findings: enlarged pancreas (kappa .22; SE .07), hypoechoic pancreas (kappa .26; SE .07), irregular margins (kappa .12; SE .61), hyperechoic pancreas (kappa .03; SE .04), mixed-echoic pancreas (kappa .14; SE.07), hyperechoic mesentery (kappa .13; SE .06), dilation of common bile duct (kappa .04; SE .05), dilation of pancreatic duct (kappa .08; SE .05), pancreatic cysts (kappa .02; SE .03), pancreatic masses (kappa -.02; SE .03), and free fluid (kappa .2; SE .07). The maximum agreement for an enlarged (kappa .4; SE .09) or hypoechoic pancreas (kappa .37; SE .08) was found with a Spec fPL cut-off > 31 lg/l, and > 17 lg/l respectively.

In conclusion ultrasonographic findings commonly attributed to pancreatitis do not agree convincingly with serum Spec fPL concentrations. Cisapride is commonly used as an adjunctive treatment for canine esophagitis due to the assumed positive effects on the lower esophageal sphincter (LES) pressure. However no studies have ever validated this concept in veterinary medicine, notably the effects of cisapride on the LES have never been followed over the course of time. High-Resolution Manometry (HRM) is a new, non-invasive, promising tool for measuring esophageal pressure profiles in human medicine. We recently assessed the feasibility in healthy dogs and the aim of the current study was to evaluate the duration of effect of oral cisapride administration on LES pressure in awake healthy dogs.

The study population consisted of 6 healthy adult Beagle dogs (median BW 13.5 kg, age 21 months), the study protocol was approved by the local ethics committee. Cisapride was given orally at a dose of 0.5 mg/kg of body weight. HRM was performed with a solid-state manometry catheter equipped with 40 pressure sensors spaced 1 cm apart with the dog in a sitting position after an overnight fast. Real-time pressure imaging on screen during catheter intubation enabled accurate placement of the catheter with its tip lying intragastrically. Following nasal insertion of the catheter, measurements were started after a 5 minutes adaptation time. The LES pressure was recorded for a period of 20 minutes; the same protocol was repeated 1, 4 and 7 h post cisapride administration.

The LES pressure was calculated with a virtual e-sleeve over a median period of 10.6 min out of the total measuring time. Spontaneous deglutitive LES relaxations and post contractions as visualized on screen were excluded from the analysis. Results (baseline vs. 1, 4 and 7 h post cisapride administration) were compared using the Wilcoxon test. Statistical significance was set at p < 0.05.

The median values (ranges) were i) baseline pressure 29.1 mmHg (23.6-48.4) ii) 1 h post cisapride 44.4 mmHg (38.6-77.3) iii) 4 h post cisapride 50.7 mmHg (44.9-72.8) iiii) 7 h post cisapride 44.3 mmHg (43.1-54.9). Significant differences compared to baseline pressure were found for 1 h (P = 0.0313) and for 4 h post cisapride administration (P = 0.313).

In conclusion, cisapride significantly increases the LES pressure over at least 4 hours. Due to lacking effects 7 h after medication, a more narrow administration interval may be considered in patient with severe esophagitis, in order to avoid potentially harmful reflux episodes. The gastrointestinal (GI) microbiota plays an important role in host health. The combination of an altered composition of the GI microbial community (dysbiosis), underlying host genetic susceptibility, and environmental factors (i.e., diet) are suspected to contribute to the pathogenesis of idiopathic inflammatory bowel disease (IBD). However, only limited information is available about the fecal microbial composition in dogs with IBD. The aim of this study was to characterize the fecal microbiota in dogs with IBD before and after medical treatment for this disorder.

Fecal samples were collected from healthy dogs (n = 10) at one time point, and from dogs with IBD (n = 12) before and after 21 days of standard medical treatment (e.g., elimination diet and administration of anti-inflammatory drugs). The disease status was assessed using the clinical disease activity index (CIBDAI). Fecal DNA was evaluated for quantitative PCR (qPCR) analysis for selected bacterial groups: total bacteria, Bacteroidetes, Fusobacteria, Ruminococcaceae, Bifidobacterium spp., Blautia spp., E. coli, Faecalibacterium spp., Lactobacillus spp., and Turicibacter spp. Differences in bacterial abundance among healthy dogs and IBD dogs (before treatment) were evaluated using a Student's t-test or a Mann Whitney test where appropriate. Samples collected before and after treatment from each affected dog were compared using a paired t-test or a Wilcoxon signed rank test where appropriate. The Benjamini & Hochberg's False Discovery Rate was used to correct for multiple comparisons and an adjusted p < 0.05 was considered to be statistically significant. The CIBDAI was compared in IBD dogs before and after treatment using a Wilcoxon signed rank test.

Before treatment, the abundance of Blautia spp., Faecalibacterium spp., and Turicibacter spp. was significantly decreased in dogs with IBD (p = 0.037, 0.048, 0.029, respectively) when compared to the healthy dogs. E. coli appeared to be increased in feces of IBD dogs and approached statistical significance (p = 0.051). There were no significant differences in the abundance of the other selected bacterial groups between dog groups. CIBDAI was significantly decreased after treatment in the dogs with IBD (median [range]: 1 [0 -5]) compared to before treatment (6 [1 -13]), p = 0.002). However, no significant differences in the abundance of any bacterial groups were found pre-versus post-treatment in IBD dogs.

In conclusion, fecal dysbiosis was observed in dogs with IBD. Although the CIBDAI decreased significantly after 21 days of medical treatment, the abundance of selected bacterial groups did not change significantly during the trial. This observation suggests that a longer period of treatment or a different approach to medical therapy (i.e., pre-/probiotics) may be needed for dogs to recover from fecal dysbiosis associated with IBD. Gastrointestinal (GI) issues are associated with numerous functional changes within the GI tract, including changes in the GI microbiome. The objective of this controlled, cross-over clinical study of 15 cats with naturally-occurring diarrhea was to evaluate changes in GI microflora using metagenomic pyrosequencing, and to correlate those changes with fecal scores (FS) of cats before and after receiving dietary therapy. The study evaluated the response to 2 canned therapeutic diets (Diet X: Hill's â Prescription Diet â i/d â Feline; or Diet Y: Purina Veterinary Diets â EN Gastroenteric â brand Feline Formula), with 4 weeks allowed per therapeutic diet followed by cross-over. FS were recorded during the final week of each period using a 7 point FS system. FS improved over baseline with both diets, but were significantly better after Diet Y versus Diet X. Fecal DNA samples were extracted and the V1-V2 hypervariable regions of the microbial 16S rRNA gene were amplified using primers suitable for 454-pyrosequencing, generating 384,255 sequences. Dominant bacterial phyla included the Bacteroidetes (34%) and Firmicutes (30%), followed by Fusobacteria (18.8%), Proteobacteria (7.7%), Tenericutes (6.6%) and Actinobacteria (2.6%). Orthogonalpartial least squares (OPLS-DA) clustering showed microbial differences between cats when fed Diet X versus Diet Y, and with Diet Y versus baseline. No microbial differences were found between baseline and following Diet X. Significant correlations were found between metagenomic data and FS for the two diets. The data suggest that alterations in intestinal microflora are associated with improvement in diarrhea: further research is needed to determine cause versus effect. It is estimated that~15% of foster kittens die or are euthanized due to illness before 8-wks of age. Most are reported to have clinical signs or post-mortem evidence of small bowel enteritis. The enterococci play conflicting roles as commensal flora, probiotics, and opportunistic pathogens in kittens. A better understanding of what constitutes "normal" enterococcal flora in the small bowel and the impact of severe illness on this microbiota is needed. The purpose of this study was to determine the species identity, virulence attributes, and prevalence of enteroadherent and mucosa-associated enterococci in the small bowel of kittens and their association with mortality.

Fifty apparently healthy kittens euthanized by animal control (Group A) and 50 foster kittens that died or were euthanized due to severe illness at an SPCA (Group B)(all kittens 12-wks of age and 1 kg body weight) underwent necropsy and FISH for identification of enteroadherent bacteria. The mucosa-associated enterococci in the ileum were cultured, speciated, characterized by genotype and phenotype for virulence, and tested for multidrug resistance.

Seven (14%) kittens in Group A and 30 (60%) kittens in Group B had clinical or post-mortem evidence of enteritis; most commonly of small bowel origin. The ileum mucosa-associated enterococci of apparently healthy kittens was predominated by E. hirae. Overt enteroadherence of E. hirae to the small bowel epithelium was common and extensive in healthy kittens. Isolates of E. hirae generally lacked phenotypic or genotypic determinants of virulence. Kittens that died or were euthanized due to illness had a significant increase in colonization of the ileum mucosa by non-E. hirae enterococci, notably E. faecalis. E. faecalis isolates possessed a high level of gelatinase activity, strong biofilm formation on polystyrene, multiple virulence genotypes and antimicrobial resistance. Attachment of enteropathogenic E. coli to the intestinal epithelium was significantly associated with terminal illness and was not documented in any kitten for which enteroadherent E. hirae was observed.

These findings identify a significant dysbiosis of the small bowel enterococci in association with terminal illness in foster kittens and suggest that E. hirae may represent an important commensal bacterium with the potential to confer colonization resistance to enteropathogenic E. coli and ostensibly pathogenic E. faecalis. Given the significance of small bowel disease as a cause of mortality in young kittens, these findings have important implications toward identifying species of probiotics having a potential to significantly impact foster kitten survival. Confocal endomicroscopy (CEM) is an endoscopic technology permitting in vivo cellular and subcellular imaging. In people CEM aids real-time, clinical assessment and diagnosis of various gastrointestinal diseases and improves diagnostic yield, by assisting specific lesion targeting for biopsy. The objective of this study was to determine the feasibility of CEM to evaluate gastric mucosal topologic morphology in dogs.

Six clinically healthy colony dogs underwent standard endoscopic evaluation of the gastric mucosa followed by CEM. The fluorophore acriflavine (0.05% solution) was administered topically via an endoscopic spray catheter to provide fluorescent contrast for imaging. A minimum of five gastric sites were assessed and at each location, sequential adjustment of imaging depth allowed collection of a three-dimensional volume equivalent to an 'optical biopsy'. CEM guided pinch biopsies were obtained for histologic comparison.

CEM provided high quality in vivo histologically-equivalent images to a depth of 50-70 lm. CEM images were easily obtainable from the gastric body and pyloric antrum, but reduced flexibility of the endoscope tip limited imaging of the cardia and fundus. Topical acriflavine preferentially stained cellular nucleic acids, allowing evaluation of nuclear morphology, but also provided cytoplasmic detail and aided identification of Helicobacterlike organisms (HLOs) in 5/6 dogs. Gastric villous morphology was altered in 4/5 sites assessed in the dog negative for HLOs, and histologically this dog was diagnosed with chronic, lymphocytic gastritis.

Confocal endomicroscopy provides in vivo histologic-equivalent images during endoscopy. Identification of altered villous morphology in one dog has implications for real-time in vivo diagnosis of gastrointestinal pathology. Measures of disease activity are essential in defining disease burden at diagnosis and for determining the effect of treatment in canine inflammatory bowel disease (IBD). Since GI endoscopy is usually performed for direct visualization of the mucosa and acquisition of biopsies, endoscopic findings might be used to measure disease activity. Previous studies have failed to determine whether standardized endoscopic evaluation is useful in defining the severity and extent of inflammatory lesions in affected dogs. The aim of the present study was to evaluate the inter-and intra-observer agreement in the assessment of endoscopic activity in canine IBD.

Archived images from consecutive duodenoscopy procedures performed in dogs diagnosed with IBD between 2009 and 2012 were reviewed. A total of 35 endoscopic images of normal and inflamed duodenal mucosa from 29 dogs were selected. The endoscopic variables evaluated were hyperemia, erosions, granularity, friability, and lymphatic dilation. Four expert and 4 trainee endoscopists were invited to assess these images for inflammatory activity. Expert endoscopists were experienced and familiar with disease activity of IBD as identified through endoscopy; the trainees had performed few endoscopic procedures. All images were displayed to each endoscopist independently and scored. The assessment of inflammatory changes was repeated 1 month later using the same images, although the endoscopists were not informed that they would review a test set of images prior to their second critique. Weighted kappa statistics were used to estimate inter-and intra-observer variation. A mixed effects logistic regression model used mean values to compare intra-and interobserver agreement between novice and expert endoscopists. A P value < 0.05 was regarded as indicating statistical significance.

Among the 35 images of the test set obtained during duodenoscopic examination in IBD dogs, 6 were of normal mucosa (some images obtained post-biopsy), 6 showed friability, 5 showed hyperemia, 6 showed granularity, 7 showed erosions, and 5 showed lymphatic dilation. The mean intra-observer weighted kappa value for expert endoscopists was 0.11 (P > 0.20) indicating the degree of variability in mucosal assessment between skilled operators was small. In contrast, novice endoscopists showed significant variability (j-value < 0.01; P < 0.05) between operators which was attributed to one operator who scored near the expert range. The weighted kappa score (j = 0.21; P > 0.6) for the other 3 novice endoscopists showed little intra-observer variation in endoscopic assessment. Regression analysis showed a significant (P = 0.03) difference between groups regarding lesion assessment with expert endoscopists having less chance of disagreement.

Accurate assessment of IBD disease activity from endoscopic findings benefitted from experience. Acceptable agreement rates might be obtained by endoscopists under training using welldefined endoscopic appearances. Lipoproteins transport lipids in the blood stream and are classified into 6 classes: chylomicrons, very low density lipoproteins (VLDL), intermediate density lipoproteins (IDL), low density lipoproteins (LDL), high density lipoproteins (HDL), and lipoprotein (a) based on their density. Based on the human literature, altered proportions of lipoprotein classes have been described in certain disease states, such as cardiovascular disease, metabolic disease, and inflammatory diseases. Exocrine pancreatic insufficiency (EPI) results in inadequate production of pancreatic digestive enzymes leading to maldigestion. The aim of this study was to compare serum lipoprotein profiles and its correlations with serum cholesterol and triglyceride concentrations in dogs with confirmed EPI and healthy dogs.

Analysis of serum lipoprotein profiles were performed in 31 healthy dogs and 25 dogs with confirmed EPI (cTLI 2.5 lg/L) using a density gradient ultracentrifugation technique. The image of each tube following ultracentrifugation was converted to a density profile using a commercially available software program. In addition, serum cholesterol and triglyceride concentrations were measured using a commercially available clinical chemistry analyzer (SIRRUS â , STANBIO, Boerne, TX, USA). Dogs with EPI were divided into two groups: those being treated with enzyme supplementation (n = 15) and those not yet treated (n = 10) at the time of serum sample collection. Data were analyzed using a Wilcoxon rank sum test. Significance was set at p < 0.05.

Triglyceride-rich lipoproteins (TRL), LDL and HDL levels in healthy dogs were significantly higher than in EPI dogs (p = 0.0150, p = 0.0021, and p = 0.0002, respectively Homocysteine (HCY) is an amino acid that is metabolized by 2 pathways: 1) methylation to methionine via the methionine synthase pathway, which requires cobalamin and folate (or betaine in an alternate pathway), and 2) transsulfuration to cystathionine via cystathionine-b-synthase, which is pyridoxal-5-phosphate dependent. Hyperhomocysteinemia is commonly reported in humans with inflammatory bowel disease, and is thought to be due to cobalamin and/or folate deficiency. The prevalence of hyperhomocysteinemia in dogs with chronic enteropathies (CE) is unknown. The aim of this study was to measure serum HCY concentrations in dogs with CE.

Serum samples were collected from 20 dogs with CE that had histopathologically confirmed intestinal inflammation, in which exocrine pancreatic insufficiency had been excluded by measurement of serum trypsin-like immunoreactivity. Serum concentrations of cobalamin (reference interval [RI]: 252-908 ng/ L), folate (RI: 7.7-24.0 lg/L), and HCY (RI: 30.7 lmol/L) were measured in all 20 dogs. Serum HCY concentrations were compared between hypo-and normocobalaminemic dogs using a Mann-Whitney U test. A correlation between serum cobalamin, folate, and HCY concentrations was evaluated using Spearman's rank correlation. Statistical significance was set at p < 0.05.

The median serum HCY concentration in all 20 dogs was 9.5 lmol/L (range: 2.7-71.4). One dog had a HCY concentration higher than the upper limit of the RI. The median serum cobalamin concentration in all 20 dogs was 297 ng/L (range: 149-1,000). Eight dogs were hypocobalaminemic, 3 of which had undetectable cobalamin concentrations ( 149 ng/L), including the dog with an increased serum HCY concentration. There was no significant difference in HCY concentrations between hypoand normocobalaminemic dogs (p = 0.91). Serum folate concentrations (median [range]: 13.8 lg/L [7.0-21.5]) were within the RI in all but 1 dog. No correlation between serum HCY and cobalamin or folate concentrations was identified.

These data suggest that hyperhomocysteinemia may not be common in dogs with CE. While increased serum HCY concentrations in human patients with IBD are believed to be primarily due to cobalamin and/or folate deficiency, no difference in serum HCY concentrations was seen between hypo-and normocobalaminemic dogs with CE in this study, indicating no major effect of cobalamin on serum HCY concentrations in this group of dogs. It is possible that hypocobalaminemia was not of sufficient severity to cause an increase in HCY concentration. However, in humans, HCY is a sensitive marker for both cobalamin and folate deficiency, thus it is interesting that HCY did not differ between hypo-and normocobalaminemic dogs. Serum folate was decreased in only 1 dog, and statistical analysis was therefore not possible. It is conceivable that the betaine or transsulfuration pathways in dogs compensate more efficiently for folate and/or cobalamin deficiency than in humans, which warrants further studies. Tritrichomonas foetus (TF) is a protozoan that parasitizes the feline ileum and colon resulting in chronic diarrhea. TF is distributed worldwide, and no commercially available drugs consistently treat the infection. The mechanisms by which TF colonizes the feline intestine and causes diarrhea are largely unknown. Thus, defining these cellular mechanisms holds promise for development of novel treatment strategies to prevent or ameliorate the infection. Parasitic proteases have been identified as key virulence factors in venereal infection of cattle and women by TF and T. vaginalis, respectively. Because these proteases can be pharmacologically inhibited, they present potentially attractive therapeutic targets. The aim of this study was to determine which classes of proteolytic activity are characteristic of feline TF and to determine the role of these proteases in TF survival, adherence to intestinal epithelial cells and cytopathogenicity.

The class-specific protease activities of protein extracts obtained from 3 feline TF and 1 feline Pentatrichomonas hominis isolate were identified by means of substrate-polyacrylamide gel electrophoresis in the presence or absence of cysteine (E64 0.3-0.6 mM), metallo-(EDTA 0.5 mM), serine (PMSF 0.5-1.5 mM; DFP 0.01-10 mM; AEBSF 1-4 mM) and aspartic (pepstatin A 0.01-0.1 mM) protease inhibitors. The effect of protease inhibitors or their diluents on TF growth and adhesion was determined using feline TF labeled with [ 3 H]thymidine that were allowed to adhere to monolayers of porcine intestinal epithelial cells (IPEC-J2) in co-culture. The cytopathogenic effect of TF was quantified by immunoblotting TFinfected IPEC-J2 cells for the M30 antigen of cleaved cytokeratin 18, which is used as a marker of apoptosis. A minimum of 3 replicates were performed for each experiment. Data were analyzed using Systat software (p < 0.05).

Patterns of substrate-gel proteolysis, demonstrated by 3 different feline isolates of TF, were similar and revealed the presence of multiple high and low molecular weight proteases. On the basis of pharmacological inhibition, these activities were identified as serine and cysteine proteases, respectively. P hominis produced little protease activity compared to TF and had no cysteine protease activity. Serine protease inhibition resulted in death of TF while cysteine protease inhibition blocked adhesion of TF to the intestinal epithelial cells and significantly inhibited enterocyte apoptosis.

These studies establish that serine and cysteine proteases are the predominant mediators of feline TF proteolytic activity and identify the specific involvement of serine proteases in TF survival and cysteine proteases in TF adhesion and cytopathogenicity to intestinal epithelial cells. These results provide strong support for the further study of pharmacological serine and cysteine protease inhibitors for potential prevention, treatment or amelioration of feline TF infection. Recent molecular studies have revealed that the canine gastrointestinal tract (GIT) harbors a highly complex microbial ecosystem. Gut microbes play a very important role in the development and regulation of the host immune system, which is believed to be mediated in part through the production of immunomodulatory metabolites, such as short-chain fatty acids (e.g., butyrate, propionate, acetate). Alterations in the small intestinal microbiota of dogs with chronic enteropathies have recently been demonstrated using 16S rRNA gene sequencing. However, limited information is available about any potential changes in the predominant bacterial groups in dogs with acute diarrhea. In previous studies, Blautia spp., Fusobacterium spp., and Faecalibacterium spp. have been shown to be abundant groups in the canine GIT and have been shown to be producers of immunomodulatory metabolites in humans and rodent models. Therefore, the aim of this study was to establish quantitative polymerase chain reaction (qPCR) assays for these bacterial groups and to evaluate differences in the abundance of these groups between healthy dogs and dogs with acute hemorrhagic diarrhea (AHD).

Fecal samples were collected from healthy dogs (n = 15) and dogs with AHD (n = 15). DNA was extracted using a commercial DNA extraction kit (ZR Fecal DNA Kit TM , Zymo Research Corporation). Oligonucleotide primers were designed for caninespecific Blautia spp. and Fusobacterium spp., based on the alignment of their 16S rRNA gene sequences that are available from the Ribosomal Database Project. The qPCR assay for Faecalibacterium spp. was adapted based on previously established inhouse primers. All qPCR assays were performed using a fluorescent dye (SsoFast TM EvaGreen â supermix; Biorad Laboratories). Differences in log amount of DNA between the groups were evaluated using a nonparametric Mann-Whitney test. Statistical significance was set at p < 0.05.

The abundance of Faecalibacterium spp. organisms was significantly decreased in fecal samples from dogs with AHD when compared to healthy dogs (median log DNA 4.56 vs. 6.64; p = 0.0025). The abundance of Blautia spp. organisms was decreased in fecal samples from dogs with AHD when compared to healthy dogs, but this difference did not reach significance (median log DNA 8.93 vs. 9.63; p = 0.0512). The abundance of Fusobacterium spp. organisms was not significantly different in fecal samples from dogs with AHD when compared to healthy dogs (median log DNA 7.62 vs. 7.58; p = 1.0).

In conclusion, this study demonstrates a significant decrease in the abundance of Faecalibacterium spp. in fecal samples from dogs with AHD when compared to healthy dogs. Further studies are warranted to evaluate the resulting functional effects in affected dogs. Idiopathic inflammatory bowel disease (IBD) is an important disease in dogs and a dysregulated innate immunity plays a major role in its pathogenesis. S100A12 (A12) is an endogenous damage-associated molecular pattern molecule that is associated with phagocyte activation and was shown to be increased in serum/fecal samples from dogs with IBD. Fecal A12 concentrations were also associated with endoscopic disease severity in dogs with IBD. A12 binds to the receptor of advanced glycation end products (RAGE), a pattern-recognition receptor, and results of studies in human IBD suggest a role of RAGE in chronic inflammation. Soluble RAGE (sRAGE), a decoy receptor for inflammatory proteins (e.g., A12) was shown to be decreased in human IBD patients and may be a potential therapeutic target in chronic inflammation. This study aimed to evaluate serum sRAGE and serum/fecal A12 concentrations in the diagnosis and treatment of canine IBD.

Serum and fecal samples were collected from 20 dogs with IBD before and after initiation of medical treatment and from 13 healthy control dogs. Serum sRAGE and serum and fecal A12 concentrations were measured by ELISA, and were compared using a Wilcoxon rank sum test between dogs with IBD vs. healthy controls, and between dogs with positive outcome (i.e., clinical remission, n = 14) vs. those that were euthanized (n = 6). A Spearman rank sum correlation coefficient was used to assess the potential relationship of serum sRAGE concentrations with clinical disease activity (using the CIBDAI scoring system), serum and fecal A12 concentrations, and histologic disease severity (using a 4-point semi-quantitative grading system).

Serum sRAGE concentrations were significantly lower in dogs with IBD (median [IQR]: 52 [52-309] ng/L) than in healthy controls (460 [279-577] ng/L; p = 0.001) but were not associated with the severity of histologic changes (p = 0.776), the CIBDAI score before (p = 0.072) and after treatment (p = 0.107), the serum A12 concentration before (p = 0.726) and after treatment (p = 0.701), and the individual outcome (p = 0.407). Clinical remission and the change in serum sRAGE concentration after treatment were not significantly associated (p = 0.278); however, serum sRAGE concentrations increased only in IBD dogs with complete clinical remission. Also, dogs that were euthanized had significantly higher fecal A12 concentrations (1,760 [758-3,162] ng/g) than dogs that were alive at the end of the study (77 [12-1,062] ng/g; p = 0.032).

This study showed that serum sRAGE concentrations are decreased in dogs diagnosed with IBD compared to healthy dogs, suggesting that sRAGE/RAGE may be involved in the pathogenesis of canine IBD. Lack of correlation between sRAGE and A12 concentrations is consistent with sRAGE functioning as a non-specific decoy receptor. Further studies need to evaluate the gastrointestinal mucosal expression of RAGE in healthy and diseased dogs, the formation of A12-RAGE complexes, and the impact of pre-treatment fecal A12 concentrations on outcome. The defecation behavior of dogs is a frequent end-point in clinical trials involving veterinary diets. Surprisingly, to the authors' knowledge, the number of bowel movements per day has not been documented in healthy privately-owned dogs.

A standardized questionnaire was designed to evaluate the daily number of bowel movements. The study parameters included signalment (including breed, age, sex, weight, body condition score), dietary history, housing conditions (including free or limited access outside, walked freely or on a leash), the number of bowel movements per day and fecal scoring. The questionnaire was documented by clients of the primary care clinic of the Veterinary Teaching Hospital of the University of Toulouse under the guidance of final year veterinary students. A Generalized Linear Model was used to evaluate the effect of predictor variables on the number of bowel movements.

One hundred and ninety five questionnaires were adequately documented. The average number of daily bowel movement in the study population was 2.4 (95% Cl: 2.2-2.5). No statistically significant effect of the different predictors was observed.

In contrast to what is reported in cage-housed research dogs, the number of daily bowel movements in privately-owned dogs as evaluated by a retrospective standardized questionnaire was not influenced by age, format, sex or diet. The aim of the present study was to examine the expression patterns of claudins protein, major components of tight junction, in the duodenal mucosa from dogs with IBD.

Duodenal biopsy specimens from 12 dogs with IBD were used in this study. IBD dogs were scored based on canine chronic enteropathy clinical activity Index (CCECAI) to assess the severity of the disease. A histological severity of the biopsy samples were scored based on WSAVA standards. Normal duodenal mucosae were obtained from 6 healthy dogs. The expression of claudin-1, -2, -3, -4, -5, -7, and -8 in the duodenal mucosa were analyzed by Western blot. Band densities were determined using densitometer. A ratio of claudins to b-actin was calculated for each sample. Claudins/b-actin ratio in duodenal samples from dogs with IBD was correlated with CCECAI and WSAVA score.

The median CCECAI score of dogs with IBD was 6 (range, 3-17) and median total WSAVA score was 4 (range, 1-8). Claudin-3, -5 and -7 protiens were detected in the duodenal mucosa from both IBD dogs and healthy control dogs. There is no significant difference in claudin-3, -5 and -7/b-actin ratio between dogs with IBD and control dogs. However, claudin-7/b-actin ratio was lineally correlated with CCECAI score (r s = 0.6149, P = 0.03) but not with WSAVA score. Claudin-3 and -5/b-actin ratio were not correlated with CCECAI and WSAVA score. Expression of claudin-1, -2, -4 and -8 proteins could not be detected in duodenal mucosa of any dog.

In the present study, there is no significant difference in claudins protein expression between dogs with IBD and control dogs. However, claudin-7/b-actin ratio was correlated with CCECAI score. Further studies are needed to elucidate whether increased claudin-7/b-actin ratio is truly associated with clinical severity and could be a useful for the diagnosis and monitoring of canine IBD. Cortisol is the major stress response hormone produced by the adrenal cortices under control of the hypothalamus-pituitaryadrenal axis (HPA) and has been measured in fecal samples from various wildlife species in order to assess stress levels. Simplicity, ready availability of sample material, and lack of animal handling, as well as low cost are the most important advantages for measuring cortisol in fecal samples rather than in serum samples in this setting. However, an assay for the measurement of cortisol in fecal samples from dogs has not yet been analytically validated. Therefore, the goals of this study were to simplify a cortisol extraction method based on a previously published protocol and to analytically validate a commercially available radioimmunoassay (RIA) kit for the measurement of cortisol in canine feces.

Fecal samples were collected from 8 dogs within 15 AE 5 minutes after morning defecation, homogenized, extracted, and aliquots stored at -20°C until use. The extraction protocol was simplified by adjusting operating temperature and time. Using 100% methanol instead of 90% ethanol for the final extraction step allowed for the omission of the time-intensive vacuum evaporation and sample reconstitution. A commercially available RIA kit was used for analytical validation (Coat-A-Count, Siemens, Los Angeles). This RIA kit has previously been validated for the measurement of cortisol concentrations in urine and serum samples from humans. Intra-assay variability for 4 separate extracts per sample from 4 different samples was measured in order to test extraction repeatability. Analytical validation of the RIA consisted of dilutional parallelism (4 different samples with dilutions of 1:1, 1:2, 1:4, and 1:8), spiking recovery (cross-combining 4 samples with known cortisol concentrations in a 1:1 dilution), and intra-and inter-assay variability (4 samples with 6 to 10 aliquots each and 8 repetitions for 4 samples, respectively). Supplied standards covered a range from 1 through 50 lg/dL cortisol and samples for intra-and inter-assay validation were chosen to represent the lower half of the working range of the assay, encompassing 5 out of 6 control standards.

The intra-assay coefficient of variation for fecal extracts used to test variability of the extraction protocol ranged from 2.5 to 8.7%.Observed-to-expected ratios for dilutional parallelism and spiking recovery ranged from 81.8 to 134.7% and 101.5% to 106.5%, respectively. Coefficients of variation for intra-and interassay variability ranged from 4.1 to 10.0% and 4.4 to 12.2%, respectively.

In conclusion, the simplified extraction method in combination with the Siemens Coat-A-Count RIA kit for the measurement of fecal cortisol in dogs is linear, accurate, precise, and reproducible. Further investigation of fecal cortisol concentrations in dogs under varying stress situations is warranted to validate this approach clinically. Several studies in humans have reported that prebiotic and probiotic supplements have a benefit in the management of various gastrointestinal diseases. One of the hypothesized beneficial effects of pre-and probiotics is an enhancement of the host's natural defense system by boosting the proliferation of beneficial bacteria in the gut, and in turn inhibiting proliferation of pathogenic and potentially pathogenic bacteria. The purpose of this study was to determine the effect of a new test formulation of a nutritional supplement (Viyo International NV, Belgium) containing 0.4% fructooligosaccharides (FOS) on the fecal microbiota of healthy dogs.

Ten healthy dogs without any clinical abnormalities on physical examination and routine blood work (CBC, serum biochemistry) and normal serum cobalamin and folate concentrations were enrolled into this study. Fecal samples were collected on days 1, 8, 16, and 24 and frozen as soon as possible. From the 9 th to the 24 th day of the study, the dogs received the supplement at a calculated dose of 3.2 ml/kg bodyweight once a day (or twice a day if the daily volume exceeded 50 ml) before the meal. Before offering the regular food, the supplement had to be fully consumed. On day 24, the physical examination and general bloodwork was repeated. Also, owners were asked to complete a daily questionnaire concerning the clinical signs observed. Bacterial DNA was extracted from all samples and analyzed by 16S rRNA gene 454-pyrosequencing. Subsets of 1,800 randomly selected sequences per sample were analyzed. A non-parametric Friedman's test was performed and the resulting p-values were corrected for multiple comparisons based on the Benjamini-Hochberg False discovery rate. Statistical significance was set at p < 0.05.

Overall, the acceptance of the new test formulation was good to excellent, but one of the 10 dogs showed poor acceptance on two separate days. Most dogs showed no gastrointestinal side effects. However, one dog had a yellow feces and some flatulence during supplementation and another one had foulsmelling feces. While there was a trend from day 1 (timepoint 1, TP1) to day 24 (timepoint 4, TP4) for some bacterial groups on the family and genus level (median (range) Bifidobacteriaceae: TP1 0 (0-0) and TP4 0 (0-0.27), p = 0.024; Veillonellaceae: TP1 0.04 (0-0.14) and TP4 0.06 (0.03-0.18), p = 0.026; Clostridium XVIII: TP1 0.12 (0-1.10) and TP4 0.07 (0-0.84), p = 0.013; Catenibacterium: TP1 2.45 (0-8.09) and TP4 5.02 (0-62.33), p = 0.041) in response to product administration, this effect did not reach statistical significance when adjusting for multiple comparisons. For all timepoints, five bacterial phyla made up almost 100% of all obtained sequences: Firmicutes (26-93% of all sequences), followed by Bacteroidetes (0-66%), Actinobacteria, Proteobacteria, and Fusobacteria (~4% for these three phyla together).

These results suggest that in healthy dogs there is no significant statistical effect of this nutraceutical test formulation on the intestinal microbiota. The neuroendocrine system of the gut plays a significant role in regulating intestinal motor and sensory functions, as well as modulating inflammation in the intestine. The intestinal mucosa is the dominant site of serotonin (5-HT) synthesis in mammals. More than 95% of the intestinal 5-HT is present in the enterochromaffin cells (ECC). When released from these cells, 5-HT acts locally on the intestinal tract, affecting intestinal motility and nociception. 5-HT excess is associated with autonomic hyperactivity of the intestinal tract. Chromogranin-A (CgA) is a protein that is co-stored and co-released with monoamines and peptide hormones of various endocrine cells, including those of the gastrointestinal tract. CgA thus serves as a marker for intestinal endocrine cells. The aim of this study was to compare the expression of both 5-HT and CgA in the small intestine between a group of dogs diagnosed with inflammatory bowel disease (IBD), and a control group of healthy dogs. We hypothesized that both 5-HT and CgA expression would be increased in the small intestine of IBD dogs versus that of the control population.

The medical database at the Oregon State University Veterinary Teaching Hospital was searched for dogs diagnosed with IBD. Small intestinal mucosal biopsy specimens from thirteen IBD dogs and 8 healthy dogs from an unrelated project were selected and evaluated. Serotonin and CgA expression was determined by immunohistochemistry (IHC). For each dog, the number of CgA and 5-HT positive cells was counted in 20 random high power fields (hpf). Results were expressed as the average number of positive cells/hpf. Differences in mean CgA+/hpf and 5-HT+/hpf were compared using Student's t-test. Pearson's correlation coefficient was calculated for 5-HT+/hpf vs CgA+/hpf.

Both the IBD dogs and control dogs showed enterocytes with strong staining for 5-HT and CgA. The mean positive cells/hpf were significantly increased for both 5-HT and CgA immunopositive cells in the IBD dogs compared to the control group. (5-HT mean 1.0 cells/hpf vs. 2.3 cells/hpf, p < 0.01; CgA 1.7 cells/hpf vs. 2.7 cells/hpf, respectively, p < 0.05). There was a significant correlation between CgA and 5-HT positive cells/hpf across all dogs (Pearson r 2 = 0.2433 p = 0.016). We concluded that, in this study at least, there is increased expression of 5-HT and CgA in the small intestine of dogs with IBD. These results suggest that upregulation of 5-HT expression in the small intestine of dogs with IBD may have an important role in intestinal dysmotility and visceral nociception in this disease. REGURGITATION IN 43 DOGS (2010 -2012 . P Sattasathuchana 1,2 , JA Lidbury 1 , JS Suchodolski 1 , JM Steiner 1 . 1 Gastrointestinal Laboratory, Texas A&M University, College Station, TX, 2 Department of Companion Animal Clinical Sciences, Faculty of Veterinary Medicine, Kasetsart University, Bangkok, Thailand.

Regurgitation is a common clinical sign in dogs with esophageal disease and may be caused by a wide variety of underlying causes. The objective of this retrospective study was to evaluate the underlying causes of regurgitation in dogs.

Medical records of 43 dogs presenting with regurgitation that underwent a work-up at the Small Animal Hospital at Texas A&M University between January 2010 and October 2012 were reviewed retrospectively. For each dog the signalment, history, clinical signs, diagnostic test results, and final diagnoses were recorded. Cases were divided into 2 groups: group 1 comprised of dogs with acute regurgitation (<2 weeks duration) and group 2 was composed of dogs with chronic regurgitation ( ! 2 weeks duration). The association between group and the reported underlying cause of regurgitation was evaluated using Chisquared tests. Significance was set at P < 0.05.

Sixty three percent (27/43) of dogs had acute regurgitation, while 37% (16/43) had chronic regurgitation. In dogs from group 1, the most common causes of regurgitation recorded were esophagitis in 10/27 (37%), other extra-esophageal gastrointestinal (GI) diseases in 5/27 (19%), esophageal dysmotility without esophageal dilatation in 3/27 (11%), intraluminal obstruction in 1/27 (4%), megaesophagus (ME) in 1/27 (4%), but the cause of regurgitation could not be identified in 26% (7/27). In dogs from group 2, the most common causes were recorded as being extra-luminal obstruction in 4/16 (25%), ME in 4/16 (25%), other extra-esophageal GI diseases in 3/16 (19%), esophageal dysmotility without esophageal dilatation in 2/16 (13%), esophagitis in 1/16 (6%), intraluminal mass in 1/16 (6%), and gastroesophageal reflux in 1/16 (6%). Overall 5/43 dogs (12%) had a diagnosis of ME recorded, but in 2 of these the cause of ME remained unknown due to an incomplete workup, 1 had myasthenis gravis, 1 had another degenerative neuropathy, and 1 had idiopathic ME. The odds of esophagitis being the cause of regurgitation in dogs from group 1 was 9 times higher than for those in group 2 (P = 0.03). The odds of extra-luminal esophageal compression being the cause of regurgitation in dogs from group 2 was 11 times higher than in those from group 1 (P = 0.02). Although, the odds for megaesophagus being the cause of regurgitation in dogs from group 2 was 5 times higher than for those in group 1, this association did not reach statistical significance (P = 0.29).

Esophagitis was the most common diagnosis in dogs with acute regurgitation and was more common in this group of dogs than in dogs with chronic regurgitation. The most common causes for chronic regurgitation were extra-luminal compression and ME. Dogs with chronic regurgitation were more likely to have an extra-luminal esophageal compression as the cause of their regurgitation than those with acute regurgitation.

A-C Duchaussoy 1 , B Bacci 1 , MJ Sharman 1 , P Delaney 2 , C Mansfield 1 . 1 Faculty of Veterinary Science, The University of Melbourne, Werribee, VIC, Australia, 2 Optiscan, Nottinghill, VIC, Australia.

Confocal endomicrosopy (CEM) is a diagnostic tool involving a laser light allowing real time microscopy and optical sampling of a three-dimensional volume equivalent to an 'optical biopsy' of the intestinal mucosal layer, with high resolution of cellular, subcellular and even subnuclear structures. It could therefore be thought of as "in vivo histology". Whilst several publications have assessed CEM to be a relevant and reliable tool to investigate the gastrointestinal tract in people, its use in veterinary medicine is recent. Our group has presented data from the stomach of dogs, but no previous images of the colon of dogs have been described. The staining characteristics are considered to be different due to different cellular structures.

Four healthy, adult client-owned dogs (two males, two females) were recruited. Dogs were prepared as standard for colonoscopy, and standard white light endoscopy images recorded. CEM was performed using a protoptype confocal endomicroscope developed by Optiscan Imaging. This confocal endomicroscope delivers 488 nm (blue) laser light via a single optical fibre. Its characteristics allow acquisition of images from the surface to a depth of 250 lm at 3 lm increments and a variety of frame rates and resolutions ranging from 0.8 frames/sec at 2.1 megapixels, to 6 frames/sec at 0.25 megapixels. Fluorescence is detected in the range of 405-590 nm wavelengths. Images obtained reveal the appearance of goblets cells, connective tissue, inflammatory cells, capillaries and bacteria within the colon. Three dye protocols were assessed: cresyl violet topically, acriflavine topically and fluorescein intravenously. Six pinch biopsies were performed at 60 cm, 40 cm and 20 cm to perform histology, quantitative PCR and FISH.

Cresyl violet showed a lack of fluorescence at low concentration (0.15%) and formation of precipitates at higher concentration (0.3%). The best images were obtained using a combination of intravenous fluorescein (10%) and topical acriflavine (0.05%). Fluorescein alone highlighted vessels, cells, connective tissue, and potentially bacteria between the crypts immediately after injection. Acriflavine stained nuclei and helped to visualize the cytoplasm. Inflammatory changes concordant with descriptions in the human literature were identified in one dog. There was good correlation with histological samples.

CEM is a promising real time tool to assess the integrity and histology of the colon in dogs. A combination of fluorescein IV and acriflavine topically appears best for visualization of subcellular structures in vivo. Confocal endomicroscopy (CEM) is an endoscopic technology that permits in vivo cellular and subcellular imaging of the gastrointestinal mucosa. In people CEM aids real-time, clinical assessment and diagnosis of a range of gastrointestinal diseases and also improves diagnostic yield, by assisting specific lesion targeting for biopsy. The objective of this study was to determine the feasibility of CEM to evaluate small intestinal mucosal topologic morphology in dogs.

Six clinically healthy colony dogs underwent standard endoscopic evaluation of the small intestine followed by CEM. Two fluorophores previously evaluated in people, and shown to be effective and safe in dogs in a previous study, were used to provide contrast. Fluorescein (10% solution, 15 mg/kg) was administered intravenously followed by topical acriflavine (0.05% solution) via an endoscopy spray catheter. A minimum of five sites within the small intestine were assessed and at each location sequential adjustment of imaging depth allowed collection of a three-dimensional volume equivalent to an 'optical biopsy'. CEM-guided pinch biopsies were obtained for histologic comparison.

CEM provided high quality in vivo cellular and subcellular histologically-equivalent images. Intravenous fluorescein provided sufficient fluorescent contrast to allow assessment of the vasculature, cellular cytoplasmic features and goblet cell numbers and distribution. Topical acriflavine preferentially stained cellular nucleic acids, allowing evaluation of nuclear morphology. Quality of captured images was occasionally affected by motion artefact, but improved with operator experience.

Confocal endomicroscopy provides in vivo histologic-equivalent images allowing assessment of mucosal morphology during endoscopy. This has implications for aiding in vivo diagnosis of gastrointestinal pathology. Probiotics are routinely administered to feline patients with evidence of gastrointestinal disease. Most veterinary probiotics contain between 100 9 10 6 and 500 9 10 7 bacteria per dose. The probiotic VSL#3 contains Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus bulgaricus, Streptococcus thermophilus and is licensed for use in human patients with irritable bowel syndrome, ulcerative colitis and an ileal pouch. Recommended doses of VSL#3 for people range from 450 9 10 9 to 360 9 10 10 bacteria per day. High dose probiotic administration has not been reported in healthy cats or cats with chronic gastrointestinal disease. The aim of this study was to evaluate the safety and palatability of VSL#3 probiotic when administered to healthy cats at a dose of 224 9 10 9 bacteria per day.

Eight client-owned clinically normal cats were given the probiotic VSL#3 (112 9 10 9 bacteria per dose) by mouth twice daily for 21 days. Body weights ranged from 4.00 to 5.79 kg, with a median of 4.95 kg. A physical examination, CBC, chemistry panel, and measurement of serum folate, cobalamin, feline pancreatic lipase immunoreactivity, and total thyroxine concentrations were performed on all cats prior to administration and on day 21. Owners were asked to keep a diary with daily observations of fecal score, attitude, appetite, thirst, defecation frequency, fecal volume, and presence of clinical signs of gastrointestinal disease for 8 days prior to probiotic administration and during probiotic administration.

The probiotic was well tolerated, though two cats had occasional soft stool and three cats vomited once during the administration period. Three cats had an increased appetite while receiving the probiotic and two had a diminished appetite, though the cats maintained or gained weight during the administration period. The measured laboratory parameters showed no evidence of toxicity or adverse effects of the probiotic VSL#3.

The probiotic VSL#3 appears to be safe in healthy cats with acceptable palatability at the high dose described. Side effects may include vomiting, loose stool, and changes in appetite. Further study is warranted in cats with chronic gastrointestinal disease. The WSAVA Gastrointestinal (GI) Standardization Group developed a histopathological monograph that attempts to standardize interpretation numerous morphologic and inflammatory features in endoscopic biopsies (EB) from stomach, duodenum and colon. However, even with this standardized scheme there has been considerable disagreement between pathologists. The purpose of this study was to evaluate agreement between pathologists using the WSAVA GI template and determinate histopathological changes of the GI tract in healthy cats.

Nine young healthy cats were included in the study, 5 specific pathogen free (SPF) and 4 client-owned indoor cats. SPF cats were euthanized for other reasons and full-thickness biopsies (FTB) were obtained from stomach, duodenum, jejunum, ileum and colon during necropsy in each cat (15 slides). EB from stomach and duodenum (2 and 3 biopsies respectively) were obtained in each indoor cats during sterilization procedure (8 slides) authorized by owners and approved by Ethics Committee. All tissue samples were stained with hematoxylin and eosin and reviewed by 3 board-certified pathologists (NM, JM, AR) who were blinded to the clinically normal status of the cats. Furthermore, all samples were evaluated according to the WSAVA GI template. The average (l), standard deviation (SD) and percent-age coefficient of variation (%CV) were calculated for the different scoring.

Stomach EB and FTB scoring ranged from 0 to 2 and 0 to 13, and l was 0.55 and 6.5, respectively. Duodenal EB and FTB scoring ranged from 0 to 10 and 0 to 8; l was 5.0 and 2.3, respectively. Considering the overall EB and FTB from stomach and duodenum, range was from 0 to 13 and 0 to 10; l was 2.52 and 3.66; SD was 3.25 and 2.79, and%CV 77.36 and 131%, respectively. FTB from jejunum, ileum and colon scoring results obtained the following results, ranged from 0 to 5, 0 to 3 and 0 to 7; l was 1.71, 1.33 and 1.4; SD was 1.68, 0.97 and 2.06, and% CV was 101, 136 and 67%, respectively.

Our results suggest that there is a clearly disagreement between the pathologists using the scoring template proposed recently by WSAVA in this small group of healthy cats. A major variation was observed in the interpretation of duodenal and ileal biopsies, followed for jejunal biopsies. Surprisingly, mild inflammation (mainly lymphoplasmacytic) was observed in at least one anatomical region of the GI tract in all cats. Nevertheless, we must consider the low number of cats in this study and small number of EB obtained from indoor cats for ethical reasons. Also, this scoring template only considers EB and duodenum from small intestine. Furthermore, we used SPF cats that may present potential differences with pet cats related to their genotypic variability, husbandry, differences in intestinal/environmental flora and stress. In conclusion, a new standardized monograph may be necessary to improve agreement between pathologists and consider EB and FTB as well as all anatomical regions of the small intestine. Finally, an inflammatory response may be present in clinically healthy cats. Stress-related mucosal disease (SRMD) is a significant cause of morbidity and mortality that affects 60-100% of critically ill human patients. The prevalence of SRMD in dogs is unknown but is expected to affect canines similarly. Critical illness disrupts gastroprotective mechanisms leading to the mucosal barrier dysfunction of SRMD. Traditionally, SRMD is treated using acid suppressants that increase gastric pH. Consequently, gastric bacterial proliferation predisposes patients to tracheobronchial colonization and pneumonia. Sucralfate, a sucrose/aluminum salt complex, does not increase the risk of these complications. We examined the protective effect of sucralfate on acid-injured ex vivo canine gastric mucosa, which exhibits similar barrier dysfunction as SRMD.

Gastric antrum was collected immediately post-mortem from 17 random-source dogs scheduled for euthanasia by an animal control facility. After equilibration on Ussing chambers, acid Ringer's solution (pH 1.2) was applied to the mucosa for 45-minutes. Sucralfate (100 mg/ml) was added to the mucosal bathing reservoir commensurate with and subsequent to acid. Gastric barrier function was assessed by calculating the transepithelial electrical resistance (TER) and permeability to 3 H-mannitol. Hematoxylin and eosin stained mucosa from each treatment was examined via light microscopy.

Mucosal application of acid Ringer's solution significantly decreased TER and increased flux of 3 H-mannitol. Sucralfate ameliorated acid-induced changes in barrier function (TER p < 0.001 Figure 1 , flux p = 0.008) and accelerated recovery after injury (TER p < 0.035). Histologic disruption of the superficial gastric epithelium induced by acid was prevented by sucralfate administration. These findings suggest that sucralfate has utility in treatment of SRMD mediated by acid injury to gastric mucosa. Chronic diarrhea is an important and often frustrating condition in dogs and cats. While multifactorial, chronic diarrhea can be associated with profound changes in the intestinal microbiome, which might play critical roles in causing or potentiating disease. Recent evidence has indicated that stool transplantation (fecal bacteriotherapy) can be highly effective for treatment of recurrent Clostridium difficile infection in humans through restoration of a normal intestinal microbiota. The objective of this study was to provide preliminary assessment of a fecal transplantation regimen in dogs and cats.

Case 1 was a 3 yr old dog with eosinophilic inflammatory bowel disease of 2 yrs duration that was only moderately controlled with conventional therapy. Case 2 was a 6 yr old domestic shorthaired cat with vomiting and diarrhea of 16 months duration and unremarkable intestinal biopsies that was poorly response to antimicrobials, anti-inflammatories, probiotics and diet changes. Fresh fecal samples were obtained from healthy donors that were screened for a range of enteropathogens. Following a warm water enema, 10 ml/kg of fecal suspension was administered by enema, with 45 minutes retention time. The cat was also treated with 3 ml fecal suspension into the duodenum via endoscope. Fecal samples were collected from the cases prior to and after treatment for microbiome assessment via next generation sequencing of 16s rRNA gene PCR products. Sequences were classified into operational taxonomic units (OTUs) and bacterial structures compared by the Classical Jaccard measure of dissimilarity.

Fecal consistency of the dog was improved within 24 h and he has been clinically normal for 3 months post-treatment, apart from a bout of diarrhea following enrofloxacin administration that was responsive to metronidazole. The cat fared similarly, with rapid improvement in fecal consistency, attitude and appetite. Vomiting and diarrhea have been absent and clinical normalcy has been maintained for 3 months. 42580 sequences were obtained and subsampled based on the lowest number of sequences for a single sample (4981). By day 2, fecal samples from the cases clustered phylogenetically with the donors, not the animals' own baseline samples. Species richness was highest (322 OTUs) in the feline baseline sample, along with greater microbial diversity, perhaps indicating a less adapted and structured microbial population. Species richness decreased after treatment. In contrast, species richness was lowest in the baseline sample in the dog, with an increase after treatment.

While preliminary, this treatment regimen shows promise for the treatment of chronic diarrhea. The clinical response is consistent with the marked shift in the fecal microbiome following treatment. Optimal species richness and population diversity are unknown and require further study. (2002 -2012) . LE Ferguson, SA Wennogle, CB Webb. Colorado State University, Fort Collins, CO.

Bilious vomiting syndrome (BVS) is a diagnosis associated with early morning vomiting of bile. This condition is thought to result from a reflux of duodenal fluid into the gastric lumen causing mucosal irritation.

Medical records were searched for "canine" and "bilious vomiting syndrome" between the years of 2002 and 2012. Visual inspection confirmed that a diagnosis of BVS had been applied at some point during the recorded history.

The following parameters were recorded: signalment, presenting complaint, duration of clinical signs, prior treatments, response to treatment, and diagnostics performed. The diagnosis remained BVS for the duration of the dog's contact with CSU in 17 cases. Therapy involved frequent small feedings, prokinetics, and gastroprotectants. In 4 cases a novel protein diet was initiated. 12 dogs improved with therapy. 5 dogs did not improve or were lost to follow-up. The diagnosis of BVS was supplanted in 3 cases, including gastric adenocarcinoma, dietary indiscretion, and hepatopathy.

The canine most likely to present with clinical signs consistent with BVS was a young, mixed breed, male-castrated dog with a chronic history of vomiting bile. The most frequent clinical signs were vomiting bile, hyporexia, and nausea. The most common therapies that produced a response were metoclopramide, antacids, and small frequent feedings.

Results of this study suggest that BVS continues to be a diagnosis of exclusion. Response to therapy suggests an underlying abnormality in gastrointestinal motility. Further evaluation of dogs diagnosed with BVS using Smart Pill technology may prove informative. Tenets of gastrointestinal physiology hold that gastric emptying is allowed optimally when isotonic effluent is available to be delivered to the duodenum. Therefore presence of water in food should allow more rapid delivery of metabolites to the duodenum. Absorption of metabolites is often indicated by an increase in their concentration in the plasma. Concentration of plasma glucose has been shown to remain constant in the plasma despite its absorption. Because some glucose is converted to lactate during its absorption, increased plasma lactate concentration indicates glucose absorption (1). Amino acids are also increased in plasma with absorption. We investigated the timing of postprandial metabolite concentrations as an indication of the timing of absorption from the intestine.

We fed four dogs meals of either science diet, adult original dry plus 1.5 times the volume of water (64% water) (D + W); adult original dry without added water (10% water) (D); or canned chicken science diet (C) (78% water) (C). Baseline venous samples were taken ½ hour prior to feeding, just before feeding, ½ hour, 1 hour, and 1½ hours after feeding, then hourly for 8 hours. Plasma samples were analyzed for glucose, lactate, insulin and total amino acids (alpha-amino N).

There was no significant change in mean glucose concentration after ingestion of any ration. Mean insulin increased significantly by 1½ hours after ingestion of W+D, and 1 hour after ingestion of D and C. Amino acid concentrations rose in all dogs and remained elevated throughout the sampling period, in agreement with previous studies (2,3). However, mean alpha-amino N increased significantly within 1½ hours of ingestion only when dogs ate D+W. In agreement with others, lactate concentration changes occurred soon after feeding in all dogs and tapered off four hours after ingestion. Mean lactate increased significantly within one hour of ingestion of D+W and within 2.5 hours of ingestion of C but did not significantly change after ingestion of D.

These data suggest that increased percentage of water ingested with food promotes gastric emptying. 1. Abumrad, N.N. et al., AJPhysiol 242:E398, 1982 . 2. Davis, M.A. et al., JPhysiol 247:E362, 1984 . 3. Goriya, Y. et al., AJPhysiol 240:E54, 1981 .

A RETROSPECTIVE STUDY (2004) (2005) (2006) (2007) (2008) (2009) (2010) (2011) (2012) . RJ Tulipan, J Duval, D Duval, N Berryessa, L Roth-Johnson. Georgia Veterinary Specialists, Sandy Springs, GA.

The purpose of this study was to investigate the relationship between biliary mucocele and inflammatory bowel disease and to determine if dogs with biliary mucocele were more likely to have histopathologic evidence of inflammatory bowel disease.

Medical records from Georgia Veterinary Specialists were reviewed for this retrospective case series. The experimental group consisted of patients that underwent surgery for biliary mucocele from January 2004 to July 2012, and had intestinal biopsies reviewed by a selected pathologist.

The control group comprised patients that underwent gastrointestinal surgery, and were not suspected of having biliary disease, during the same period, and had intestinal biopsies reviewed by the same pathologist. Information pertaining to breed, age, concurrent diseases, liver biopsy results and intestinal biopsy results was obtained for all animals included in the study. Statistical analyses were performed to determine differences between groups.

Histopathologic lesions consistent with inflammatory bowel disease were significantly more prevalent in dogs with biliary mucocele (92.5% of dogs with biliary mucocele, 40% of control animals; P < 0.0001), and control animals were significantly more likely to have an eosinophilic component to their inflammation (2.7% of dogs with biliary mucocele, 29.2% of dogs selected as controls; P = 0.0028).

In the present study, a significantly greater proportion of the control animals had an eosinophilic component to their intestinal inflammation, indicating an element of allergy or hypersensitivity. A positive association between biliary mucocele and inflammatory bowel disease was found. In the present study we were unable to determine whether this relationship is causal or correlational. This positive association between biliary mucocele and inflammatory bowel disease provides insight into each of these complex and poorly understood diseases, and provides impetus for further study. The antineoplastic agent doxorubicin commonly induces gastrointestinal signs such as diarrhea in canine patients. In some patients, gastrointestinal signs can be dose limiting. The severity of gastrointestinal signs typically peaks at 3 days; these gastrointestinal signs are associated with increased serum C-reactive protein (C-RP) concentrations. We hypothesized that treatment with E. faecium SF68 would reduce the frequency, duration and severity of diarrhea and blunt changes in serum concentrations of C-reactive protein in dogs receiving doxorubicin chemotherapy as part of their therapy for naturally occurring neoplastic disease.

Twenty-four dogs (12/treatment arm) were enrolled into the study from the Oncology services at Oregon State University, Corvallis, OR and Veterinary Specialty Hospital, San Diego, CA. All dogs had naturally occurring neoplastic disease and were scheduled to receive doxorubicin as part of their planned chemotherapy protocol. E. faecium SF68 (Fortiflora â ) and placebo (Fortiflora without SF68) were compounded into cellulose capsules and randomly assigned to numbered containers, information regarding the contents of each treatment container was maintained by an investigator with no patient interactions. Dogs admitted to the study began receiving their assigned treatment (2 capsules/dog per os SID) starting two days prior to the planned day of doxorubicin administration. Serum samples for C-RP determination were collected on days 0, 3 and 7 postdoxorubicin, while owners maintained diaries of fecal consistency daily for days 0-10, using a 7 point scale with 7 representing watery diarrhea.

Serum C-CRP was significantly increased over baseline at day 3 in the placebo group, but not in the treatment group (Two-Way ANOVA with Tukey's HSD post-test, p < 0.05). Fecal scores altered significantly in both groups (Repeated Measures ANOVA, p < 0.05), consistent with prior observations, however there was no significant effect noted from E. faecium treatment on duration of diarrhea or probability of diarrhea. The detected difference in number of days with fecal score >5 approached significance (Treated group median of 0 days (range 0-4), Placebo group median 2.0 days (range 0-6), p = 0.077, Mann-Whitney U Test). Using the treatment effect detected, we calculated that a study with at least 29 patients per treatment arm would be necessary to achieve 80% power to detect statistical significance for this parameter. Number-needed-to-treat calculations based on decrease in number of days with fecal score >5 show that four patients need to be treated with E. faecium SF68 for one to show benefit. No adverse effects were noted in this study attributable to use of E. faecium SF68. In human inflammatory bowel disease (IBD), colonic macrophage population is known to increase with high levels of proinflammatory cytokines. However, there is no clear evidence for the up-regulation of pro-inflammatory cytokines from the colonic mucosa of dogs with inflammatory colorectal polyps (ICRP) in miniature dachshunds. ICRP is recently recognized in Japan and is thought to be a novel form of canine IBD as opposed to lymphocytic-plasmacytic colitis (LPC), a major form of canine IBD. It is characterized by the formations of multiple small polyps (small polyp) and/or solitary space-occupying large polyp (large polyp) with histopathological features of infiltrating inflammatory cells such as macrophages and neutrophils. Therefore, the aim of the current study was to investigate key proinflammatory cytokines involved in the formation of colorectal polyps in ICRP.

Colorectal tissue samples were obtained endoscopically and/or surgically from dogs with ICRP (large polyp, n = 9; small polyp, n = 5), dogs with LPC (n = 6), and healthy dogs (control, n = 8). Pro-inflammatory cytokine (IL-1b, IL-6, TNF-a, IL-8, IL-12p35, IL-12/23p40, and IL-23p19) mRNA expressions were quantified by real-time PCR. IL-8 protein levels in colorectal mucosa of dogs with ICRP were evaluated by ELISA and its location were investigated by immunofluorescence microscopy.

The expressions of all inflammatory cytokines mRNA were significantly increased in large polyp of dogs with ICRP compared to control. The expressions of IL-1b, IL-6, IL-8, and IL-12p35 mRNA were significantly increased in small polyp compared to control. IL-8 mRNA expression was increased 10,000fold in large polyp and 100-fold in small polyp when compared to control. IL-8 protein levels were significantly elevated in large polyp compared to small polyp and control, and IL-8 was localized in macrophages.

These results suggest that pro-inflammatory cytokines may play an important role in the pathogenesis of ICRP. Furthermore, IL-8 secreted by macrophages may induce infiltrations of neutrophils resulting in formation of colorectal polyps. Acute pancreatitis is the most common disease of the canine exocrine pancreas and accurate noninvasive diagnosis can be challenging. Ultrasound (US) is useful for diagnosis, however when unremarkable cannot rule out presence of pancreatitis. Contrast-enhanced ultrasonography (CEUS) with contrast agents such as Sonazoid â is a major breakthrough for US imaging and provides real-time assessment of organ microvascularization as its contrast-enhancement correlates well with the degree of tissue perfusion. The purpose of this study was to elucidate the perfusional derangements of the pancreas in cerulein-induced pancreatitis in dogs using CEUS.

6 adult female beagles received 7.5 lg/kg/h of cerulein infusion for 2 hours. CEUS of the pancreas was performed at 0, 2, 4, 6 and 12 hours post-infusion. Qualitative and quantitative analyses of perfusion parameters were compared to baseline values (0 hour).

Peak intensity (PI) of the pancreas was increased from 82.6 AE 15.1 MPV at baseline to 101.0 AE 9.1 MPV (P < 0.01) and 99.4 AE 13.9 MPV (P < 0.001) at 2 and 4 hours respectively after cerulein infusion. Area under the curve (AUC) peaked at 4 hours post-infusion (19.6 AE 5.0^10 3 MPV.s, P < 0.001) compared to baseline (14.0 AE 4.6^10 3 MPV.s). Prolonged wash-out time at 4 hours post-infusion (283.3 AE 67.4 s, P < 0.0589) approached significance.

CEUS can be used in dogs for detecting presence of pancreatic inflammation based on contrast-enhancement as a measure of microvascular derangement. Quantifying the PI and AUC can provide valuable information in differentiating acute edematous pancreatitis from the normal pancreas. Prolonged hyperechoic enhancement on CEUS indicates active pancreatitis. The veterinary literature conveys the concept that serum lipase activities measured by traditional catalytic assays are of no clinical value. Currently the Spec fPL assay is believed to be the most sensitive test for the diagnosis of feline pancreatitis, although data are scarce. A colorimetric assay for lipase determination, based on the use of 1,2-o-dilauryl-rac-glycero-3-glutaric acid-(6'methylresorufin) ester (DGGR) as the substrate, is used in the routine diagnostic at the authors' institution, and has recently been evaluated in dogs with acute pancreatitis. However, no information about this test has been published in cats so far. Therefore the aims of this study were to evaluate the agreement of the DGGR-lipase activity and Spec fPL concentration in cats with suspicion of pancreatitis, as well as assessing the effect of concurrent azotemia.

Between November 2008 through to August 2012 DGGRlipase activity and Spec fPL concentration were measured from the same blood sample in cats undergoing investigation for pancreatitis. DGGR-lipase activity was measured using a colorimetric assay (Cobas Integra 800, Roche). A reference interval in our laboratory had been established previously using 80 apparently healthy cats of various breeds and all genders (reference range, 8-26 U/l). Spec fPL was measured through IDEXX Laboratories. The agreement between DGGR-lipase and Spec fPL at different cut-offs was assessed using Cohen's kappa coefficient (SPSS Version 11). Values of kappa from 0.00-0.20 are considered a slight, 0.21-0.40 a fair, 0.41-0.60 a moderate, 0.61-0.80 a substantial, and 0.81-1.00 an almost perfect agreement.

DGGR-lipase (cut-off 26 U/l) and Spec fPL (cut-off 5.4 lg/l) had a kappa of 0.68 (standard error (SE) 0.046). DGGR-lipase (cut-off 26U/l) and Spec fPL (cut-off 3.4 lg/l) had a kappa of 0.60 (SE 0.05). Maximum agreement of Spec fPL (cut-off 5.4 lg/ l) with DGGR-lipase was found when a cut-off of 34U/l was used, and had a kappa value of 0.76 (SE 0.042). In 169 nonazotemic cats, agreement of DGGR-lipase (cut-off 26 U/l) and Spec fPL (cut-off 5.4 lg/l) had a kappa of 0.72 (SE 0.053). In 46 azotemic cats, agreement of DGGR-lipase (cut-off 26 U/l) and Spec fPL (cut-off 5.4 lg/l) had a kappa of 0.61 (SE 0.125).

We conclude that results of both tests agree substantially. Concurrent azotemia does not seem to have a major effect on agreement of both methods of serum lipase determination. Based on these results the DGGR-assay seems a comparably useful and cost-efficient method for the determination of serum lipase in feline pancreatitis cases.

A.Abrams-Ogg, K Ruotsalo, F Reggeti, S Nykamp, A Downie. University of Guelph, Guelph, ON, Canada.

Pancreatic lipase immunoreactivity (Spec cPL, IDEXX) and abdominal ultrasonography are considered the most sensitive tests for the noninvasive antemortem diagnosis of pancreatitis in dogs (Vet Clin Pathol, 2012; 41:312-24) . The Spec cPL test does not have 100% specificity for pancreatitis, but factors affecting specificity are not fully described. As total serum lipase activity may be increased in dogs with reduced renal function, we investigated this effect on Spec cPL using serum creatinine level as a marker of glomerular filtration. Dogs were prospectively enrolled from all cases admitted to the Ontario Veterinary College from November 2009 to November 2012 that had an elevated creatinine level and no evidence of pancreatitis on abdominal ultrasound (in an effort to reduce the effect of pancreatitis on Spec cPL). Creatinine was measured in the Animal Health Laboratory on a cobas â 6000 c501 analyzer (Roche Diagnostics). All ultrasound examinations were performed or reviewed by board-certified radiologists. Findings supportive of pancreatitis included pancreatic enlargement and/or irregularity, decreased echogenicity, increased echogenicity of peripancreatic fat, focal abdominal effusion and extrahepatic bile duct enlargement not attributable to another cause. Spec cPL was determined at the IDEXX laboratory in Markham, ON, Canada, using the same serum sample in which creatinine had been measured.

Fifty-eight dogs (median age 9 years, range 1-13, various breeds) with 70 evaluations were enrolled. Presumptive or definitive diagnoses were: chronic kidney disease (22 dogs); acute kidney injury (15 dogs), post-renal urinary obstruction (1 dog), and disorders other than those of the primary urinary tract (20 dogs), including neoplasia, liver disease, heart disease and endocrinopathies. Serum creatinine level ranged from 155-1328 umol/L (median 217 umol/L, reference interval 20-150). Spec cPL ranged from <30 ->1000 ug/L (median 294 ug/L, reference interval: normal < 200, 200-400 elevated, >400 suggestive of pancreatitis). There was a weak but significant overall correlation between creatinine and Spec cPL (rho = 0.26, P = 0.03), but no significant correlations when data were stratified by 1) Spec cPL as <200, 200-400, >400 ug/L; 2) creatinine as <440 or >440 umol/L; or 3) diagnoses. For dogs with Spec cPL < 30 ug/L (n = 7), creatinine ranged from 156-395 umol/L. For dogs with Spec cPL >1000 ug/ L (n = 8), creatinine ranged from 155-663 umol/L. For the dog where creatinine was 1328 umol/L, Spec cPL was 232 ug/L. Eight dogs had repeated measurements; parallel changes occurred in creatinine and Spec cPL in 5 dogs, while in 3 dogs creatinine and Spec cPL changes occurred in opposite directions.

In conclusion, in dogs without ultrasound evidence of pancreatitis, decreased renal function did not have a major effect on Spec cPL. Whether elevated Spec cPL was due to pancreatitis not detectable by ultrasound or other factors affecting Spec cPL specificity is not known. The weak correlation between creatinine and Spec cPL, and parallel changes in creatinine and Spec cPL in some dogs, may indicate that pre-renal status and renal function have a minor effect on Spec cPL. Little information is only available on the role of adipokines in the development and progression of acute pancreatitis (AP) in dogs. In human medicine, it has been shown that among the adipokines, resistin stimulates inflammatory cytokine(s) expression, and conversely is up-regulated by those cytokines. The objective of present study was to evaluate the change of the levels of serum resistin and inflammatory cytokines in dogs with AP. Total 19 dogs were enrolled in this study. Among the 31-clientowned dogs newly diagnosed with AP, only 12 dogs with severity organ score 2 and normal body condition score (BCS) were selected (Group A). Seven healthy dogs with normal BCS were included as the control group (Group B). All of the dogs in Group A were reassessed at the point of discharge with resolution of clinical signs.

The circulating levels of resistin, tumor necrosis factor (TNF)-a, interleukin (IL)-6, and IL-1b were significantly higher in Group A dogs compared to those of Group B dogs. However, there were no significant differences on the levels of adiponectin, IL-10, and IL-18 between Group A and B dogs. In the Group A, after the disappearance of clinical signs from AP, the levels of TNF-a were significantly decreased; however, the mean values of IL-1b, IL-6, IL-10, and IL-18 were still higher than those of Group B. These results suggested that the up-regulation of resistin levels as well as TNF-a, IL-6, and IL-1 b might play an important role on inflammatory responses associated with AP in non-obese dogs. Pancreatic lipase immunoreactivity (fPLI) and abdominal ultrasonography are considered the most sensitive tests for the noninvasive antemortem diagnosis of pancreatitis in cats (Vet Clin Pathol, 2012; 41:312-24) . Nonspecific serum lipase activity is considered to be insensitive, but this is based on a limited number of reports, and methodologies and reference intervals (RI) vary between laboratories. To examine the sensitivity of lipase at the Animal Health Laboratory, University of Guelph, records were retrieved for all cases where: 1) fPLI had been ordered (either by original RIA, Texas A & M University, or by ELISA [Spec fPL], IDEXX, Markham, ON, Canada); and/or 2) an antemortem diagnosis of pancreatitis had been made on the basis of clinical signs plus abdominal ultrasonography and/or fPLI; and 3) lipase had been measured using the LIPC lipase colorimetric assay (Roche Diagnostics). Lipase had been measured on a Hitachi 911 (Roche Diagnostics) analyzer prior to February 2009, at which time a cobas â 6000 c501 (Roche Diagnostics) analyzer was used. A RI of 12-32 U/L was determined for the Hitachi 911 using 40 mature cats, and was revalidated for the cobas â using 60 mature cats. All ultrasonography had been performed/ reviewed by board-certified radiologists. Findings supportive of pancreatitis included pancreatic enlargement, decreased pancreatic echogenicity, increased echogenicity of peripancreatic fat, and common bile duct enlargement not attributable to another cause.

Fifty-one cats with 62 evaluations were identified from January 2007 to November 2012. Nineteen cats with 20 evaluations had positive ultrasounds and elevated fPLI (RIA 7.1-326.1 ug/L [RI 2-6.8 ug/L], Spec fPL 7.1->50 ug/L [RI 3.5]; lipase was elevated in 16 evaluations (80%, range 39-259 U/L). Six cats with 8 evaluations had negative ultrasounds but elevated fPLI (RIA 7.4 ug/L, Spec fPL 4.1-44 ug/L); lipase was elevated in 3 evaluations (38%, 34-73 U/L). Nine cats with 14 evaluations without ultrasound had elevated fPLI (RIA 30.9-330.8 ug/L, Spec fPL 3.9->50 ug/L); lipase was elevated in 9 evaluations (64%, 46-594 U/L). Five cats with 8 evaluations had positive ultrasounds but normal fPLI (RIA 2.7-5.6, Spec fPL 1.1-1.5 ug/L); lipase was low-to-normal in all (7-23 U/L). Three cats had negative ultrasounds and normal fPLI (Spec fPL 1.0-2.8 ug/L); lipase was low in all (10-11 U/L). One cat without ultrasound had normal fPLI (Spec fPL 1.0 ug/L) and low lipase (11 U/L). Lastly, in 8 cats with positive ultrasounds but no fPLI measurement, lipase was elevated in 6 cats (75%, 67-488 U/L). Lipase was correlated with fPLI-RIA (n = 11, rho= 0.79, P = 0.004) and with Spec fPL (n = 40, rho=0.92, P < 0.001).

If positive ultrasound plus elevated fPLI together is taken as the gold standard for noninvasive antemortem diagnosis of feline pancreatitis, lipase had a sensitivity of 80%. For cases where fPLI was measured, positive ultrasound plus increased lipase had a 100% positive predictive value for elevated fPLI, and negative ultrasound plus low lipase had a 100% positive predictive value for normal fPLI. We conclude that total lipase is useful for the diagnosis of feline pancreatitis. Exocrine pancreatic insufficiency (EPI) is a syndrome characterized by the inadequate synthesis and secretion of pancreatic digestive enzymes resulting in maldigestion. Hypoalbuminemia or hypoproteinemia are clinicopathological abnormalities not usually associated with EPI in dogs. Alpha 1 -proteinase inhibitor, a serum proteinase inhibitor, is lost into the gastrointestinal tract at a rate comparable to that of albumin, and serves as an endogenous marker of gastrointestinal protein loss. The aim of this study was to evaluate fecal a 1 -proteinase inhibitor (fa 1 -PI) concentrations in dogs with EPI.

Surplus fecal samples from 20 dogs enrolled in an unrelated clinical trial at the Gastrointestinal Laboratory were utilized. To be included into the study, the dogs had to be at least 1 year of age, have clinical signs of EPI (i.e., polyphagia, weight loss, steatorrhea, and/or loose, voluminous and/or malodorous stools), have a serum cTLI concentration 2.5 lg/L, not be pregnant or lactating, and be free from any clinically apparent disease other than EPI. Three naturally voided fecal samples collected over three consecutive days, which were frozen immediately after collection, were used. Fecal a 1 -proteinase inhibitor was measured using an in-house radioimmunoassay, and was compared with an established reference interval for healthy dogs; a mean three-day fa 1 -PI of ! 13.9 lg/g feces or a maximum fa 1 -PI of one individual sample of ! 21.0 lg/g feces was considered abnormal. Serum total protein and albumin concentrations were measured using a chemistry analyzer (SIRRUS â Clinical Chemistry Analyzer, Stanbio Laboratory, TX). Serum folate and cobalamin concentrations were measured using previously validated chemiluminescence immunoassays (IMMULITE â 2000, Siemens, IL).

There was considerable day to day variation of fa 1 -PI concentrations (median coefficient of variation [range]: 43.8% [8.6-117.0]) observed in 18 of the 20 dogs; for two dogs three separate fecal samples were not available. Sixty percent (12/20) of the dogs had increased fa 1 -PI concentrations (median [range]: 23.4 lg/g feces [9.1-78.8]). Seventy percent (14/20) of the dogs had a serum total protein concentration below the lower limit of the reference interval (5.7-7.8 g/dL; median [range]: 5.5 g/dL [4.7-6.2]). However, none of the dogs had a serum albumin concentration below the lower limit of the reference interval. Only 50% (7/14) of the dogs with a decreased serum total protein concentration had an increased fa 1 -PI concentration. No correlations were observed between the mean three-day fa 1 -PI concentrations and serum concentrations of total protein, albumin, cobalamin, or folate.

This study shows that fecal a 1 -proteinase inhibitor concentrations may be increased in dogs with EPI. Further studies are necessary to elucidate the mechanisms that are responsible for this finding. Gastroduodenal motility was assessed via ultrasound in 8 healthy dogs by monitoring antral and duodenal contraction amplitude and frequency in both the fasted and post-prandial state after IV administration of metoclopramide, cisapride, erythromycin (0.5 mg/kg each), or ranitidine (2 mg/kg), and saline as a negative control. A randomized double-blind crossover design was used. Gastric and duodenal motility were evaluated by calculating the amplitude of antral and duodenal contractions in triplicate for each time period. Contraction frequency was counted over 3 minutes. Results were evaluated using a mixed model repeated measures ANOVA with dog as a random effect and treatment and time as fixed effects. Statistical significance was set at p < 0.05.

After placebo antrum contraction amplitude (CAA) increased over the first 30 min. post-prandially (pp), then reached a plateau. Cisapride was the only drug that elicited a higher CAA at 30 minutes pp. After placebo antrum contraction frequency (CFA) was increased for the first 60 minutes pp, then reached a plateau. The CFA for erythromycin was greater than placebo for all measurement times. Metoclopramide CFA was greater than placebo until 60 min. pp. No consistent prokinetic effect of drugs was noticed on amplitude and frequency of duodenal contractions.

Ultrasound measurement of contraction frequency and amplitude in the pyloric antrum and duodenum allowed rapid, noninvasive assessment of motility in these segments. In the gastric antrum, tested prokinetics elicited an earlier increase of fasted and postprandial motility when compared to placebo. Some prokinetics appear to cause a sustained increase in antral motility. Rivaroxaban is an oral, direct factor X inhibitor used in human thrombotic disorders. It has promising characteristics for use in dogs: our previous in vitro study showed concentrationdependent effects of rivaroxaban on several coagulation parameters. To test the hypothesis of an in vivo anticoagulant effect in this species, we documented the pharmacodynamics of rivaroxaban.

Healthy dogs were randomized into 3 groups (n = 8) and received orally: placebo, or 2 mg/kg rivaroxaban once or twice daily (8 h, 16 h) respectively. Fifteen blood samples were collected over a 30 h period, and blindly assayed for prothrombin time (PT), partial thromboplastin time (PTT), tissue factor induced thrombin generation (TG) and anti-fXa activity. Thromboelastography (TEG) was evaluated at 0, 1, 4, 8 and 24 h. Peak/baseline anticoagulant effect ratios were analyzed with generalized linear models using beta distributions, and times to return to baseline with survival analyses (a = 0.05).

Treatment groups only significantly differed in pre-treatment albuminemia (p = 0.0425). Peak anticoagulant effect ratios of PT, PTT, anti-fXa activity, TG and R(TEG) differed significantly between placebo and rivaroxaban groups (p < 0.0001). Median return to baseline times for peak TG, time to peak TG, and PT were respectively 19.8 h, 18.0 h, and 11.6 h in dogs treated once with rivaroxaban, and 26.5 h, 27.1 h, and 17.3 h in dogs treated twice. Differences between once and twice daily administration were significant for PT and time to peak (p 0.0026). Other parameters are being analyzed.

These results confirm our hypothesis. Rivaroxaban may be administered twice daily to maintain 24 h anticoagulant efficacy. No adverse effects were recorded. Immune thrombocytopenia (ITP) is a relatively prevalent disease in dogs with significant morbidity and mortality. Canine ITP is clinically analogous to human ITP, with heterogeneity in bleeding manifestations in individuals with similar platelet counts. With a view to ultimately investigate this bleeding heterogeneity, we set out to develop a canine model of ITP. There are currently no existing large animal models of ITP. An induced canine ITP model would be representative of ITP without the confounding co-morbidities seen in clinical cases. Since spontaneous ITP occurs in both dogs and humans, the dog is an ideal translational model. We hypothesized that 2F9, a murine IgG2a monoclonal antibody to the canine platelet glycoprotein GPIIb (a common target of autoantibodies in ITP), would induce predictable dose-dependent thrombocytopenia (TCP) in healthy dogs.

A dose titration (2 dogs) and a dose repeatability study (3 dogs) were performed in healthy adult research dogs by repeated intravenous infusion ( 6 doses) of 2F9 antibody until a target nadir of 5-30 9 10 3 platelets/ll was reached. The following were evaluated daily until platelet count recovery: CBCs, physical examination, serum cytokines and chemokines (INFc, Interleukin (IL) 2, 6, 7, 8, 10, 15, 18, KC, IP-10, MCP-1, GM-CSF, TNFa). Specificity of the 2F9 effect was confirmed by IV infusion of isotype control antibody to 3 dogs at the highest cumulative effective dose of 2F9 (167 lg/kg).

Within 2 hours of a median cumulative 2F9 administration of 63 lg/kg (range 50.0-166.6 lg/kg), all dogs developed profound TCP (range 11-28 9 10 3 /ll). Platelet nadir was in our target range and platelet count remained < 40 9 10 3 /ll in all 2F9-treated dogs for 24 hours. Dosing was predictable: in each dog, after an initial dose of 50 lg/kg 2F9, the second dose needed to reach the target nadir could be accurately calculated from the initial platelet decrease.

2F9-treated dogs developed a range of clinical bleeding from few petechiae to melena and hematuria. Dogs had no changes in vital signs or demeanor and did not require any transfusion support. 2F9 infusion also generated negligible systemic inflammation, as assessed by WBC count and serum cytokine measurement. Unexpectedly, serum IL8 tracked faithfully with platelet count, demonstrating that platelets might be a major source of serum IL8 in dogs. Platelets have not been previously described as a significant serum IL8 source. Since IL8 is an important neutrophil chemokine, our finding may illuminate a novel mechanism of platelet-neutrophil cross-talk.

We have developed a novel large animal ITP model that is representative of the spontaneous disease. As in naturally-occurring ITP, dogs demonstrate bleeding heterogeneity despite similar platelet counts. We expect our model to lead to further insights into bleeding mechanisms in ITP. Gastric dilatation-volvulus (GDV) is a critical illness and may be associated with coagulation derangements, including disseminated intravascular coagulation (DIC). This study's purpose was to evaluate baseline thromboelastography (TEG) in dogs with GDV and determine if TEG abnormalities are associated with mortality.

Dogs with GDV admitted for surgical therapy were enrolled. Blood was collected on admit for kaolin-activated TEG, fibrinogen, and platelet count. TEG was evaluated in all dogs and values were compared between survivors and non-survivors. Groups were compared using a Mann-Whitney Rank Sum with a p value of < 0.05 considered significant. The subset of dogs with abnormal TEG values was evaluated, and survivors compared with nonsurvivors. A Fisher's Exact Test calculated mortality between this subset and dogs without TEG abnormalities.

Twenty-seven dogs were enrolled. The median results for TEG for all 27 dogs are shown below.

Twenty-one dogs survived, 5 were euthanized, one died for overall mortality of 22%. Median TEG values did not differ between survivors and non-survivors amongst all dogs. Dogs with baseline TEG values outside the reference range had higher mortality (p = 0.044) than dogs without abnormalities.

Coagulation derangement was documented on presentation in dogs with GDV and is associated with survival. Pregnancy in women is associated with the development of a hypercoaguable state, which is considered protective against severe hemorrhage during labor and delivery. Multiple studies using thromboelastography have confirmed this in-vivo phenomenon. The purpose of the study was to evaluate if pregnant cats have similar changes.

Queens were recruited from the TNR program run by the Shelter Medicine program during June 2012; duration of pregnancy was not known specifically in cats. EDTA and citrated blood samples were collected from cats while under anesthesia for ovariohysterectomy and from non-pregnant cats as controls. Pregnancy was visually assessed as early, mid or late term. Kaolin-activated thromboelastography was performed using citrated whole blood. Platelet counts and hematocrit were determined from EDTA -anticoagulated blood samples. TEG variables, including R time, K time, angle, maximum amplitude (MA) and G, packed cell volume, and platelet count were compared between pregnant and non-pregnant cats using the Mann-Whitney U and Wilcoxon W test, with a P < 0.05 considered significant.

Nine mid and late term pregnant cats were enrolled and compared with 10 non-pregnant controls. There were no differences between hematocrit, platelet count or R time between pregnant cat and non-pregnant cats. Pregnant cats had significantly shorter K time (p = 0.01), greater angle (p < 0.01), increased MA (p < 0.01), and higher G (p < 0.01) compared to controls.

Pregnancy results in a hypercoagulable state as assessed by thromboelastography in cats. The relationship of these changes to surgery or complications of pregnancy is still to be determined. Large volume fluid resuscitation is commonly used in the emergency veterinary setting for treatment of hypovolemia. The purpose of the study was to investigate the effects of in-vitro hemodilution with lactated Ringers solution (LRS), 6% hydroxyethyl starch (HES), and fresh frozen plasma (FFP) on whole blood coagulation in dogs as assessed by kaolin-activated thromboelastography.

Six healthy dogs were used. Platelet count and hematocrit were normal. Whole blood was collected and diluted in-vitro at a 33% and 67% dilution of either LRS, HES or FFP. Kaolin-activated thromboelastography was performed on each sample as well as a simultaneous control. Thromboelastographic parameters R(min), angle(deg), and MA(mm) were measured and compared to the sample control for each dilution using mixed model methodology. Pairwise post-hoc comparisons were tested using Tukeys adjustment for multiple comparisons. A p value of < 0.05 was significant.

Prolongation in coagulation times were seen at both dilutions with LRS and HES. There was no significant difference in R times at the 33% dilution, but R time was significantly prolonged at the 67% dilution with HES (p = 0.004). MA was significantly decreased for LRS at both dilutions (p = 0.013, p < 0.001) and more profoundly decreased for HES (p < 0.001, p = 0.006). No significant difference in any parameter was found for FFP.

In-vitro hemodilution of whole blood with both LRS and HES but not FFP resulted in significant effects on coagulation with HES having a more profound effect. In-vivo evaluation of changes in coagulation with various resuscitation fluids is warranted and may be clinically relevant. Hemophilia A is a recessive sex-linked coagulopathy in dogs caused by deficiency or dysfunction of Factor VIII. We evaluated the characteristics of a group of dogs identified with hemophilia A.

We recruited information via an internet survey distributed to the ACVIM and ACVECC list serves, and to the VIN community about dogs identified with hemophilia A based upon a Factor VIII coagulant activity (FVIII:C) 25% of control canine plasma. Additionally, the relationships between% FVIII:C activity and outcomes (survival, bleeding episodes and severity, transfusions) were examined.

We included data from 41 dogs. Mixed breeds, German shepherd dogs and Labrador retrievers were most commonly affected, although 27 purebreds were reported. Clinical bleeding events, associated with teething, minor trauma, vaccination and elective surgical procedures most commonly prompted FVIII:C testing. Fourteen dogs had <2% FVIII:C, 11 dogs had 2-5% FVIII:C, 16 dogs had 6-25% FVIII:C. The owners of 37/41 dogs elected to treat after the diagnosis was reached. Affected dogs displayed similar rates and severity of spontaneous bleeding, without clear relationship to FVIII:C activity. Twenty-nine dogs received one or more transfusions; FVIII:C did not appear to predict transfusion need or number. Survival times were varied, with 6 having been treated for >5 years at the time of this survey.

Limitations of this survey include the potential for confounding factors contributing to hemorrhage and lack of long-term follow-up on many dogs after initial diagnosis. This study supports that hemophilic dogs are varied in their clinical course and FVIII:C activity may not predict the severity of clinical signs, transfusion needs, or long-term survival. Complete blood count (CBC) abnormalities described in cats with intestinal lymphoma (IL) may include anemia of chronic disease or regenerative anemia secondary to intestinal blood loss, neutrophilia secondary to inflammation, neoplasia, or stress, and monocytosis for chronic inflammation. The aim of this study was to evaluate hemoglobin and reticulocytes changes in cats with IL.

Medical records of cats with histopathologically confirmed IL and CBC performed by ADVIA-120 at the moment of presentation were retrospectively reviewed from 2009 to 2012. Complete blood counts performed to 20 healthy cats previously to sterilization procedure were used as control group (CG).

Eight cats were included in the study (5 neutered males, 2 spayed female and 1 entire female). The age ranged from 8 to 16 years. All were domestic short hair cats. Feline leukemia virus and feline immunodeficiency virus status were negative in all cats. High grade lymphoma (HGL) was diagnosed in 4 cats and low grade lymphoma (LGL) in other cases. Immunophenotyping was determinate only in 3 cats and were T cell lymphoma. Anemia was described in only 2 cats due to stomach ulcer.

Statistical analysis was performed by Kolmogorov-Smirnov and Mann-Whitney U tests. Hematological differences (p < 0.05) were found between both groups. Cats with IL compared to the CG showed a higher white blood cells (mean: 15,945 x10 3 /lL), neutrophils (mean: 11,687 x10 3 /lL) and monocytes (mean: 1,252 x10 3 /lL) count. A statistically significant lower hemoglobin (mean: 9,212 g/dL), hemoglobin distribution width (mean: 2,578 g/dL) and reticulocyte mean hemoglobin content (mean: 15,787 pg) were observed compared to control cats.

These results showed a decrease of hemoglobin levels in red blood cells (RBC) and reticulocytes in IL cats. No differences were described in reticulocytes or RBC number. The rest of the hematological changes observed were expected. This decrease of hemoglobin levels in RBC may be due to iron deficiency for chronic blood loss, insufficient gastrointestinal iron absorption or secondary to systemic inflammation. Further studies are needed to assess the signification of these findings in cats with IL. Limited information exists regarding the prevalence of anemia in hospitalized small animals. In-hospital development of anemia is recognized commonly in people, impacting hospitalization length, transfusion requirements and survival. The aim of this prospective study was to investigate the prevalence of anemia (PCV<35% dogs, PCV<30% cats) in critically ill dogs and cats in veterinary teaching hospitals.

A daily census of PCV and total solids (TS) of 388 critically ill animals (300 dogs, 88 cats) with numerous diseases (121 surgical, 267 medical) was performed. Patient information recorded included signalment, underlying disease, daily PCV/TS, estimated blood volume drawn and transfusion requirements. PCV/TS were recorded in 363 (93.5%) patients at admission.

Median PCV was 40% (range 9-80%), with 112/363 patients (30.9%) being anemic at admission (79/282 dogs, 33/81 cats). More than two PCV/TS were recorded in 201 patients (55%), with 111/201 (55%) developing anemia during hospitalization. In total 70/388 (18%) patients required blood products.

This pilot data illustrates that anemia is frequently documented in critical patients and that in-hospital development of anemia is under-reported. The development of anemia is likely multifactorial, dependent on underlying diseases, medications and phlebotomy techniques. Change in clinical practices (e.g. microsampling) has the potential to reduce the prevalence of anemia in the critically ill, which may improve outcome. Some operators report difficulty with insertion, advancement and specimen retention for bone marrow biopsy in cats using a standard 13ga Jamshidi needle, which has a bevel-tip cannula and bevel-tip stylet. We previously reported use of a 15ga needle with a trephine-tip cannula and trochar-tip stylet in cats 1 , which we hypothesized would be more easily inserted and advanced. The procedure was easier to perform than 13ga biopsy, but an acceptable specimen was only captured in 30% of cases 1,2 . A novel 13ga x 3.5in Jamshidi needle (CareFusion TJM3513) also has a trephine-tip cannula and trochar-tip stylet, as well as a Marrow Acquisition Cradle (MAC) placed into the cannula to promote specimen retention. We evaluated this needle for biopsies of the humeral head and femoral trochanteric fossa, and compared them to biopsies obtained from the femur with two other needles. Biopsies were taken under anesthesia from 16 healthy cats weighing 2.9-6.2 kg with body condition scores of 5-8 (scale 1-9). Biopsies were performed in randomized order by one investigator. Two biopsy sites were used with each cat; there was a maximum of two attempts at each site. Facility of site localization, needle insertion and needle advancement were scored from 1 (hardest) to 5 (easiest). Biopsy sections were reviewed by a pathologist unaware of biopsy site and cat, and quality scored from 0 (no hematopoietic tissue) to 5 ( ! 5 intertrabecular spaces free of artifact). Post-procedure, cats were given tramadol (4-5 mg/kg, PO, q12 h) for 3 days, and assessed for pain ( Significantly different by t-test (symbols refer to comparisons within columns).

The 13ga humeral biopsies with the novel needle were of higher quality than the femoral biopsies obtained with either the novel needle or with the 15ga needle, and of higher quality than the previously reported 15ga humeral biopsies. 1 There were no differences in facility scores. There was no swelling or reaction to palpation at biopsy sites (pain scores = 0 for all cats). In conclusion, in comparison to 15ga humeral biopsies previously obtained under the same conditions, 13ga humeral biopsies with the novel needle were as easily obtained and as well tolerated, but resulted in higher quality specimens. There were no differences between the novel 13ga needle and 15ga needle for femoral biopsy, both yielding lower quality specimens than 13ga humeral specimens. Thromboprophylaxis is an integral part of therapy in dogs with IMHA. 2,3-dinor-thromboxane B 2 (dTXB 2 , the primary metabolite of platelet-derived thromboxane B 2 in dogs) is a relevant indicator of ongoing in vivo platelet activation. We aimed to 1) compare in vivo platelet activity in dogs with IMHA to a reference population of normal dogs; and 2) evaluate suppression of platelet function in response to thromboprophylaxis in dogs with IMHA.

Dogs with IMHA received either low-dose aspirin (LDA, 0.5 mg/kg/d) or individually adjusted heparin (IAH, based on anti-Xa assay) as part of a prospective, randomized, doubleblinded clinical trial. Urine was collected at baseline and on day 7, and evaluated for dTXB 2 by ELISA. dTXB 2 concentrations were corrected for urine creatinine.

Urinary dTXB 2 /creatinine was markedly (10-18 fold) higher in IMHA dogs than in normal dogs. In contrast to normal dogs, there was no correlative relationship between platelet count and urinary dTXB 2 . No dog in either the LDA group (n = 8) nor the IAH group (n = 9) demonstrated suppression of platelet activity (as indicated by a > 50% decrease in urinary dTXB 2 ) on day 7 as compared to baseline.

Platelet activity is increased in dogs with IMHA as compared to of normal dogs. Neither LDA nor IAH suppresses platelet thromboxane release in vivo in dogs with IMHA. Babesia canis subtype vogeli is endemic in parts of the US, particularly in racing greyhounds. Infected greyhounds often appear clinically healthy, and in such dogs the organism can be difficult to detect. Since transfusion from infected donors can cause severe babesiosis in previously unexposed recipients, identification of occult infection in potential donor greyhounds is essential. In the past, splenectomy was used as a means of exposing occult infection in potential donor dogs. Our study was designed to assess the utility of using liposomal clodronate, a drug that causes marked but transient depletion of macrophages (in essence, a transient 'medical splenectomy'), to expose Babesia infection in greyhounds strongly suspected to harbor the organism.

Four healthy greyhound dogs known to have either recently harbored (2 dogs PCR positive on repeat testing, 1 kennel-mate dog transiently seropositive) or been exposed to (1 kennel-mate dog) Babesia canis, but currently negative when tested by blood smear examination, serology, flow cytometry and PCR, were confirmed to be in good health based on physical examination, complete blood count (CBC), serum biochemistry, and urinalysis. Consistent baseline negative status on assays for Babesia upon repeat testing was be confirmed over a 2 week period by daily smear examination, and flow cytometry and PCR approximately every second day. Flow cytometry (hydroethidine stain for the presence of the organism on red blood cells) and blood smear examinations (modified Wright's stain) were performed at MSU, and serology (IFA) and PCR were performed at the NCSU Vector Borne Disease Diagnostic Laboratory.

The 4 dogs were then given intravenous liposomal clodronate at a single low dose of 0.5 ml/kg. Potential side-effects were evaluated by daily physical examination, daily CBCs, and weekly biochemistry. Babesia status was monitored over a 2 week period post-treatment by daily smear examination, and by flow cytometry and PCR approximately every second day. The same study design was then repeated at a medium liposomal clodronate dose of 1 ml/kg and then, finally, at a high liposomal clodronate dose of 2 ml/kg except that, after the final dose of liposomal clodronate, Babesia status was monitored for an extended post-treatment period of 4 weeks by daily smear examination, by flow cytometry and PCR every second day, and by repeat serology.

Intravenous liposomal clodronate was well-tolerated apart from a transient mild to moderate fever (1-3 day duration), slight inappetance and mild diarrhea in several dogs, particularly at higher drug doses. No evidence of Babesia infection was detected by blood smear examination, flow cytometry, PCR or serology at any point during the study period.

Based on the results of our study, liposomal clodronate does not appear to be a reliable means of exposing occult babesiosis in greyhounds with a recent history of harboring the organism. Normal Dachshunds, like Greyhounds, are reported to have higher hematocrits than other dog breeds; however, there appears to be no objective information to support this observation. The purpose of this study was to determine whether red blood cell indices and numbers of circulating cells differ between Dachshunds and mixed breed dogs.

Retrospectively, packed cell volume and other red blood cell indices, total solids, white blood cell and platelet numbers were compared between 61 healthy Dachshunds and 60 mixed breed dogs that presented for wellness evaluations, dental prophylaxis or neutering to a university and a private clinic.

Dachshunds had a higher mean packed cell volume (52% versus 50%; P = 0.047), mean hematocrit (52% versus 48%; P = 0.0003), mean red blood cell count (7.7x10 6 /lL versus 7.1x10 6 /lL; P = 0.0004), and mean hemoglobin concentration (18.2 g/dl versus 16.8 g/dl; P = 0.0003) than mixed breed dogs. There were slight differences in hematocrit and hemoglobin concentration between clinics (P < 0.05). There was no evidence of a difference in the mean cell volume, mean cell hemoglobin concentration and total solids between breeds (P > 0.5). More Dachshunds than mixed breed dogs had red cell parameters outside the reference range: 29% versus 2% for hematocrit (P = 0.001); 40% versus 7% for hemoglobin (P = 0.0006); and, 26% versus 5% for RBC (P = 0.01). There were statistically significant but clinically trivial differences in some white cell numbers.

Dachshunds have higher packed cell volume, hematocrit, red blood cell count, and hemoglobin concentration compared with mixed breed dogs. Veterinarians should consider this variation when interpreting complete blood counts. The purpose of this prospective study was to compare the effects of three intravenous platelet-containing products on hemostasis. Nine client-owned dogs diagnosed with lymphoma who underwent an allogenic or autologous hematopoietic cell transplantation and developed severe radiation induced thrombocytopenia (< 20,000 platelets/lL) were randomized to receive a transfusion of fresh whole blood (FWB), apheresis derived platelet concentrate (APC) or frozen platelet concentrate (FPC). Data was collected and calculated at hospital admission, daily post thrombocytopenia (< 20,000 platelets/lL), immediately prior to platelet transfusion and at 1, 24 and 72 hours post-transfusion and included: vital parameters, clinical bleeding score, platelet count, kaolin-activated thromboelastography (TEG), prothrombin time, activated partial prothrombin time, fibrinogen, post transfusion platelet increment and post transfusion corrected count increment.

Differences in outcomes for FPC, APC and FWB products were tested using repeated measures analysis of variance (ANOVA) across all time collections. In the first stage of analysis, differences amongst all three products were tested. The second stage of post hoc tests were performed to evaluate the differences amongst the groups. After controlling for multiple comparisons, there was a significant difference (p < 0.003 after a Bonferroni correction) in the platelet counts between the three products, driven by the difference between the frozen (FPC) and fresh products (APC and FWB), with much higher platelet counts in the fresh products. These results have clinical impact on the selection of platelet transfusion products in critically ill thrombocytopenic patients. The platelet's importance in haemostasis, inflammation, and cancer metastasis is well established. However, new roles in several physiologic and pathological processes have only recently been unravelled. Human and rodent platelets have been shown to be key players in cancer angiogenesis, epithelial to mesenchymal transition, and the molecular balance of their granular contents has been shown to have prognostic and diagnostic potential. New mechanisms of platelet interaction with cancer cells potentiating metastasis have been reported. To understand comparative roles of canine platelets in these and other pathways, we analyzed the non-activated platelet proteome of healthy dogs.

Highly pure platelet samples were obtained from three dogs, and differential detergent fractionation was used for protein extraction. Mass spectrometry analysis yielded approximately 6,000 unique proteins. Of these, only 384 had previous experimental evidence of in vivo expression in any species. This yield is 3-fold larger than previously reported in human platelets, 12-fold greater than the rat, and 8-fold greater than the pig. Gene Ontology annotations and sub-cellular fractionation analysis of proteins was performed. Canine platelet subcellular organization did not appear to drive separation by different detergents as reported in other cells and tissues, yet most proteins tended to be extracted from a single fraction. By employing new fractionation procedures, we believe the sensitivity was improved leading to greater proteome coverage. This initial study contributes to a broader understanding of canine platelet biology, provides a new proteome for resting platelets and aids functional research identifying potential treatment targets and biomarkers. Hereditary factor VII (FVII) deficiency in Beagle and Alaskan Klee Kai dogs reportedly results from the missense mutation (p.G96E) in exon 5 of canine FVII genomic DNA. We have previously identified not only the G96E mutation in exon 5, but also a novel mutation (p.G285D) in exon 8 in a mixed breed dog (Papillon and Toy Poodle) that presented with spontaneous severe bleeding with markedly decreased FVII activity. The present study investigated the prevalence of these two mutations (G96E and G285D) in Papillon and Toy Poodle dogs. Blood DNA samples obtained from 23 Papillon and 64 Toy Poodle dog patients referred to Nihon University Animal Medical Center between October 2010 and April 2012 were investigated for the two mutations. Exons 5 and 8 of canine FVII were amplified from blood DNA samples using gene-specific primers constructed within introns. The mutation in exon 5 was also examined by direct sequence of the amplicon, while the mutation in exon 8 was analyzed by restriction fragment length polymorphism using VpaK11B I.

The G96E mutation in exon 5 was only observed as a heterozygous mutation in two Papillon dogs. On the other hand, the G285D mutation in exon 8 was not identified in dogs of either breed. Congenital FVII deficiency should thus be considered in Papillon dogs. The clinical impact of transfusing red blood cells (RBCs) after longer durations of storage is a major issue in human transfusion medicine, as some studies have found an association between administration of "older" RBCs (generally defined as >14 days of storage) and increased mortality, multiple organ dysfunction, and/or sepsis. This important issue has not yet been investigated in veterinary medicine.

The purpose of this retrospective study was to determine if the duration of packed RBC (PRBC) storage was associated with increased morbidity and mortality in dogs receiving PRBC transfusions at the Veterinary Hospital of the University of Pennsylvania between January 2001 and December 2010. Review of the Penn Animal Blood Bank logbook identified 3198 dogs transfused with PRBCs during this period. Medical records for 2155 patients have been reviewed, with the interim data analysis presented here. For this part of the study, survival was defined as alive/dead 30 days after the date of transfusion of the oldest PRBC unit to that patient.

Hemorrhage was the most common reason for PRBC transfusion (67%), followed by hemolysis (17%) and ineffective erythropoiesis (IE) (16%). Immune-mediated hemolytic anemia (IMHA) accounted for 91% of the dogs with hemolysis. The median duration of PRBC storage was 22 days (range 0-40 days). The median total PRBC volume administered during a hospitalization was 15 ml/kg (range 5-163 ml/kg). Survival was greatest for dogs with hemolysis (59%) followed by hemorrhage (51%), and IE (42%) (P 0.004). There was no difference in 30-day survival when comparing dogs that received only PRBC units stored <14 days ("fresh") to those receiving PRBC units stored >14 days ("old"). When considering all dogs regardless of cause of anemia, there was no association between duration of PRBC storage or total volume of PRBCs administered and 30-day survival following oldest PRBC transfusion. However, longer duration of PRBC storage was associated with development of new or progressive multiple organ dysfunction (P = 0.03), coagulation failure (P = 0.0017), disseminated intravascular coagulation (P = 0.02), and thromboembolic disease (P = 0.007) post-transfusion. In addition, a logistic regression model indicated that for the subset of dogs with hemolysis, longer duration of PRBC storage is a negative risk factor for survival; for every 7 day increase in storage, there is a 0.64 lesser odds of 30-day survival (95% CI 0.51-0.804; P < 0.001). Duration of PRBC storage does not appear to be a major contributing factor to mortality in the general canine population receiving PRBC transfusions, although it may be associated with progression of multiple organ dysfunction and coagulopathies. Also, duration of PRBC storage may be an important consideration for mortality in patients with hemolysis, particularly IMHA. Prospective studies are warranted to further investigate the clinical impact of duration of PRBC storage on morbidity and mortality post-transfusion in dogs. Transfusion medicine is a component of high quality veterinary care. Approximately 55 ml of blood can be removed safely from domestic cats, resulting in one approximately 20-25 ml unit of packed red blood cells (pRBC). Because of this, one pRBC unit is often inadequate to effectively treat a patient. When a patient receives pRBC from multiple donors, there is an increased risk of transfusion reactions; this is particularly true in patients needing multiple transfusions.

Domestic and non-domestic cats share the A-B blood type system. If blood from non-domestic felids is compatible with that of domestic cats, a feline patient could be administered a therapeutic dose of pRBC from only one donor, thus decreasing risk of transfusion reactions. The purpose of this study was to evaluate compatibility of domestic and non-domestic cat blood for transfusion purposes.

Blood from 49 domestic and 25 non-domestic cats was typed using Rapid Vet H â typing cards. Domestic cats were 43 type A, 5 type B and one could not be typed due to hemolysis. Nondomestic felids were 22 type A and 3 type B. All the latter were cougars. Blood from each domestic cat was crossmatched with blood from 1-6 non-domestic felids. One hundred eleven crossmatches were performed; 78 were compatible and 33 incompatible (tables below). Flow cytometry documented no donor immunoglobulin bound to the surface of recipient RBC.

Results suggest that non-domestic felids may be compatible blood donors for domestic cats, if appropriate cross matching is performed first.

Compatible crossmatch tests between blood type A and B domestic and non-domestic felids Blood transfusions have gained an important role in feline intensive care medicine which, in turn, has led to an increased demand for point-of-care blood-typing methods.

The objective of this prospective study was to evaluate a novel immunochromatographic blood-typing test (RapidVet â -H IC feline) (IC) for the feline AB blood group system. EDTA-anticoagulated blood samples from 105 sick and healthy cats were typed comparatively with the IC test and 2 other laboratory methods, a tube agglutination (TUBE) and a gel column test (DiaMed ID â ) (GEL). The samples were between 0 and 10 days old (median 3) and were tested for hemolysis and agglutination; the hematocrit ranged from 7 to 57% (median 40).

The TUBE and the GEL method were in agreement for all blood types in 100% of the tests. 85 A, 17 B and 3 AB samples were determined. The IC test correctly identified 80 A samples, 4 were typed as AB and 1 was inconclusive (weak indicator line). All 17 B samples and 2 AB samples were correctly typed; one of the AB samples was inconclusive. The sample quality had no influence on test performance.

The agreement of the IC-test with the control methods was 96.1% for the 103 conclusive tests. It is suggested that AB results be reconfirmed with a TUBE or GEL laboratory method and that a "back-typing" will be performed with plasma from B-samples to detect the presence of alloantibodies. Given its accuracy and ease of use, the IC test can be recommended for clinical settings. The aim of the present study was to evaluate hematologic changes in the red blood cells (RBC) and reticulocytes count in stray cats with or without sedation. In previous studies, RBC count, haemoglobin and packed cell volume (PCV) resulted significantly decreased after treatment with different sedative drugs (midazolam, butorphanol, dexmedetomidine, ketamine), but reticulocytes were not reported. Sedation is frequently necessary to handle stray cats, since they are often wild, aggressive and unmanageable. Blood sampling can be frustrating and time consuming, even injurious for both the employee and the animal. In this study, when manual restrain resulted satisfactory, sedation was avoided. (1,26) P value 0,818 0,809 0,000 0,146 0,522 64 stray cats were clinically classified as healthy or ill animals (the most common diseases were dermatophytosis, chronic rhinitis and conjunctivitis, gingivitis, unspecific wounds). Blood was obtained with manual restrain or medetomidine sedation. No serious problems regarding safety occurred. Blood samples were drawn from the jugular, cephalic or femoral vein and kept in EDTA tubes. The RBC and reticulocyte count were performed using the laser-based haematology analyser ADVIA 120 + (Siemens Diagnostics), that permits automated and accurate reticulocyte count, using lasers mark cell samples with fluorescent dye which marks RNA. A Student t Test was used to evaluate differences between groups.

No statistical significant differences were observed between healthy and ill animals for any parameter studied. Cats under sedation had statistically significant lower reticulocytes count than non sedated ones, no significant changes were observed on any other RBC parameter. Accumulation of the circulating blood cells in the spleen or other reservoirs have been seen in animals due to a decrease in the sympathetic activity. A previous study in dogs attested the use of medetomidine leads to an increase in splenic volume, but no correlation between PCV and splenic size was found, supposedly due to sequestration of RBC in nonsplenic sites. Also, there may be a shifting of fluid from extra to intravascular compartment in order to maintain normal cardiac output during sedation. RBC sequestration and fluid shifting may contribute to the reticulocyte count decrease observed in sedated cats. Previous studies have shown that canine patients with congenital portosystemic shunts (CPSS) have changes in serum concentrations of both pro and anti-coagulant proteins. The objective of this study is to conduct thromboelastography (TEG) analysis to determine the coagulation status in dogs with CPSS. Twentyone client owned dogs with CPSS confirmed by ultrasound or nuclear scintigraphy were enrolled prospectively. Complete blood count, biochemical panel, conventional hemostatic tests (platelet count, prothrombin time (PT), activated partial thromboplastin time (aPTT), quantitative fibrinogen, antithrombin activity (AT), protein C (PC), d-dimers, and factor VIII) as well as kaolin activated TEG were measured. Dogs with CPSS had significantly shorter k values and increased angle, maximum amplitude (MA) and G values compared to normal dogs. On conventional coagulation testing, dogs with CPSS had significantly prolonged PT, lower platelet counts, AT, and PC activity and increased concentration of d-dimers and factor VIII. G value was significantly associated with platelet count and fibrinogen level. Evaluation of G value defined 9/21 dogs with CPSS that were hypercoagulable. These dogs were more likely to have uncontrolled hepatic encephalopathy than CPSS dogs that were normocoagulable. TEG analysis detected hemostatic abnormalities consistent with a hypercoagulable state in dogs with CPSS. The presence of hepatic encephalopathy may increase the risk of a hypercoagulable state in these dogs. CRITERIA (2005 CRITERIA ( -2012 . A Wayne, P Ewing, M Whelan. Angell Animal Medical Center, Boston, MA.

Hy's Law defines clinical pathology criteria for severe druginduced hepatotoxicity that predicts a case fatality rate of 10% or higher without liver transplant. To date, Hy's Law has been used exclusively in human medicine. There are no studies in animals to determine whether or not similar criteria and outcomes can be identified in animals. Hy's Law is defined as:

Serum alanine transaminase (ALT) or aspartate transaminase (AST) ! 3 times the upper limit of normal (ULN) AND serum total bilirubin levels ! 2 times the ULN without evidence of cholestasis (defined as serum alkaline phosphatase (ALP) activity > 2 times ULN) SECONDARY TO drug therapy (known drug history) when no other reason can be found to explain the combination of increased ALT, AST and total bilirubin (viral hepatitis, preexisting liver disease, etc).

In people, simultaneous elevation of ALT and total bilirubin is a sensitive marker for severe liver injury and is associated with high mortality rates. The goal of the study is to determine if Hy's Law can be identified in dogs and, if so, if the prognosis is similar to that in humans. We extended our evaluation to include dogs that met the clinicopathologic criteria, but were not considered to have a drug-induced hepatopathy in order to better define prognostic indicators for a variety of severe hepatic disorders in dogs. We predicted that patients with greater elevations in their liver enzymes would have less favorable outcomes than those with mild elevations in liver parameters, especially bilirubin elevations.

Laboratory data at Angell Animal Medical Center from January 1, 2005-July 1, 2012 was searched for dogs that met Hy's Law laboratory criteria (ALT or AST ! 3 ULN and serum total bilirubin ! 2 times the ULN and excluding dogs with ALP activity > 2 times ULN). For identified patients, medical records were searched to determine the underlying cause (if able) and outcome (survival to discharge). Mean or median laboratory values, including AST, ALT, total bilirubin and GGT, were compared between survival groups. 566 dogs met the laboratory criteria of AST ! 3 times the ULN and serum total bilirubin levels ! 2 times the ULN. When dogs with ALP activity > 2 times ULN were excluded, 70 cases remained. Three cases were excluded because medical records could not be located. 49/67 (73%) of dogs that met the laboratory criteria survived to discharge. 18/67 (27%) did not survive to discharge. 3/67 dogs were on medications that were clinically suspected as the cause of their liver enzyme elevations. Of the 3 dogs, two survived and one did not. ALT was found to be statistically significantly higher in the group that did survive to discharge when compared to the group that did not survive to discharge (median value 374 vs 169 respectively, p value 0.02). AST, GGT and total bilirubin were not statistically significantly different between the survival groups.

Our data indicate that Hy's Law laboratory criteria, when applied to dogs, does carry a high mortality rate, as reported in humans. There were too few dogs with drug exposure to draw any definitive conclusions about mortality in dogs with drug exposure. The ALT was higher in survivors than non-survivors, which was unexpected. Further research is necessary to elucidate the cause. Hy's Law as defined in people was applied to this data set, however future studies to include dogs with ALP activity > 2 times ULN may be more clinically relevant due to the steroid induced elevations of ALP seen in dogs. Mortality rates in people that meet the laboratory criteria for Hy's Law and have known drug exposure ranges from 10-50%. In this study of 67 dogs that met the Hy's Law laboratory criteria, the mortality rate is 27%. Many dogs with liver disease have increased hepatic copper (Cu) concentrations. Abnormal hepatic Cu may be due to metabolic, cholestatic or nutritional factors. In the 1930s, Cu concentrations were reported to be 50-75 ppm dry weight (DW), which is similar to the normal range for humans. Today, normal canine hepatic Cu concentrations range from 200-450 ppm DW. A dog consuming an average commercial maintenance diet ingests about twice the amount of Cu compared to that contained in the average human diet. We hypothesized that the increase in hepatic Cu levels over time may be due to excessive Cu in commercial dog foods.

Liver biopsies obtained from age-and sex-matched purposebred dogs (n = 9) and feral dogs (n = 9) from Malaysia and Nicaragua were analyzed for Cu and Zn levels via flame atomic absorption spectroscopy. Both groups had similar body condition scores (3.5-4/9). The purpose-bred dogs had no evidence of liver disease based on measurement of liver enzymes and bile acids. The feral dogs were presumed to survive on a diet of foraged food and human food scraps.

Median hepatic Cu levels for the purpose-bred dogs versus the feral dogs were 472 ppm (199-997 ppm) and 152 ppm (69-370 ppm), respectively (p = 0.004). Median hepatic Zn levels for the purpose-bred versus the feral dogs were 181 ppm (157-320 ppm) and 249 ppm (153-417 ppm), respectively (p = 0.039). There was no correlation between Cu and Zn levels for either group. The maintenance diet fed to the purpose-bred dogs contains 23.35 mg Cu per kg of diet on a dry matter (DM) basis and 242.16 mg Zn per kg of diet on a DM basis, both within AAFCO guidelines.

Hepatic Cu concentrations in normal purpose-bred dogs were approximately three times that of the feral dogs. In 6 of the 9 purpose-bred dogs, hepatic Cu concentrations were outside the normal range with one dog approaching a Cu level at which Cu chelation therapy might be considered in a dog with hepatitis. These findings suggest that dietary Cu content influences hepatic Cu concentrations and may play a role in Cu-associated hepatic disease. Autosomal dominant polycystic kidney disease (ADPKD), the most common inherited disease in cats, is also a common inherited disorder in humans. Initially described in cats of Persian ancestry, it also occurs in cats of other lineages. A causal point mutation in Polycystin 1 (PKD1) is diagnostic. Feline ADPKD varies in phenotypic expression but usually is evident using ultrasonographic renal imaging (cystic lesions). While renal ADPKD manifestations are well documented, a subset of cats from the initial ADPKD colony concurrently demonstrated biliary fibrosis and hyperplasia consistent with a fibrocystic biliary malformation. Rare single biliary cysts were also described (lined by flattened or dysplastic biliary epithelium). To our knowledge, fibrocystic biliary disease consistent with a ductal plate malformation in the absence of polycystic renal lesions has been defined in humans, but has not been definitively associated with the feline PKD1 mutation. We hypothesize that additional fibropolycystic genetic variants exist in cats. Using DNA extracted from paraffin embedded tissue (n = 36), whole blood (n = 6), and fresh frozen tissue (n = 1) histopathologically confirmed hepatic fibropolycystic phenotype (n = 29), polycystic kidney phenotype (n = 8), and unaffected controls (n = 6), we investigated whether the point mutation causal to feline AD-PKD (PKD1 exon-29) exists in cats with fibropolycystic hepatic malformations lacking cystic renal lesions. A feline validated restriction fragment length polymorphism test for the PKD1 and DNA sequencing were used to detect heterozygotes. Findings confirm that some cats with fibropolycystic liver (88.5%) or polycystic kidney (62.5%) lesions lack the PKD1 point mutation. In dogs, hepatocellular carcinoma (HCCA)/adenoma (HCA) are slowly progressive, locally invasive, and rarely metastasize. Androgen exposure affiliates with HCCA in humans, as suspected in Scottish Terriers. This retrospective study investigated relationship of HCCA/HCA with vacuolar hepatopathy (VH), adrenomegaly, and adrenal hyperactivity in diverse dog breeds (excluding Scottish Terriers).

Cornell's electronic databases identified 94 dogs with HCCA/HCA. Signalment, clinical signs, clinicopathologic features, hepatic and adrenal ultrasonographic images, histologic findings, and adrenal function tests were recorded. Of 33 breeds (4% were small [<10 kg], 53% medium [10-20 kg], 43% large [>20 kg] dogs), 38 were males (2 intact) and 56 females (1 intact). Mixed breeds (29/94) and Golden Retrievers (11/94) were most common. Median age was 12 (2-23) yrs and weight 23.9 (6.8-42.0) kg. Clinical features included: lethargy (28%), inappetence (28%), hepatomegaly (27%), abdominal mass (19%), PU/PD (18%), weight loss (17%), restlessness (15%), vomiting (13%), diarrhea (9%), fever (7%), and distended abdomen (5%). Median hematologic parameters were within reference intervals. Median liver enzymes exceeded reference intervals, notably ALP (9x); 76%, 51%, 95%, and 67% had increased ALT, AST, ALP, and GGT, respectively. Hepatic imaging disclosed hyperechoic parenchyma with variably-sized masses (0.5-30 cm; 73% >5 cm). Lesions were more common in left (39.5%) and right (25.9%) than central (11.1%) lobes; 23.5% were multifocal. Cytology confirmed HCCA/HCA in 27% of aspirates. VH occurred in 83% and adrenomegaly (>0.6 cm dogs 10 kg; >0.7 cm dogs >10 kg) in 29/43 (67%). Adrenal hyperactivity (cortisol >22 lg/dL 1-hr after 5 lg/kg ACTH IV, cortisol >1.4 lg/dL 8-hrs after 0.015 mg/kg dexamethasone IV, and/or >2-fold increased baseline or ACTH-stimulated androstenedione, progesterone or 17-OH progesterone) was confirmed in 15/22 (75%) dogs; 9/11 dogs (82%) had increased androstenedione.

Paraneoplastic enzyme activities, tumor affiliation with diffuse VH, adrenomegaly, and adrenal hyperactivity (including sex hormones) suggest possible association with steroidogenic hormones. This phenomenon warrants further investigation. The liver plays an important role in vitamin-B metabolism. In humans acute hepatitis, cirrhosis, and hepatocellular carcinoma are associated with increased serum concentrations despite a reduction in hepatic cobalamin stores. The effect of hepatic disease on canine cobalamin metabolism has not previously been studied. The objective of this study was to evaluate the cobalamin status of dogs with different kinds of hepatic disease.

Thirty-five dogs undergoing hepatic biopsy and 5 dogs euthanized due to hepatic disease at Gulf Coast Veterinary Specialists were prospectively enrolled into the study between 3/1/12 and 10/30/12. Dogs that had received cobalamin supplementation were excluded. The history, physical examination findings, results of the diagnostic evaluation, and diagnosis were recorded for each patient. Dogs were divided into 3 groups-group 1: vascular anomalies; group 2: non-neoplastic hepatic disease; and group 3: hepatic neoplasia. Serum cobalamin concentration was measured using a chemiluminescent assay (Immulite 2000, Siemens). Serum methylmalonic acid (MMA) and homocysteine (HCY) concentrations were measured using in-house gas chromatography-mass spectrometry assays. Serum cobalamin, MMA, and HCY concentrations were compared between the groups of dogs using ANOVA or Kruskal-Wallis tests as appropriate. Post-hoc testing was performed using Tukey's tests or Mann-Whitney U tests. Statistical significance was set at p < 0.05.

Dogs were assigned to the 3 groups as follows: 7 (17.5%) to group 1, 20 (50%) to group 2, and 13 (32.5%) to group 3. Overall 15 dogs (37.5%) had serum cobalamin concentrations greater than the upper limit of the reference interval (908 ng/L). Of these, 4 dogs (26.7%) had serum MMA concentrations greater than the upper limit of the reference interval (1193 nmol/L) suggesting cobalamin deficiency on a cellular level. The median (min-max) serum cobalamin concentrations were 1000 (293-1121), 642 (316-1099), and 398 (237-2311) ng/L for groups 1, 2 and 3 respectively (p = 0.13). The median serum MMA concentrations were 410 (277-2806), 936 (446-2184), and 1011 (477-1792) nmol/dL for groups 1, 2, and 3 respectively (p = 0.25). The median serum HCY concentrations were 4.8 (2.4-8.0), 9.2 (0.9-23.2), 18.8 (6.6-45.8) lmol/L for groups 1, 2, and 3 respectively. Group 3 had a significantly higher HCY concentration than groups 1 (p = 0.001) and 2 (p = 0.007). There was no significant difference between groups 1 and 2 (p = 0.26).

Increased serum cobalamin concentrations were common in dogs with hepatic disease. However, the relevance of this and of dogs with concurrently increased serum cobalamin and MMA concentrations is not known. The groups of dogs with vascular anomalies and non-neoplastic hepatic disease may have had lower mean HCY concentrations than the hepatic neoplasia group as they were more likely to have had altered amino acid metabolism due to hepatic insufficiency. The purpose of the study was to determine if the administration of ursodeoxycholic acid affects biochemical parameters and bile acid profiles in healthy adult canines.

Twenty dogs owned by students, faculty, and staff of the veterinary teaching hospital were enrolled in the study. Signed owner consent was obtained. Dogs were determined to be healthy based on normal physical exam, serum biochemical profile, complete blood count, urinalysis, ultrasound of the liver and biliary system, and fasting and post-prandial bile acids (time 0). All dogs received between 9.9 and 15.7 mg/kg of ursodeoxycholic acid once daily for 6-8 weeks. Doses were administered at night. The morning following the last dose (time 1), dogs were re-evaluated via physical exam, serum biochemical profile, complete blood count, and bile acid profile. Time 0 and time 1 values were compared for ALT, ALP, triglycerides, cholesterol, total bilirubin, and bile acids. A paired t-test was used with null hypothesis of zero mean difference between paired observations. The test assumes that the differences are normally distributed.

Only fasting bile acids at time 1 differed significantly from time 0 (p = 0.032, mean difference: -1.55 and 95% confidence interval -2.9547, -0.1453), although only 2/20 dogs had a fasting bile acids value outside of the reference range (11.7 and 10 umol/ L, reference range 0 -9 umol/L). On repeat bile acid profile 72 hours later, all values were normal.

Long-term administration of ursodeoxycholic acid does not significantly alter serum biochemical parameters or bile acids in healthy canines. The optimum method of liver biopsy in the dog is unknown. The potential for increased morbidity and cost of more invasive methods of biopsy must be balanced with the amount of tissue obtained. The purpose of this study was to identify the method of liver biopsy that most consistently produced adequate samples and accurate histopathology results.

We tested the hypothesis that liver biopsy specimens obtained from the left lateral liver lobe with an 8 mm surgical punch, laparoscopic biopsy forceps, and percutaneous 14 gauge trucut needle have similar histopathologic diagnoses to larger excised liver samples. The study was conducted using 70 cadaver dogs undergoing necropsy. After biopsy acquisition via the three techinques, two excised large tissue samples (at least 2 cm x 2 cm x 1 cm) were collected near the center of the left lateral lobe to serve as a gold standard to which other biopsies were compared. All biopsy samples were collected from within 5 cm of each other. All samples were evaluated by a board certified veterinary pathologist who assigned a diagnosis, recorded the histopathologic features, and numbers portal triads in each sample. If the two large gold standard samples had differing diagoses, the samples from that case were excluded from analysis.

The mean number of portal triads produced using each sampling method included 2.9 in trucut biopsies, 3.4 in laproscopic forceps biopsies, 12 in surgical punch biopsies, and 30.7 in the gold standard samples. The proportion of biopsies in which the histological diagnoses were in agreement with the gold standard samples were 66% of trucut biopsies, 60% of laproscopic cup samples, and 69% of surgical punch samples. Cohen's kappa coefficients for each technique were similar (trucut, laparoscopic, and surgical biopsies were 0.59, 0.52, and 0.62, respectively). None of the biopsy methods had a corresponding kappa coefficient indicating strong agreement.

Each biopsy method evaluated resulted in modest agreement with the gold standard. These findings call into question the accuracy of any single liver biopsy. Obtaining multiple samples from a liver lobe may be of greater imporance than the method of biopsy. Age-related immunosenescence (ARI) occurs in many species, including cats. This study explored differentially expressed genes (DEG) within feline leukocytes that might be related to ARI. Following an overnight fast, blood samples were collected from 36 healthy, adult, neutered cats equally divided by age as young (<7 yrs), middle-age (7-11 yrs) and old (>11 yrs). Leukocyte differentials were determined on fresh blood, and leukocyte RNA was purified and stored (-80C) until analysis via microarray. Gene ID, signal median, background median and quality control flag information were extracted from the raw data, with gene expression determined as the difference between signal and background noise. Age effects were determined via ANOVA, with p < 0.05 considered significant. Neutrophil percentages increased (46.6 a AE 4.1, 54.6 a AE 3.4, and 69.3 b AE 2.8, respectively for young, middle-age and old; p < 0.01), and lymphocytes decreased (43.9 a AE 5.2, 37.4 a AE 4.0, and 23.3 b AE 2.8, respectively for young, middle-age and old; p < 0.05), with age. Age-related DEG was identified for 104 genes (p < 0.05):15 differed between young and middle-age, 45 between young and old, and 88 between middle-age and old. They fit into 5 groups based on their biological pathways, although with considerable overlap: cell cycle control; immune/ inflammation; metabolism; cell signaling; and all others. Among the affected pathways, ALOX5 (up-regulated with age) is part of the leukotriene process and functions in immunosenescence; DNAJA2 and Sirtuin6 (downregulated with age) are implicated in T-cell quiescence and cellular longevity, respectively. These data demonstrate how agerelated changes in leukocyte gene expression contribute to immunosenescence in cats. Non-erosive immune-mediated polyarthopathy (IMPA) in dogs is diagnosed with arthrocentesis and synovial fluid analyses, and serial joint arthrocentesis is recommended to monitor response to therapy. However, repeated arthrocentesis is invasive and requires sedation, so treatment adjustments are often made without any objective measures of disease remission. We hypothesized that one of several pro-inflammatory molecules, C-reactive protein (CRP), interleukin-6 (IL6), interleukin-8 (IL8 or CXCL8), and tumor necrosis factor alpha (TNFa) would show promise as a non-invasive plasma marker of joint inflammation, and resolution, in dogs with IMPA.

10 dogs with cytologically confirmed neutrophilic polyarthritis (IMPA) were enrolled in this pilot study. Plasma in EDTA was drawn at baseline for measurement of CRP, IL6, IL8, and TNFa, as determined by sandwich ELISA, with 6 healthy dogs used as controls. Owner questionnaire, force plate analysis, and accelerometry collars were used to assess lameness and mobility. All dogs were treated with 50 mg/m 2 of prednisone. At 2 and 4 weeks of treatment, joint fluid analyses, plasma markers and lameness evaluations were repeated, along with joint fluid analyses.

After 2 and 4 weeks of treatment, 5 of 10 dogs had normal joint fluid cytology (TNCC < 3000/ul) in all joints examined. Of the remaining 5 dogs, all had a reduction in the number and severity of affected joints. Mobility as measured by accelerometry significantly increased in all dogs over 4 weeks. Plasma CRP was significantly increased in IMPA dogs at the time of diagnosis (median 51.0 ug/ml, range 30.0-70.8), compared to healthy controls (median 7.7 ug/ml, range 0.7-11.5; P = 0.006). CRP concentrations fell dramatically during treatment in all dogs, and were essentially undetectable by week 4 (median 0.0 ug/ml, range 0.0-2.2). IL-6 was also significantly increased in IMPA dogs at the time of diagnosis (median 38.8 pg/ml, range 13.7-867.5), compared to healthy dogs (median 0.4 pg/ml, range 0.0-11.0; P = 0.005). With treatment, Il-6 concentrations fell in all dogs and were significantly decreased by 4 weeks (median 0.0 pg/ml, range 0.0-83.1; P = 0.008). In contrast, IL8 was not different in IMPA dogs (median 147.7 pg/ml, range 18.8-1000 pg/ml), vs. healthy dogs (median 147.1 pg/ml, range 83.3-1049 pg/ml; P = 0.84), and did not change at the 2 and 4 week evaluations. TNFa was not detected in the sera from any of the IMPA or control dogs.

The results of this pilot study indicate that serum CRP and IL-6 show promise as objective but non-invasive markers of clinical disease in dogs with IMPA. These changes parallel improvements in clinical status and mobility by accelerometry, but precede complete cytologic resolution of synovial inflammation in some dogs. These results provide the rationale for a prospective study to determine the value of using plasma CRP and IL-6 concentrations to stage disease, provide prognostic information, and guide adjustments in immunosuppressive drug therapy in dogs with IMPA. Age-related immunosenescence (ARI) has been documented in many species, including dogs. The objective of this study was to explore differentially expressed genes (DEG) within canine leukocytes that might be related to ARI. Following an overnight fast, blood samples were collected into lithium-heparin tubes from 10 young (2-4 years of age) and 7 old (10-11 years of age) healthy, neutered Labrador retriever dogs. Leukocyte RNA was purified and stored at -80C until analysis via microarray. Gene ID, signal median, background median and quality control flag information were extracted from the raw data, with gene expression determined as the difference between signal and background noise. Age effects were determined via ANOVA, with p < 0.05 considered significant. Principal component analysis (PCA) of the data was performed using Partek software. Among the 147 DEG (p < 0.05) by age, 111 could be identified via existing annotation. They fit into 6 groups based on their recognized functions, although with considerable overlap: cell cycle control and cell proliferation, including up-regulation of TGF-beta and SUV39H1; immune/ inflammation; metabolism, especially glucose metabolism; stress, including genes involved in apoptosis; cell signaling, with 20 different genes involved in the Notch pathway differentially expressed; and all others, including 30 genes involved in RNA splicing and 13 involved in catabolic processes. PCA of the transcriptomic data showed a clear separation between age groups. Many of the identified changes are associated with decreased proliferation and senescence-associated processes. These data demonstrate how agerelated changes in gene expression contribute to immunosenescence in dogs. Immunosuppressive doses of glucocorticoids are considered first-line treatment for canine primary immune-mediated polyarthritis (IMPA) but adverse effects are common and can negatively impact patient quality of life. There is a need for alternative treatments. Cyclosporine inhibits T-cell function and has gained widespread use in dogs to treat severe or refractory immune-mediated disease. The purpose of this study was to compare the efficacy of oral cyclosporine to that of oral prednisone for the treatment of canine primary IMPA.

This was a prospective, randomized clinical trial. Twenty client-owned dogs diagnosed with IMPA were randomized into two groups. Group 1 dogs were treated with prednisone (1 mg/kg, PO, q12 h). Group 2 dogs were treated with cyclosporine (5 mg/ kg, PO, q12 h). Group 2 dogs were also treated with carprofen (2.2 mg/kg, PO, q12 h) for 7 days for analgesia. History, physical examination and arthrocentesis were performed on days 0, 14, 45, and 90. Trough cyclosporine plasma levels were determined for Group 2 dogs on day 7. Treatment failure end points for either group were failure of clinical improvement by day 14, lack of cytologic improvement by day 45, or a need to cross over due to adverse effects.

Clinical treatment success occurred in 7/10 prednisone-treated dogs and 6/10 cyclosporinetreated dogs (P = 1.00). Complete cytologic resolution on day 45 occurred in 5/10 prednisonetreated dogs and 8/10 cyclosporine-treated dogs (P = 0.35). Adverse effects of cyclosporine treatment were primarily gastrointestinal.

Cyclosporine monotherapy appears to be a suitable alternative to prednisone for treatment of IMPA. IL-17-producing helper T cells (Th17) were recently recognized in humans and in mice as a fourth component that regulates adaptive immune responses. Th17 cells function by promoting inflammation that counterbalances the activity of regulatory T cells (Tregs). The relationships of Th17 cells to health and disease remain poorly understood, but data from laboratory animals suggest that Th17 cells could contribute to a variety of immune-mediated diseases. The existence of Th17 cells in the dog has not been conclusively established. Although expression of mRNA encoding orthologs of IL-17 and the IL-17 receptor was documented in tissues from dogs with arthritis, inflammatory bowel disease, and lymphoma, there was no association between IL-17 gene expression and disease phenotype. However, robust assessment of the role of Th17 cells will require methods that can measure the frequency of these cells in health and disease in balance with other lymphocyte subsets. For this reason, we validated three antibodies against human and murine IL-17 to recognize IL-17 producing canine T cells. Peripheral blood mononuclear cells were stimulated with the mitogen Concanavalin-A (ConA) for 4-6 hr, secretion was inhibited by disrupting Golgi function, and production of IL-17 was examined in T-cell subsets using intracellular stain-ing. Accumulation of intracellular IL-17 was observed in stimulated CD4 and CD8 T cells using each of the three antibodies tested. The effect of inflammatory and regulatory cytokines on this Th17 response, as well as the relationships between Th17 and Tregs in canine peripheral T cells will be discussed. Immune-mediated hemolytic anemia (IMHA) is an autoimmune disease in which the immune system reacts against red blood cells just as it would against a foreign bacteria or virus. Although the causes of IMHA are still not well understood, IMHA can result from a primary immune system problem or result secondary to some other condition. Naturally occurring CD4 + , CD25 + regulatory T cell (Treg) that is produced by the normal thymus as a functionally mature T-cell subpopulation has a function that maintain immunologic self-tolerance and inhibit immune response. We performed this study to investigate the level of Treg in spontaneously occurring canine IMHA.

Blood from 7 IMHA dogs having no previous immune suppression treatment history were evaluated by flow cytometry. CD4 + / CD25 + /FoxP3 + lymphocytes were regarded as canine Tregs using flow cytometry. Interestingly, the percentage of CD4 + / CD25 + /FoxP3 + T cells in total lymphocytes from 7 IMHA dogs were increased (24.64%AE10.28%) compared with healthy 8 beagle dogs (7.66%AE1.65%, p < 0.01). Based on many article review. Canine sterile neutrophilic dermatitis is a rarely reported disease in dogs that resembles Sweet′s syndrome (SS) in humans. SS is described as an acute febrile neutrophilic dermatosis with cutaneous and extracutaneous manifestations. The diagnosis is based on the presence of two major criteria (acute onset of typical cutaneous lesions and compatible histopathological findings) and two minor criteria (history of a previous disease/vaccination/pregnancy associated with that syndrome; fever; leukocytosis; and rapid response to glucocorticoid therapy).

Two dogs, English Mastiff(dog A) and Labrador (dog B), male and female, seven and six months old, respectively, were referred to the Veterinary Teaching Hospital at University of São Paulo with sudden onset of multiple erythematous papules and nodules, perypheric neutrophilia, fever, lameness, arthralgia and myalgia. Conjunctivitis and blepharitis were also observed in the dog B. Skin biopsies of lesions (A) revealed neutrophilic dermatitis and large number of non-degenerate neutrophils was seen in the FNA cytology of skin nodules of the second dog (B). Microorganisms were not observed in both cases nor isolated on bacterial culture. One dog presented immune mediated hemolytic anemia shortly before the onset of cutaneous lesions. No other abnormalities could be found on physical examination, images or serum biochemistry. Glucocorticoid (prednisolone 2 mg/kg BW bid) resulted in improvement of cutaneous lesions over the first 48hs and complete clinical remission within the first week of treatment. As a common feature, the vaccination schedule has been recently completed, with canine vaccine (DHPPiCL, three dosis), Bordetella bronchiseptica vaccine, rabies vaccine and Giardia vaccine. The rapid onset of cutaneous and extracutaneous signs occurred soon after the last dose of Giardia vaccine (five to seven days) and progressively worsened throughout fifteen days. The two cases fit the criteria recommended for diagnosis of Sweet syndrome in humans and no trigger conditions as neoplasia, concurrent disease nor association to drugs could be identified other than a strong temporal relationship to vaccination with multiple antigens.

A SERIAL CD4 + /CD25 + /FOXP3 + REGULATORY T CELL CHANGES IN CANINE SEPTIC MODEL. Jun-hwan Kim 1 , Ruhui Song 2 , Hyunseok Lee 2 , Yun-hye Kim 2 , Dae-seung Baek 2 , Chul Park 2 , Jinho Park 2 . 1 Western Animal Medical Center, Seoul, Korea, 2 College of Veterinary Medicine, Chonbuk National University, Jeonju, Jeonbuk, Korea.

Naturally occurring CD4 + , CD25 + regulatory T cell (Treg) that is produced by the normal thymus as a functionally mature T-cell subpopulation has a function that maintain immunologic self-tolerance and inhibit immune response. In septic patients, however, the percentage of circulating Tregs is markedly increased, which may cause sepsis-induced immunosuppression. In a report, sepsis surviving-mice that express persistent expansion of Tregswas correlated with a long-term adaptive immune dysfunction and susceptibility to secondary infection.We performed this study to investigate serial changes of regulatory T cells in canine septic model. 4 beagles in experimental group were injected with high dose LPS (1 mg/kg) diluted in 5 ml saline for canine septic model. 4 beagles in control group were injected with saline of the same dose for experimental group.CD4 (+)/CD25 (+)/FoxP3 (+) lymphocytes were regarded as canine Tregs using flow cytometry. Changes of regulatory T cells in peripheral blood were investigated for 7 days.

The percentage of CD4 (+)/CD25 (+)/FoxP3 (+) T cells in total lymphocytes decreased in 1 day after LPS injection (0.04% AE 0.04%, p < 0.05). And after that, it increased in gradually at 3 and 7 days after LPS injection compared with control group (respectively 0.61% AE 0.18%, p < 0.01 and 1.39% AE 0.49%, p < 0.05). Figure 1 . Changes of percentage of CD4 (+)/CD25 (+)/FoxP3 (+) T cells in total lymphocytes during 7 days of LPS injection. The percentage was measured by flowcytoemtric analysis. The percentage of CD4 (+)/CD25 (+)/FoxP3 (+) T cells was decreased in 1 day after LPS injection, and increased in rapidly after 1 day. Data represent meanAE S.D *p < 0.05, **p < 0.01 versus baseline. Cytokines, adipokines and hormones are the principal mediators orchestrating physiological response to stimuli/stress and are therefore 'signatures' of health or disease. Evaluating static as well as temporal changes in these molecules provides a comprehensive understanding of the status of a biological system and its response to stressors and interventions [nutritional & therapeutic] . Here we report the development of a set of novel multiplex assay capable of simultaneously measuring multiple biologically relevant feline proteins. These panels are assayed on a Luminex xMAP platform and will enable simultaneous quantitative measurement of~26 different feline proteins consisting of several cytokines, adipokines and hormones, using very small quantities of serum/plasma samples. In conjunction with appropriate pattern recognition and pathway analysis techniques these panels will help predict / evaluate functional outcomes of physiological stressors and interventions.

. P Naranatt 1 , S Moroff 1 , J Millet 2 , B Licitra 2 , G Whittaker 2 . 1 Antech Diagnostics, Irvine, CA, 2 Cornell University, Ithaca, NY.

Feline coronaviruses (FCoVs) occur as two pathotypes, feline infectious peritonitis virus (FIPV) and feline enteric coronavirus (FECV). FECV is common in cats, causing mild transient enteritis in kittens, but is usually asymptomatic in adult cats. In contrast, FIPV is sporadic but invariably lethal. The purpose of the present study was to evaluate the performance of RT-PCR based on spike gene mutations to differentiate FIPV from FECV.

Identification of genetic differences conferring pathogenicity of feline coronaviruses has been elusive. Studies have indicated sequence differences in spike protein (S gene), membrane protein, envelope protein and NSP 3c and 7b. Bioinformatics studies from Whittaker laboratory identified presence of 2 proteolytic cleavage sites in the Spike protein of FCoV type I strains. These include a furin cleavage site motif (R-R-S/A-R-R-S) at S1/S2 and a second cleavage motif (K-R-S) at S2'. Studies indicated that S1/S2 consensus was mutated in all cases of cats with confirmed FIP. Similarly, in some FIP cases, the S2′ site was mutated to change the dibasic motif to a monobasic amino acid. For type II FCoV strains, cleavage occurs at the Y-R-S sequence, found at the S2′ site.

We developed RT-PCR assays that amplify genetic mutations encoding cleavage motif(s) of type I and type II viruses, to detect FIP and differentiate FECV from FIPV. SYBR green PCR identified positive samples by dissociation and melting temperatures. Amplicons were sequenced by dye-terminator chemistry. The sequences with good Phred scores were aligned using Codon Code Sequence Aligner. Contiguous sequences were translated to identify the motif(s) and to differentiate FECV and FIPV. A total of 89 clinical samples were tested using PCR: 35 ascites, 40 blood samples, 11 formalin fixed paraffin embedded tissues (FFPE) tissues, and 3 thoracic effusions. Diagnosis of FIP was achieved by necropsy or surgical biopsy and, in most cases, demonstration of intralesional coronavirus positive cells by immunohistochemistry (IHC) or combination of cytologic diagnosis, ascitic electrophoresis and clinical findings compatible with FIP. In 2 cases, paired blood and ascites were positive. Test performance of FIP assay compared to the above FIP diagnostic criteria was computed to be: sensitivity (86.36%), specificity (98.53%), PPV (95%) and accuracy (95.56%). 20 of 23 samples were identified to have mutations in either S1/S2 or S2' site(s) and were categorized as FIPV. Three samples did not reveal any mutations and were called FECV. 38 blood samples (negative for antibodies to FCoV), 26 cavitary effusions (from cats with diseases other than FIP), and 3 FFPE tissues (negative for FCoV by IHC) were tested by PCR and all were found to be negative.

Thus, PCR amplification of spike gene mutations encoding S1/ S2 and/or S2' cleavage sites in blood, cavitary effusions, CSF and formalin fixed paraffin embedded tissues differentiates FIPV from FECV. For many reasons, the diagnosis of canine vector-borne infectious diseases (CVBDs) can be challenging for clinicians. CVBDs are caused by a diverse array of pathogens with varying biological behaviors. CVBDs result in a wide spectrum of clinical presentations and laboratory abnormalities. For these reasons, clinicians continue to search for the "holy grail" of CVBD diagnostic testing. There is an ongoing debate over the relative value of serological and molecular assays. The purpose of this study was to compare serological and PCR CVBD testing results in healthy dogs and in sick dogs suspected to have CVBDs.

We prospectively tested healthy dogs (n = 30) using a panel of serological and molecular assays for evidence of exposure to, or infection with, 10 genera of organisms that cause CVBD. Using the same panel, we retrospectively tested 3 additional groups of sick dogs: 1) dogs that were seronegative and only had serologic testing requested by the attending clinician (n = 25), 2) dogs that were seroreactive against at least one organism and only had serologic testing requested by the attending clinician (n = 25), and 3) dogs that were PCR positive for at least one organism and only had PCR testing requested by the attending clinician (n = 24).

In these representative populations a panel combining serological and molecular assays run in parallel would have resulted in 4-60% increased recognition of exposure to, or infection with, CVBD. We conclude that the "holy grail" remains elusive and serological and PCR assays should be used in parallel to maximize CVBD diagnosis. Bartonella clarridgeiae has frequently been grown or its DNA amplified from Ctenocephalides felis collected from naturallyexposed cats. However, transmission of this agent among cats exposed to C. felis, using an experimental model, has not been reported. The purpose of this study was to co-house cats experimentally infected with B. clarridgeiae with na€ ıve cats while maintaining the cats in either a flea-free environment or with an infestation of laboratory-reared C. felis to determine whether this flea could be a vector.

Specific pathogen free cats (n = 18) were purchased from a commercial vender and managed under an approved institutional animal care and use protocol. A field strain of Bartonella clarridgeiae was used to infect one kitten, which became strongly PCR positive by Day 19 after IV inoculation. This cat was then gang housed with nine other B. clarridgeiae na€ ıve kittens in the absence of C. felis for > 150 days. Blood from the primarily inoculated kitten was used to inoculate two additional kittens that were then gang housed with six other B. clarridgeiae na€ ıve kittens. Once the two kittens were confirmed to be PCR positive, 50 male and 50 female laboratory-reared C. felis were placed on each of the eight cats every 14 days for five infestations. Blood was collected intermittently from all cats and assessed for B. clarridgeiae DNA using a previously reported PCR assay.

While B. clarridgeiae DNA was amplified from the primarily inoculated kitten consistently, none of the nine na€ ıve kittens that were gang housed with the kitten in the absence of C. felis were ever PCR positive. In contrast, five of the six kittens living with the two experimentally inoculated kittens that were infested with C. felis became persistently B. clarridgeiae DNA positive. The B. clarridgeiae infection rates between groups was significantly different (P = 0.002). There was no clinical evidence for bite or scratch wounds in any of the kittens over the duration of the study.

The results support the hypothesis that C. felis can be a competent vector for B. clarridgeiae. As this agent is associated with disease in some people, dogs, and cats, the results support the recommendation that flea control products be administered to all pets. False-negative fecal parvovirus antigen results in dogs with parvovirosis are common, but circumstances leading to these false-negative results are still unknown.

The aim of this study was to investigate whether clinical or laboratory variables influence the results of fecal parvovirus antigen ELISA and whether dogs with false-negative results show less severe symptoms of parvovirosis or faster recovery than dogs with positive results.

Eighty dogs with parvovirosis, confirmed by presence of clinical signs and a positive fecal parvovirus PCR, were assigned to two groups according to their fecal ELISA result. Severity of symptoms, evaluated by a clinical score, and blood parameters (CBC, albumin, total protein) were compared between both groups. Forty of these dogs were treated with a standardized treatment protocol. Clinical scores, laboratory changes, and time of recovery were compared between dogs with positive and negative fecal ELISA.

Thirty eight of 80 dogs showed a positive fecal ELISA result, whereas 42/80 had a false-negative result. Dogs with positive ELISA showed significantly lower WBC counts than dogs with negative ELISA. Dogs with positive ELISA had a significantly higher clinical score and a higher frequency of defecation than dogs with negative ELISA. However, when comparing ELISAnegative and ELISA-positive dogs receiving standardized treatment, no significant difference in outcome was detectable.

False-negative fecal antigen ELISA results in dogs with parvovirosis are very common. Dogs with parvovirosis showing falsenegative antigen ELISA results seem to suffer from less severe clinical and laboratory changes. However, clinical outcome is not associated with the fecal ELISA result. A 13 year old, entire female, DSH cat was presented at the Small Animal Clinic, Faculty of Veterinary Medicine, Assiut University for investigation of a persistent skin lesion that was described by the owner as an ulcer.

After physical examination, a blood sample was collected for hematological and biochemical investigations. Sterile bacteriological swap was obtained aseptically from the lesion for bacteriological examination. The tumour was surgically removed and submitted for histopathology.

On physical examination, a mammary gland ulcerated mass over three centimetres in diameter, in left lower quarter was evident with purulent discharge from the teat canal. Physical exami-nation was otherwise unremarkable. CBC revealed monocytosis with lymphocytopenia and a severe degenerative left shift. Macrocytic hypochromic anaemia was evident. Sero-biochemical analysis revealed a marked elevation of blood urea nitrogen and creatinine. Histopathology confirmed the mass as a typical mammary adenocarcinoma. Bacteriological examination revealed suspected mycobacterial colonies on LJ media. Suspected colonies were confirmed by PCR amplification and sequence analysis of the inter transcriped spacer (ITS) target as Mycobacterium abscesses.

This is the first report of a spontaneous mammary gland ulcer secondary to M. abscesses infection in a cat with mammary adenocarcinoma. Immune status is an important factor determining the course and prognosis of opportunistic infections by rapid growing non-tuberculous Mycobacterium. The fast deterioration of this cat was not un-expected considering the size of tumour on first presentation. CBC and the persistent M. abscesses infected ulcer could reflect the immunocompromised state of the cat. However, further observations and studies are necessary to understand the potential relationship between the isolated microorganism and the development of the tumour. Canine parvovirus is a highly infectious, often fatal disease that affects young unvaccinated dogs. Lack of proper preventative care, a contributor to the disease severity, frequently occurs within lower socio-economic classes. Owners who do not provide preventative measures are typically unable to afford the recommended inpatient treatment. Establishment of a successful outpatient treatment protocol would provide an alternative option for dogs that may otherwise die or be humanely euthanized. We hypothesized that dogs with naturally occurring parvoviral gastroenteritis may be successfully managed using an outpatient treatment protocol, as determined by a similar clinical response to treatment and overall outcome, when compared to those managed with an inpatient (hospitalized) treatment protocol.

Forty naturally infected dogs with no vaccination history or prior treatment intervention were enrolled following a positive snap ELISA parvoviral test and owner consent. Upon enrollment, all dogs received intravenous fluid resuscitation to stabilize cardiovascular parameters. Dogs were subsequently randomized to receive either an inpatient (n = 20) or outpatient (n-= 20) treatment protocol. Those within the inpatient group received intravenous fluids, enteral nutrition, maropitant (1 mg/kg IV q24 h), and cefoxitin (22 mg/kg IV q8 h). Those within the outpatient group received subcutaneous fluids, enteral nutrition, maropitant (1 mg/kg SC q24 h), and cefovicin (8 mg/kg SQ once). Parameters monitored until hospital discharge included clinical severity scoring, body weight, complete blood count and electrolyte panel, frequency of vomiting, caloric intake, hydration status, visceral pain and nausea scoring, and length of hospitalization.

Overall outcome and intermediate health measures for dogs in the two treatment groups were similar. There was no difference between groups with regard to age or duration of clinical signs prior to presentation. Additionally, no difference in survival rate was observed (18/20 inpatients vs. 16/20 outpatients, p = 0.66) when comparing the two protocols. Duration of hospitalization, blood work parameters, changes in body weight, hydration status throughout hospitalization, caloric intake, or serial objective scoring of clinical severity, nausea and visceral pain were not different between treatment groups.

This study suggests that a modified outpatient protocol may be a reasonable alternative for dogs whose owners cannot afford hospitalization for treatment of canine parvoviral gastroenteritis. Such a protocol should be considered under the appropriate circumstances, but should not be considered a substitute for inpatient care when such care can be provided. Little information is available regarding use of hand hygiene and other critical basic infection control measures in veterinary clinics. The objectives of this study were to describe the use of hand hygiene, personal protective clothing (PPC) and environmental cleaning associated with routine companion animal appointments.

Observations were made in 51 clinics over 3 weeks using 2 wireless surveillance cameras in each clinic: one in an exam room, and one in a nearby backroom area. Data from 33 clinics are presented, including 874 appointments involving 1634 individual animal contacts. The percentage of cases in which hand hygiene was observed was: in the room prior to animal contact 3% (58/ 1767), in the room after animal contact 7% (127/1780), outside the room after animal contact 15% (263/1780). Of the hand hygiene attempts observed, available alcohol-based hand sanitizer (AHS) was used in 14% (42/302). Median contact time with soap was 2s (range 1-23s, n = 415); median contact time with AHS was 9s (range 1-30s, n = 37). Appropriate PPC was worn for 67% (1087/1634) of contacts. Tables were cleaned and floors were mopped after 85% (496/585) and 8% (52/641) of appointments in which the animal had contact with these surfaces, respectively. The median contact time for table disinfectant was 9s (mean 32s, range 0-1843s).

Overall hand hygiene compliance in clinics was low, and contact time with hand hygiene agents was typically below recommendations (15-20s). Use of PPC and contact time with environmental disinfectants could also be improved. Better use of these simple measures is important for infection control in veterinary clinics. Acute canine granulocytic anaplasmosis is characterized by fever, thrombocytopenia, lethargy, anorexia and joint problems, skin lesions are rare in dogs. Persistent infection in dogs is characterized by recurrent bacteremia, positive antibody titers and PCR, clinical signs, however, have not been observed.

The objective of this study was to describe the histopathological changes observed in canine skin biopsies from lesional, PCR positive skin of dogs seropositive against A. phagocytophilum and responding to treatment with doxycycline.

Paraffin-embedded lesional skin biopsies from dogs seropositive against A. phagocytophilum and responsive to treatment with doxycycline were evaluated (n = 16). Histopathology of the skin lesions did not indicate an apparent cause. DNA was extracted and a conventional PCR for partial 16s rRNA gene was performed.

In skin biopsies from 4 dogs A. phagocytophilum DNA was amplified and sequences were identical to the prototype A. phagocytophilum. A short summary of the PCR-positive cases is presented in the following We conclude that persistent A. phagocytophilum infection is characterized by a variable degree of cellular infiltrate and edema in the superficial dermis, and it should be included in the differential diagnoses as cause for skin lesions in seropositive dogs responding to doxycycline treatment.

All 218 serum samples were seronegative for E. ewingii, E. chaffeensis and B. burgdorferi. Two samples were seropositive for A. phagocytophilum, and both of these were PCR negative. Exposure to tick-borne pathogens was highest in shelter animals and military working dogs where >90% of the samples were seropositive or PCR positive for one or more organisms as compared to 51% in client owned animals. These data indicate exposure to tick-borne pathogens in Barranquilla, Colombia is extremely high with significant levels of bacteremia. This is most critical in dogs in kennel environments such as shelters or military working dog kennels. Both E. canis and A. platys are primarily associated with clinical disease in dogs. However, E. canis infection and infection with an "A. platys-like" organism has been documented in people in South America and this high level of exposure warrants precautionary measures for both dogs and people. Q fever is a zoonotic disease caused by the pathogen Coxiella burnetii, an obligate intracellular bacterium that is found worldwide. Symptomatic disease in humans is most often associated with livestock exposure, however, it has been reported from contact with parturient cats. The organism is associated with reproductive abnormalities in a number of species including cats. In a previous study, our laboratory amplified C. burnetii DNA from the uterine tissues of 8.5% of client-owned cats, but did not attempt to associate it with clinical findings. The purpose of this study was to determine if cats with reproductive abnormalities were more likely to have C. burnetii DNA amplified from tissues than normal cats.

Total DNA extracts from the uterine tissues from 26 normal cats and eight cats with histopathological evidence of uterine disease or other reproductive abnormalities were shown to be negative for T. foetus DNA in a previous study of cats housed in catteries. The total DNA extracts from the uterine tissues were stored at -80°C until shipped on dry ice to Colorado State University. A PCR assay that amplifies the repetitive transposon-like region (Trans 1 and 2) and a PCR assay that amplifies the IS-1111-insertion sequence (IS-1111) were optimized and applied to the DNA extracts. The sensitivity threshold of both PCR assays was 12 pg/ll.

Amplicons of the expected size developed in three samples in the IS-1111 assay, however, there was not enough DNA for sequence analysis. Statistical associations between positive C. burnetii assay results and reproductive abnormalities were not detected.

While the amplicons could not be confirmed as C. burnetii by sequence analysis, the PCR positive prevalence rate is similar to what has been published previously. As the overall sample size was small, these data suggest a larger study should be performed to further evaluate the role C. burnetii may play in reproductive abnormalities in cats. These interim results suggest that urine sediment examination may be useful for rapid, cost-effective diagnosis of B. dermatitidis infection in some dogs. Sample size precludes statistical analysis at this time, but it appears that identification of organisms in urine sediment is more likely in intact dogs. Vector-borne pathogens, which infect dogs throughout North America, cause a spectrum of clinical and hematological disease manifestations. As tick distributions expand through changes in ecosystem management, wildlife migration and increased travel by humans and their companion animals, regional shifts in tick-borne pathogen exposure are likely. In an effort to better delineate regional exposure to various tickborne pathogens, an expanded set of species-specific peptides were used to develop an experimental rapid, in-house diagnostic SNAP â Multi-Analyte Assay (SNAP â M-A). The assay detects specific antibodies to Ehrlichia canis (Ec), E.chaffeensis (Ech), E.ewingii (Eew), Anaplasma phagocytophilum (Aph), A.platys (Apl) and Borrelia burgdorferi (Bb) in canine serum samples. Archived serum from a population of dogs being screened for tick-borne diseases by the Vector-Borne Disease Diagnostic Laboratory from 2008 to 2012, were rescreened with SNAP â M-A to determine the species-specific seroprevalence for six tick-borne pathogens. Of the 5889 samples evaluated, 5627 (95.5%) were submitted from the USA, with Southern and Midwestern regions overrepresented (47% and 17.4%, respectively), followed by the MidAtlantic (16%), Northeast (8%), and West (7.3%). In addition, 262 samples (4.4%) were submitted from Canada. Below, seroprevalence (%) is shown for overall population and by region. Overall, the most common co-exposures were Eew + Ech (1.4%) followed by Aph + Bb (1.2%). In the MidAtlantic, Eew + Bb (1.8%) and Ech + Bb (1.6%) co-exposures were comparable to Aph + Bb (2.0%). Improved documentation of exposure to multiple tick-borne pathogens should facilitate more accurate epidemiological surveillance and can enhance the diagnostic identification of regionally neglected species and coinfections. Hand hygiene is the most important infection control measure in hospital environments. An anonymous, cross-sectional survey was designed to better understand veterinarians' hand hygiene and glove use habits and to ascertain associated factors.

The survey, offered online from June-December 2011 through SurveyMonkey and the Veterinary Information Network, contained demographic questions and questions regarding glove use and hand hygiene practices. Data were grouped by gender, years in practice, or written infection control plan (wICP) status, then analyzed using Pearson's chi-square, Kruskal-Wallis, and Jonckheere-Terpstra tests.

Eight hundred nineteen practicing veterinarians responded. Depending on patient's clinical status, 87-100% of veterinarians stated that they routinely cleaned their hands for patient examinations, 84-97% for glove use situations, 76-100% for contact with contaminated items, and 96-100% for personal hygiene. Following glove removal, 60% claimed to always wash their hands. Reported glove use was 15%-95%, with the highest rates for examining patients with known/suspected zoonotic infections, necropsies or obstetrical procedures. Gender was more often associated with hand cleaning than were years in practice or practice wICP status. Years in practice and practice wICP status were more strongly associated with glove use than was gender.

Although veterinarians report appropriate hand hygiene and glove use practices when perceived risk is high, they report a lower prevalence of these practices in other scenarios that can be associated with disease transmission. Factors such as gender, years in practice and practice wICP status are associated with hand hygiene or glove use practices, and should be considered in programs to improve compliance. Alpha-1-acid glycoprotein (AGP) is an acute-phase serum protein that is produced by the liver in response to inflammation, infection and neoplastic conditions. Feline infectious peritonitis (FIP) is one of the most important viral diseases of cats. The diagnosis of FIP is often challenging. The clinical suspicion of FIP must be supported by the analysis of effusions when present, but usually histopathological analysis is the definitive test. Nowadays, reverse transcriptase polymerase chain reaction (RT-PCR) has been used to detect FCoV and is fast and sensitive, but results must be interpreted in the context of clinical-pathological findings. The aim of this study was to determine AGP concentration in cats with clinical and laboratorial signs of FIPeffusive and non-effusive forms. Total RNA from blood and effusion samples was tested with a RT-PCR for detection of FCoV mRNA of the highly conserved M gene. Healthy cats were used as control group. AGP was measured by the use of a commercial species-specific immunoassay test (PHASE Feline alpha-1 acid glycoprotein SRID Assay Kit, Tridelta Development Ltd, Ireland) in two groups of cats: 20 healthy cats and 8 cats with FIP diagnosed by RT-PCR. For both groups, serum samples were collected, harvested and frozen at -80°C until assayed. Unpaired T test was used to compare the groups. Mean of AGP concentration was significantly higher (3579.5 AE 690.27 lg/mL) in cats with FIP at diagnosis than in healthy cats (235.22 AE 154.19 lg/mL), p < 0.0001. The results agree with other author, who studied this same protein in cats with FIP and other diseases and found that, when the pretest probability of FIP is high, based on history and clinical signs, moderate serum AGP levels (1500 to 2000ug/mL) can discriminate cats with FIP from others, while only high AGP levels (>3000ug/mL) can support a diagnosis of FIP in cats with low pretest probability of disease. In conclusion, AGP concentration was significantly higher in cats with FIP and, based on history, clinical signs and laboratorial alterations can help with FIP diagnosis. Currently available 4 serovar containing Leptospira vaccines induce detectable microscopic agglutination test (MAT) results in most dogs shortly after vaccination. However, little is known concerning the duration of positive Leptospira MAT titers in client-owned dogs and what anamnestic titers develop after booster vaccination. The objectives of this study were to determine Leptospira MAT titers of previously vaccinated dogs one year after the primary immunization series and one month after a booster vaccine.

Healthy, client-owned dogs (n = 23) were given two doses of one of four vaccines containing the Canicola, Grippotyphosa, Icterohemorrhagiae, and Pomona serovars approximately 52 weeks previously. On recheck examination, serum was collected and the same vaccine previously used was given as a booster. The one year sera as well as sera collected approximately four weeks after booster vaccination (week 56) were tested for antibodies against serovars Bratislava (B), Canicola (C), Grippotyphosa (G), Hardjo (H), Icterohemorrhagiae (I), and Pomona (P) using MAT.

Percentages of dogs developing a MAT titer of ! 1:100 varied between the weeks and amongst the vaccines (Table 1) . Vaccine Overall, 87% (20/23) dogs were negative for antibodies against all serovars approximately one year after previous vaccination (Week 52). MAT titers ! 1:800 were detected for at least one serovar for 11/23 dogs (47.8%) approximately 4 weeks following a one-year booster vaccination.

As previous studies have demonstrated that commercially available vaccinations provide at least 1-year duration of immunity, it is clear that susceptibility to Leptospire cannot be determined from serology. MAT titers ! 1:800 will not always equate to the presence of clinical leptospirosis in the event of recent vaccination. Administration of an intranasal (IN) modified live FVRCP vaccine was shown to induce significant protection against FVH-1 challenge by day four after vaccination in a previous study but the relative efficacy was less than 100%. While the earliest onset of FHV-1 immunity induced by administration of modified live FHV-1 vaccines subcutaneously (SQ) is unknown, serological responses are generally low until after a booster vaccine and relative efficacy after challenge is less than 100%. The objective of this study was to compare clinical signs and FHV-1 shedding rates between a group of kittens vaccinated with both an IN and SQ vaccine and a group of kittens vaccinated SQ alone after FHV-1 challenge inoculation Twenty-four specific pathogen free 8 week-old kittens were randomized into 3 groups with 8 kittens. One group was administered a single dose of SQ vaccine (Felocell 3; Pfizer Animal Health, New York, NY) and the other group was administered a single dose of an IN vaccine (Felocell FVR C; Pfizer Animal Health) in addition to the SQ vaccine. The third group was maintained as an unvaccinated control group. Seven days later, all 24 kittens were inoculated with the USDA challenge strain of FHV-1 intranasally. Clinical scores were collected daily starting on the day of vaccination and ending 14 days after FHV-1 inoculation. Pharyngeal swabs were collected twice weekly for evaluation in a quantitative FHV-1 PCR assay.

In the first seven days after FHV-1 vaccination, the kittens that were administered both the SQ and IN vaccines were more likely to exhibit mild clinical signs, mostly sneezing, than control cats or cats administered the SQ vaccine alone (p < 0.05). During the 14 days after FHV-1 inoculation, kittens administered both the SQ and IN vaccines had significantly less clinical signs of disease than kittens administered the SQ vaccine alone (p < 0.05). Significantly less FHV-1 DNA was shed by the kittens administered both the SQ and IN vaccines when compared to the control group and the kittens administered the SQ vaccine alone (p < 0.001).

While administration of the FHV-1 containing vaccine can lead to transient sneezing after primary vaccination in na€ ıve cats, the combination of IN and SQ vaccination significantly lessened clinical signs of disease and viral shedding after FHV-1 challenge. These results indicate that concurrent administration of both the IN and SQ vaccines was superior to administration of the SQ vaccine alone in this model. Hand hygiene is a critical, basic infection control measure, yet there is little information available regarding perceptions and use of hand hygiene in veterinary clinics. The objectives of this study were to describe the perceived importance of hand hygiene at various times and as an overall infection control measure among companion animal veterinary clinic staff, as well as potential barriers to hand hygiene compliance.

An anonymous, voluntary written questionnaire was provided clinic staff members following participation in a video-observation study evaluating hand hygiene behaviour. Surveys were completed by 356 of approximately 558 individuals (64%) from 49/51 clinics. On a scale of 1 (not important) to 7 (very important), the percentage of respondents who rated hand hygiene as 6 or 7 were: before handling any animal 75% (268/356), after handling a relatively healthy animal 88% (312/356), after glove removal 64% (229/ 356), after handling an animal with suspected gastrointestinal, respiratory or dermatological disease 99% (352/356), after contact with urine/feces 99% (354/356), before eating/drinking/smoking 98% (349/356), overall importance for preventing spread of disease in veterinary clinics 96% (340/355). The most frequently reported reason for not performing hand hygiene was forgetting to do so (40%, 141/353). Specific discussion of hand hygiene practices at work was recalled by 114/354 (32%) respondents.

Education of veterinary personnel regarding the importance of hand hygiene prior to animal contact and particularly after glove use should be considered. Hand hygiene should be emphasized more in staff training, in order to help make the practice more habitual and improve compliance. Coccidiosis is a common protozoal intestinal infection of dogs and cats and in most cases of adult infection, clinical signs are absent to mild/moderate. However, in shelters, the risk of infection increases and the control of coccidiosis is an important shelter management issue. Recent reports of drug resistance and persistent infections despite appropriate treatment with sulfadimethoxine have resulted in the increasing use of ponazuril offlabel at a variety of dose rates. The aim of this study was to assess the efficacy of ponazuril for the treatment of canine and feline coccidiosis in a prospective randomized clinical trial using 3 dose rates.

Feces were collected from animals entering an a large metropolitan adoption-guarantee shelter and coccidiosis was initially dignosed by fecal flotation (FFl), centrifugation and microscopic examination. Coccidiosis was confirmed at a diagnostic parasitology laboratory, where fecal oocyst count was performed (oocysts/g feces). On enrollment, infected animals were allocated to receive ponazuril orally at 1 of the following 3 dose rates using a computer-generated randomization scheme -50 mg/kg q24 h for 3 days (DR1), 50 mg/kg as a single dose (DR2) or 20 mg/kg as a single dose (DR3). Animals were bathed once on enrollment to remove oocysts from the hair coat. Fecal examinations were repeated at Day 3-4 and at Day 8 using the same protocol as at enrollment. Animals still infected at Day 3-4 were treated with DR1 if possible.

Fifty-four cats (approximate age range 8 weeks to 5 years; genders F n = 26, FS n = 4, M n = 21 MN n = 3) and 33 dogs (approximate age range 8 weeks to 4 years; genders F n = 16, FS n = 2, M n = 14 MN n = 1) were enrolled as follows -DR1 n = 7 cats and n = 5 dogs; DR2 n = 24 cats and n = 12 dogs; DR3 n = 23 cats and n = 16 dogs (allocation to DR1 was discontinued early and animals were randomly allocated only to DR2 and DR3 for the remainder of the trial). In the DR1 group, 2/7 cats and 1/5 dogs were FFl positive for coccidia at Day 3-4 but all were FFl negative at Day 8 after repeat treatment with DR1 at Day 3-4. In the DR2 group, 3/24 cats and 1/12 dogs were FFl positive for coccidia at Day 3-4. Two of those 3 cats and both dogs were FFl negative at Day 8 after also receiving DR1 from Day 3-4; the remaining cat received no further ponazuril but was FFl negative at Day 8. A further 2 cats and 1 dog in DR2 that were FFl negative at Day 3-4 were FFl positive at Day 8. In the DR3 group, 10/23 cats and 5/16 dogs were FFl positive for coccidia at Day 3-4; 6/10 cats and 3/5 dogs received further treatment with DR1 from Day 3-4; 3/6 cats and all 3 of the dogs were FFl negative at Day 8; the remaining 3/6 cats were FFl positive at Day 8. In the DR3 group, 4 cats and 2 dogs that were FFl positive at Day 3-4 did not receive any further ponazuril; 2/4 cats and both dogs remained FFl positive at Day 8; the remaining 2 cats were FFl negative at Day 8. A further 2 cats in DR3 were FFl negative at Day 3-4 and FFl positive at Day 8.

We conclude that extended courses of ponazuril at 50 mg/kg q24 h PO are sometimes necessary for fecal flotation results to become negative in dogs and cats with coccidiosis. Chronic kidney disease is a common cause of morbidity and mortality in cats that is often due to unknown causes. Bartonella henselae DNA has been amplified from renal tissues from experimentally inoculated cats with concurrent mild lymphocytic interstitial nephritis. The purpose of this study was to determine if there are associations between Bartonella species antibody assay results and azotemia in client-owned cats in the United States.

Sera from 71 cats with creatinine concentrations > 2 g/dl (azotemia) and 90 cats with creatinine concentrations 2 g/dl enrolled in a previous study were tested for Bartonella species IgG using a previously reported enzyme-linked immunosorbent assay with a positive cutoff value of ! 1:64. The state of origin was used to classify each case as having low flea risk (AK, AZ, CO, ID, MT, NM, NV, UT, WY) or high flea risk (all other states). Associations between proportions of positive Bartonella spp. IgG assay results and azotemia were evaluated using logistic regression with P > 0.05 considered significant. The initial logistic model controlled for age and flea risk but age was removed from the model as it was not significant.

Overall, cats from high flea risk states were more likely to be positive for Bartonella spp. IgG than cats from low flea risk states. Prevalence rates for Bartonella species IgG in cats with azotemia (23.5%, SE = 5.4%) or without azotemia (33.7%, SE = 5.8%) were not significantly different (p = 0.14) and there were no differences in titer magnitude between groups.

Bartonella spp. IgG antibodies were not associated with azotemia in this sample set. However, Bartonella spp. IgG antibodies do not indicate current infection and high prevalence rates can mask statistical associations. Future studies should also include culture or polymerase chain reaction on blood or renal tissues to further evaluate for associations between current Bartonella spp. infection and feline chronic kidney disease. Leptospira spp. DNA was amplified from the urine of 10 of 85 cats housed in Colorado animal shelters in a previous study in our Center. However, the Leptospira spp. 16S ribosomal DNA real time PCR assay used does not provide information concerning which Leptospira spp. was being shed. The purpose of this study was to clone and sequence positive samples obtained in the previous study to gather further information on which Leptospira spp. infect cats in the United States.

Appropriately sized DNA PCR products from the 10 positive samples were excised from an agarose gel and purified (Qiagen Gel Purification Kit, Qiagen, Valencia, CA). The DNA fragment was cloned into commercially available plasmid DNA (Clone-Smart Cloning Kit, Lucigen, Middleton, WI). Insert DNA was sequenced with plasmid primers flanking the insert sequence (SL1 Primer, SR2 Primer, Lucigen, Middleton, WI) at a core facility (Proteomics and Metabolomics Facility, Colorado State University). DNA sequences were aligned and analyzed using CLC Sequence Viewer 5 (CLCbio, Aarhus, Denmark) and identified by BLAST search (NCBI) in the nr database.

Sequencing was completed successfully for one of the positive samples. Alignment with GeneBank submissions showed this isolate to be most homologous with Leptospira interrogans serovar Hardjo-prajitno (99%).

Previous studies have implicated canicola, grippotyphosa, and pomona as possible infecting serovars in cats, but correctly identifying the serovars and serogroups remains a diagnostic challenge. Leptospira interrogans serovar Hardjo-prajitno primarily infects bovine species. As this cat was housed in a shelter, it is possible contact with bovine urine could have been made. Although clinical disease in cats is rarely reported, understanding which serovars and serogroups infect cats can have significant impact on understanding how cats play a role as possible reservoir or incidental hosts in the transmission of leptospirosis. Canine parvovirus (CPV) and canine distemper virus (CDV) are frequent causes of morbidity and mortality for dogs in animal shelters. Many dogs admitted to shelters do not have protective immunity against CPV or CDV. Current guidelines recommend vaccination of all dogs against CDV and CPV on shelter admission. The objective of this study was to determine the proportion of dogs responding to vaccination and the time to develop protective antibody titers (PAT) to CDV and CPV.

Blood samples collected from 845 dogs at shelter admission were tested for CDV and CPV PAT. Dogs without PAT received a vaccine containing canarypox-vectored recombinant CDV and modified-live CPV. Blood samples were collected on days 7, 14, 21, and 28 for PAT testing. Proportions of juveniles (<6 months) and adults that seroconverted at each time point were compared using Chi-square or Fisher exact tests.

At admission, 34% of adults and 84% of juveniles did not have CDV and/or CPV PAT. The proportion of adults that responded to vaccination was greater than for juveniles on days 14, 21 and 28. Development of PAT to CPV was faster than for CDV in both age groups. By day 14, 47% of adults had CDV PAT and 91% had CPV PAT, while only 21% of juveniles had CDV PAT and 54% had CPV PAT.

This study suggests that many dogs, particularly juveniles, enter shelters without immunity to CDV and/or CPV. The slow response to vaccination creates a large pool of unprotected dogs that contribute to risk for disease outbreaks. There are few reports of serum electrolyte abnormalities in canine babesiosis and none specifically in dogs with complicated babesiosis. One study showed a trend towards hypokalemia, hyperchloremia, and mild hyponatremia, whereas another study showed a trend towards hyponatremia, hypokalemia, and hypochloremia. Complicated babesiosis has a high mortality rate and is characterized by 1 or more organ dysfunctions.

The purpose of this study was to determine serum sodium, potassium, chloride, total calcium, inorganic phosphate, and albumin in 60 dogs with complicated Babesia infection. Serum albumin was measured in order to calculate the corrected calcium. To account for water balance changes the corrected chloride was calculated using the following formula: corrected chloride = measured chloride x 146/measured sodium. The complications that occurred were haemoconcentration, cerebral, acute respiratory disease syndrome, acute kidney injury, and cardiac.

Results showed that the median serum sodium was 145 mmol/ l (range 136-158), with 52 cases being within normal reference range, 6 cases of hyponatremia, and 2 cases of hypernatremia. The median serum potassium was 3.95 mmol/l (range 2.4-7.4), with 30 cases being within normal reference range, 24 cases of hypokalemia, and 6 cases of hyperkalemia. The median chloride was 123 mmol/l (range 105-141), with 11 cases being within normal reference range, 8 cases of hypochloremia, and 41 cases of hyperchloremia. The median corrected chloride was 123.5 mmol/l (range 106-142), with 11 cases being within normal reference range, 5 cases of hypochloremia, and 44 cases of hyperchloremia. The median total calcium was 2.35 mmol/l (range 1.75-3.63), with 51 cases being within normal reference range, 4 cases of hypocalcemia, and 5 cases of hypercalcemia. The median inorganic phosphate was 2.2 mmol/l (range 0.36-7.45), with 3 cases being within normal reference range, 15 cases of hypophosphatemia, and 42 cases of hyperphosphatemia. There was no correlation with the presence of hyper-and hyponatremia with serum potassium and creatinine; and no correlation with the presence of hyperkalemia and hypercalcemia with serum creatinine.

This study showed that common electrolyte abnormalities in complicated canine babesiosis were hyperphosphatemia, occurring in 70%; hyperchloremia, occurring in 68%; and hypokalemia, occurring in 40% of cases. Likely etiologies for the hyperchloremia would be metabolic alkalosis or renal tubular acidosis; the hyperphosphatemia haemolysis or reduced renal clearance; and the hypokalemia reduced dietary intake, alkalosis, or renal tubular acidosis. 2%) and 107 (5.8%) cats tested positive for FeLV and FIV, respectively. FeLVpositive results among male cats ranged from 0.7% to 2.5%. For females, FeLV-positive results were significantly fewer in 2006 (p < 0.01). FIV-positive results ranged from 9.5 to 13.1% for males and 1.1% to 2.7% for females (p < 0.01 for each year). Male cats were more likely to be FIV-positive in the later 3-year period (p < 0.3).

Overall, the prevalence of FIV was higher, and for FeLV was lower, compared with other U.S. studies. Curiously, FIV prevalence for males increased despite increased spay/neuter opportunities across these years (p < 0.03 for trend). This increasing trend suggests the need for continued testing and surveillance of FIV in free-living, community cats. The aim of this study was to investigate a possible correlation among the histological alterations and the presence of CD3 + T and CD79 + B cells with the presence of amastigotes of Leishmania infantum in renal tissue of dogs naturally affected by visceral leishmaniasis. Two groups of dogs were used: G1 with 24 dogs with amastigotes in renal tissue, and G2 with 43 dogs with no evidence of the parasite in the kidneys. The prevalence of renal disease in the population studied was 100% in G1 and 56.1% in G2. All dogs from G1 and 53.6% of the dogs from G2 had urine protein/creatinine ratio (U/PC) > 0.5, demonstrating a positive correlation between the occurrence of severe proteinuria and the immunostaining of Leishmania amastigotes in renal tissue (p < 0.001). All dogs from G1 and 88.4% of the dogs from G2 had glomerular lesions, and the presence of the parasite in the kidney was correlated with a higher prevalence and intensity of the lesions (p < 0.001). Immunostaining for CD3 + T cells were observed in 100% of the dogs from G1 and in 97.7% of G2. There was a correlation between the presence of amastigotes in renal tissue and the intensity of the inflammatory infiltrate (p < 0.0005). Immunostaining for CD79 + B cells were observed in all dogs from G1 and in 55.8% of the dogs from G2 (p < 0.0001). Dogs with parasites in the renal tissue presented a greater intensity of B cells infiltrate (p < 0.0001). The presence of Leishmania, CD3 + T and CD79 + B lymphocytes in the kidneys of dogs naturally affected by visceral leishmaniasis suggest the involvement of both the parasite, humoral and cellular immunity in the pathogenesis of renal injury. Transient fever is a common clinical finding in B. henselaeinfected cats and bacteremia can be prolonged. The bacterial load of B. henselae in blood can be estimated by a recently developed quantitative real-time (qPCR) assay, but the assay results have not been evaluated in client-owned cats. The objective of this study was to compare B. henselae qPCR results using blood from naturally exposed cats with and without fever.

New primers and a hydrolysis probe were designed for use in a new qPCR assay to amplify a segment of the groES gene of B. henselae. The assay was optimized for annealing temperature and concentrations of primers and probe and the analytical sensitivity and specificity were determined. The assay was used to estimate the bacterial load in individual samples by comparison to a standard curve. Blood samples were collected from client-owned cats presenting to veterinary hospitals in Bangkok, Thailand, and included 22 cats with fever of unknown cause and 178 cats without fever. Total DNA extracted from blood samples were initially assayed for Bartonella specific DNA using conventional PCR assay (cPCR). All samples that were positive for B. henselae DNA in the cPCR assay and 12 samples that were negative in the cPCR assay were subsequently assayed in the qPCR assay.

Bartonella henselae DNA was amplified by both PCR assays from blood samples of 6 febrile and 21 afebrile cats. The median bacterial load of febrile cats (median; 2470, range; 21 -7467 CFU/ml) was about 50 times higher than that of afebrile cats (median; 49, range 19 -3494 CFU/ml), although the difference was not statistically significant (P = 0.15).

The results from the two PCR assays had 100% agreement. The B. henselae bacterial load was numerically greater in cats with fever. Based on these results, determination of bacterial load may be more predictive than reporting positive or negative results when attempting to associate fever with B. henselae infection in cats. A larger study is indicated to further evaluate these findings. CPV and CDV antibody titers were determined in 399 serum samples using standard reference laboratory methods with defined cutoff values for PAT. The samples were also tested for CPV or CDV PAT using a point-of-care immunochromatography assay (Canine VacciCheck, Biogal). According to manufacturer instructions, samples with PAT have color density at least equal to a positive control dot. Sensitivity, specificity, and overall accuracy of the immunoassay for CPV and CDV PAT were calculated using the gold standard reference laboratory antibody titers.

Immunoassay sensitivity was 97% for CDV and 99% for CPV. Specificity was 75% for CPV and 79% for CDV. Many of the false positive reactions were in samples with antibody titers near the reference laboratory PAT cutoff. Overall diagnostic accuracy was 90% for CDV and 94% for CPV.

The Canine VacciCheck immunoassay has acceptable accuracy for detection of CPV and CDV PAT although dogs with antibody titers near the PAT cutoff can test false positive. Shelter staff can use this tool during disease outbreaks to identify dogs at low risk for infection based on presence of PAT. Several thousand new cases of feline injection site sarcoma (FISS) occur as a consequence of vaccination each year. Radical surgical excision of tumors in the limbs and torso is often disfiguring, painful, and expensive, prompting many cat owners to avoid curative treatment. The purpose of this pilot study was to assess alternatives to currently recommended vaccination sites in terms of preference by oncology practitioners, ease of injection, and serological responses.

Surgical, radiation, and medical oncology practitioners were surveyed regarding their preference for vaccination sites based on the ease of tumor resection. A 6-point Likert scale was used to measure behavioral reaction to vaccination (1 = no reaction, 6 = injection not possible) of 60 adult cats when injected subcutaneously in the distal hind limb or the distal tail. Serum collected before and 1-2 months after vaccination was tested for antibody titers against feline panleukopenia virus (FPV) and rabies virus (RV).

The vaccination sites ranked excellent by 94 oncology practitioners were a traditional site of below the stifle (46%) and a novel site of the tail (41%). A total of 31 cats were vaccinated below the stifle, and 29 were vaccinated in the distal third of the tail. More cats accepted tail vaccination with a low behavioral reaction score of 1-2 (95%) than hind limb injection (78%) (P = 0.03). Of the cats that were seronegative for FPV at the time of vaccination, 100% developed protective antibody titers ( ! 40) 1-2 months following vaccination. For cats seronegative for RV, all but one (tail vaccine) developed adequate antibody titers ( ! 0.5 IU/mL).

Both tail and hind limb vaccination were well-tolerated and elicited similar serological responses. This pilot study provides proof-of-concept for vaccination in an anatomic site that can be amputated without severe disfigurement. In addition, since tail amputation is a minor surgery that general practitioners can perform on an outpatient basis, more cats afflicted with FISS may have access to curative surgery.

PROGUANIL AGAINST BABESIA GIBSONI AND EFFI-CACY OF THE COMBINATION THERAPY FOR ACUTE CANINE BABESIOSIS. A Iguchi 1 , A Matsuu 1 , H Ikadai 2 , N Shiranaga 3 , Y Hikasa 1 . 1 Tottori University, Tottori, Japan, 2 Kitasato University, Aomori, Japan, 3 Shiranaga Animal Hospital, Yamaguchi, Japan.

Malarone â (GlaxoSmithKline, UK), which is commercially available for treatment of human malaria, contains atovaquone (ATV) and proguanil (PG). The efficacy of combination of ATV and PG against Babesia gibsoni has not been examined. The purpose of this study was to evaluate in vitro interaction of ATV and PG against B gibsoni and the clinical efficacy of this combination therapy with Malaroneâ for acute canine Babesiosis.

For in vitro examination, two culture strains including ATVsensitive and ATV-resistant B. gibsoni were prepared. A modified fixed-ratio isobologram method was employed and the sums of the fractional inhibitory concentrations (∑FICs) for ATV and PG combination were calculated. As a result, the interactions between ATV and PG against both ATV-sensitive and ATVresistant B. gibsoni strains were found as synergistic.

To evaluate the clinical efficacy of this combination therapy, three dogs were experimentally infected with B. gibsoni. The infectious experiments were performed in the Kitasato University's laboratory, and the University's Animal Care and Ethics Committee approved the use of animals. When parasites were detected by blood smears and PCV decreased to under 20%, Malarone â was administered orally (17-25 mg/kg ATV and 7-10 mg/kg PG, bid for 10 days). The parasites disappeared and the clinical signs were improved quickly, however, parasites reappeared by 26 days after the last treatment. A single nucleotide mutation in B. gibsoni cytochrome b gene (M121I) was detected commonly from all dogs after the treatment. One of three dogs relapsed on 30 days after the last treatment. For this dog, when the second Malarone â treatment was performed at the same dose and duration of administration as the first treatment, parasites disappeared quickly and clinical signs were recovered. Although severe side effects were not found in all dogs, temporary vomiting and hyperphosphatasia were recognized in each one dog.

Furthermore, Malarone â was administered to two clinical cases. Both dogs were diagnosed as acute canine Babesiosis based on the blood smear and clinical signs. Malarone â was administered to these dogs with owner's consent. One dog recovered soon after Malarone â treatment at the same dose and duration as mentioned above, but relapsed on 100 days with M121I parasites. Then, the second Malarone â treatment with doxycycline (orally 5 mg/kg, bid for 30 days) was performed. After this treatment, the recurrence was not caused for 72 days. Another dog was administered Malarone â with doxycycline at the initial visit, and the recurrence was not caused for 80 days.

These results indicated that the combination of ATV and PG was effective for acute canine Babesiosis without severe side effects, but could not eliminate the parasite completely. Combinations with another drugs such doxycycline may prevent the recurrence of the Babesiosis. Currently, the serovar Copenhageni belonging to Leptospira interrogans serogroup Icterohaemorrhagiae is the serovar most frequently found in dogs and humans in metropolitan areas of Brazil. Despite the recommendation to include in the vaccine representative for serovars most prevalent in the region, the serovar Copenhageni is not included in the vaccines for dogs commonly used in Brasil. Some authors suggest the existence of crossprotection among serovars included in the same serogroup, but this condition has not yet been sufficiently clarified for serovars Icterohaemorrhagiae and Copenhageni. In the present study, 29 adult dogs two to six-year-old, that had been vaccinated annually with commercial vaccine containing Canicola, Icterohaemorrhagiae, Grippotyphosa and Pomona bacterins were evaluated as to the immune status against leptospiral infection before and 30 days after the yearly booster vaccination. Microscopic agglutination test (MAT) and in vitro growth inhibition test (GIT) were performed to search for antileptospires agglutinins and neutralizing antileptospires antibodies, respectively, for serovars Canicola and Icterohaemorrhagiae, and additionally, for serovar Copenhageni, not included in the vaccine. The results showed that the immunity conferred by the vaccine to serovar Icterohaemorrhagiae is more lasting than that observed for serovar Canicola, since neutralizing antibody titers> 1.0 log10 (minimum protective titer) were observed before the booster vaccination with no substantial increase after revaccination. As for serovar Canicola, revaccination resulted in a considerable increase in neutralizing antibody titer when compared to that observed previously to the revaccination (p = 0.001). The analysis of the data obtained by GIT allowed us to conclude that dogs given vaccine containing Icterohaemorrhagiae bacterin did not produce enough neutralizing antibodies against serovar Copenhageni to inhibit leptopiral growth at the same level as occurred for the homologous serovar. Despite this, the GIT titer found for serovar Copenhageni before and after revaccination showed that at least, some level of protection could be expected for dogs vaccinated with serovar Icterohaemorrhagiae bacterin, not a complete cross protection, though. While cats are frequently presented to veterinarians in Singapore, the prevalence of common feline infectious agents in the region is currently unknown. The study objective was to determine the prevalence of feline leukaemia virus (FeLV) serum antigen, serum antibodies against feline immunodeficiency virus (FIV), Bartonella spp., and Toxoplasma gondii, and DNA of the vector borne pathogens Anaplasma spp., Bartonella spp., Ehrlichia spp., Mycoplasma haemofelis (Mhf), Candidatus M. turicencis (Mtc), Candidatus M. haemominutum (Mhm) and Rickettsia spp. in the blood of domestic cats in Singapore.

Whole blood was collected from 212 cats (117 client-owned cats, 95 cats from 3 rescue centers). FeLV antigen and FIV antibodies were detected by a commercial kit (SNAP â FIV/FeLV Combo Test; IDEXX Laboratories) and T. gondii and Bartonella spp. IgG were detected by ELISA. Total DNA was extracted from whole blood and previously published PCR assays used to amplify DNA of the haemotropic organisms.

Overall, 34 cats (16%) were positive for FIV antibodies and 33 (16%) were positive for FeLV antigen. Of the FeLV positive cats, 23 of 95 (24%) were shelter housed and 10 of 117 (9%) were client-owned. Dual FIV and FeLV infection was detected in 10 cats (5%). Mhm DNA was amplified from 21 cats (10%), Mhf DNA from 4 cats (2.3%), Mhf and Mhm DNA from 3 cats (1.4%), and DNA of either Mhf or Mtc that could not be sequenced was amplified from 1 cat (0.5%). DNA of B. clarridgeiae and of B. henselae were each amplified from 2 cats (0.9%). Anaplasma spp., Ehrlichia spp., or Rickettsia spp. DNA was not amplified from any cat. Toxoplasma spp. or Bartonella spp. IgG were detected in serum of 10 of 203 cats (4.9%) and 25 of 203 cats (12.5%), respectively. Overall, evidence of an infectious agent was detected in 102 (48.1%) cats.

This study, the first of its kind in the Singapore, demonstrates a high prevalence of retrovirus infection in domestic cats, with higher rates of FeLV infection in rescue center cats. Evidence for Bartonella spp., hemoplasmas and T. gondii infections was also common but numerically lower than in other similar studies in the region. Animal hoarders accumulate animals in over-crowded conditions without adequate food, water, sanitation and veterinary care. As a result, animals seized from hoarding situations frequently suffer from a variety of medical conditions including upper respiratory infections, gastrointestinal disease, parasitism, starvation, and other evidence of neglect. The purpose of this study was to determine infectious diseases in cats seized during large-scale cat hoarding responses.

Records were reviewed retrospectively from 4 large-scale seizures of cats from failed sanctuaries from November 2009 through March 2012. The number of cats seized in each case ranged from 387 to 708. Cats were tested for heartworm, FeLV, and FIV. Cats were screened for dermatophytosis in one case. A subset of cats exhibiting signs of upper respiratory disease or diarrhea were tested for infections by PCR and fecal flotation for treatment planning.

Mycoplasma spp. (78.3%), feline calicivirus (77.8%), and Streptococcus equi subspecies zooepidemicus (55.4%) were the most common respiratory infections. Feline enteric coronavirus (89.6%), Giardia spp (55.1%), Clostridium perfringens (49.3%), and Tritrichomonas foetus (39.1%) were most common in cats with diarrhea. The seroprevalence of FeLV and FIV was 7.8 and 7.7%, respectively. Microsporum canis was cultured from 9.8% of cats in one case.

Cats from hoarding cases have high risk for enteric and respiratory infections, retroviruses, and dermatophytosis. Case responders should be prepared for mass treatment of infectious diseases and should implement protocols to prevent transmission of feline or zoonotic infections during the emergency response and when transferring the rescued cats to other shelters or to adopters. Bacterial culture of refrigerated urine samples is most accurate when performed within 24 hours. Urine cultured immediately, stored under refrigeration, or stored at room temperature beyond 24 hours has not been compared. There is evidence that transport media provide accurate culture results for conjunctival scrapings and group B streptococcus isolates.

This study evaluates culture reliability of sterile urine inoculated with bacteria from clinical isolates of canine urinary tract pathogens at known concentrations and subsequently stored under various conditions for up to 7 days.

A 66 ml urine sample was obtained via cystocentesis from a healthy research dog. Urinalysis and immediate aerobic culture on Columbia blood agar and MacConkey were performed to rule out infection and sample contamination. The remaining urine was divided into 4 samples and inoculated with either Escherichia coli or Staphylococcus psuedointermedius at concentrations of 10 2 or 10 5 cfu/ml. Each portion was further divided and stored in plain sterile tubes under refrigeration at 4°C (PST-FR), at room temperature (PST-RT) or Amies transport medium without charcoal (ATM). All samples were run in triplicate at 0, 24, 48, 72, and 168 hours (7 days) .

ANOVA demonstrated that interaction between storage method and time significantly affects bacterial counts (p < 0.05) for both bacteria. Storage beyond 24 hours resulted in decreased positive cultures for PST-FR and PST-RT. Incubation for at least 24 hours in ATM resulted in superior bacterial recovery under typical aerobic culture conditions, suggesting that ATM may be a preferred storage method when delayed urine culture or low bacterial concentrations are anticipated. The Accutest Uriscreen (Jant Pharmaceuticals, Co.) is a rapid, catalase-based test for detection of bacteria and pyuria in urine. It is used as a screening test for urinary tract infections in human patients and has been evaluated for the detection of bacteriuria and bacteremia in neonatal calves. There are no published reports regarding the application of the test for the detection of urinary tract infection in domestic animals. The purpose of the study was to determine the accuracy of the test compared to urine sediment evaluation and microbial culture.

Urine samples (n = 165) obtained by cystocentesis from 141 dogs and 19 cats were submitted for urinalysis and microbial culture. Excess urine was analyzed with the Uriscreen per manufacturer's recommendations. The sensitivities, specificities, negative predictive values, and positive predictive values were calculated using culture as the gold standard for the presence of infection.

Sensitivities for the Uriscreen, confirmed microscopic bacteriuria, microscopic pyuria (>3 leukocytes/high power field), and combined microscopic bacteriuria and pyuria were 89%, 70%, 67% and 61%; specificities were 71%, 95%, 92%, and 97%; positive predictive values were 38%, 76%, 62% and 84%; and negative predictive values were 97%, 94%, 92%, and 93% respectively.

In conclusion, the Accutest Uriscreen can be used as a screening test for the presence of urinary tract infection in dogs and cats with higher sensitivity and negative predictive value than either microscopic bacteriuria and/ or pyuria. A negative result supports the absence of urinary tract infection; a positive result should be followed with microbial culture. The prevalence of urinary tract infections (UTIs) in older cats has been estimated to be between 12 and 30%. Typically these cats had concurrent disease and frequently the UTIs were clinically silent. A prospective, longitudinal study was designed to determine the prevalence, incidence and risk factors for asymptomatic bacteriuria in healthy, older cats.

Sixty seven healthy, adult cats ( ! 7 years) were tested up to four times over two years. Cats were obtained from the Feline Nutrition Unit, Massey University. Cats were group housed and weighed weekly. Healthy cats were defined as non-azotaemic and without clinical signs of urinary tract disease or persistent weight loss. Urine samples for urinalysis and quantitative urine culture were obtained by cystocentesis. Serum creatinine concentration was determined at each test. An incident infection was defined as a positive urine culture in a cat whose preceding culture was negative. Active urine sediments were defined as the presence of pyuria or haematuria (> 5 cells /high powered field).

Creatinine, body weight, urine specific gravity, sex and age were evaluated as potential risk factors for a positive urine culture. A generalised estimating equation with an exchangeable correlation structure was used to account for clustering of urine culture results within animal. Nested models were compared using the Wald test statistic.

Overall, 11 cats had asymptomatic bacteriuria during the study with 24 positive cultures from 221 samples. The prevalence of asymptomatic bacteriuria at different sampling times was between 10 and 13%. Incident infections were uncommon with only four new cases over the two years. Most positive cultures had active urine sediments (19/20 standardized urinalyses) and E. coli was the most common bacterial identified (13 of 16 of bacteria identified, 81%). Of the above variables, only sex was significantly associated with asymptomatic bacteriuria. Female cats were 16.8 times more likely to develop an occult UTI (95% CI 15.1-18.4, p < 0.001).

Asymptomatic bacteriuria was present in between 10 and 13% of healthy, older cats. Female cats were more likely to have asymptomatic bacteriuria but there was no association between a positive urine culture and body weight or serum creatinine concentration. The clinical relevance and optimal treatment strategy of asymptomatic bacteriuria requires further investigation. Leptospire species are commonly associated with acute renal failure in dogs and residual damage has been suggested as a cause of canine chronic kidney disease as well. While chronic kidney disease is common in cats, this species has been considered to be naturally resistant to the development of leptospirosis. However, in a previous study in our center, urine was collected from shelter cats and Leptospire species DNA was amplified in 10 of 85 (11.8%) showing that cats may not only harbor but potentially shed the microorganisms. The purpose of this study was to determine if there is an association between presence of antibodies to Leptospire species in cats with and without azotemia (creatinine > 2 g/dl).

In order to validate the microscopic agglutination test (MAT) for use in cats, a four serovar canine leptospiral vaccine was administered to two specific pathogen free (SPF) adult laboratory housed cats on days 0 and 14. Serum taken from the cats on days -7, 0, and weekly thereafter for a total of 12 weeks was evaluated for presence of anti-leptospiral antibodies using a commercially available MAT. MAT was performed for the Leptospira serovars Canicola, Grippotyphosa, Hardjo, Icterohaemorrhagiae, Pomona, and Bratislava with a cutoff value of ! 1:100. Sera from 71 client-owned azotemic cats and 90 client-owned non-azotemic cats enrolled in a previous study were tested for Leptospiral species antibodies. The prevalence of positive results for Leptospiral species MAT were determined as well as the mean titers. A chi-squared test was used to analyze the data and a p value of < 0.05 was considered significant.

Both SPF cats seroconverted by week nine. In the clientowned cats, there was no statistically significant difference between the compared groups. The prevalence (1/71; 1.4%) and mean titers for Leptospira grippotyphosa were higher in azotemic versus non-azotemic cats (0/90; 0.0%). In contrast, the prevalence (4/90; 4.4%) and mean titers for Leptospira canicola were higher in non-azotemic as compared to azotemic (0/71; 0.0%) cats. The prevalence and mean titers for the other serovars when compared between azotemic and non-azotemic cats was approximately the same. There does not appear to be an association between Leptospire species antibodies and azotemia in client-owned cats but further studies are needed in order to determine if leptospirosis contributes to feline chronic kidney disease. Persistent or recurrent lower urinary tract infections (UTIs) are commonly diagnosed in companion animal veterinary medicine. Urine obtained by cystocentesis is routinely cultured to identify pathogens involved in UTIs; however, these may be falsely negative because of intermittent shedding of bacteria especially from intracellular uropathogenic Escherichia coli (E.coli.) Culture of surgically-obtained full thickness bladder biopsies has been recommended to more accurately identify urinary pathogens. Transurethral cystoscopy is a minimally invasive method for examining the lower urinary tract and obtaining biopsies for bacterial culture and histopathology; however it is not known whether this technique improves bacterial identification and is worth the extra expense to the owner. Therefore, the objective of this study was to compare bacterial growth from aerobic culture of urine obtained by cystocentesis and transurethral cystoscopically-obtained mucosal bladder biopsies in the dog.

Medical records of all dogs that underwent cystoscopy and urethroscopy at a veterinary teaching hospital between the years 2002-2011 were examined. To be included in the study, the dogs must have had cultures of cystocentesis-obtained urine and bladder mucosal biopsies obtained via transurethral cystoscopy within a 2 week time period without differing antibiotic therapy. Forty dogs were included in the study. Bacteria identified from the cultures of the urine and biopsy samples were compared.

There was a significant agreement between urine and bladder mucosal biopsy culture results (P = 0.0003). Both types of cultures were negative in 66% and positive in 17% of patients. There was a 17% disagreement between groups. Although the result was not statistically different, there were two mucosal biopsy samples that revealed E.coli not present in the corresponding urine cultures. Predisposing conditions to UTI were common in the study population.

In conclusion, there was significant agreement between urine and bladder mucosal biopsy culture results both in the presence and identification of pathogenic bacteria. Therefore, transurethral cystoscopy in order to obtain mucosal biopsies for culture for pathogens not identified in a urine sample is not warranted in most dogs. The exception may be those suspected to be infected with uropathogenic E.coli. Pyonephrosis, or renal pelvis abscessation, is commonly associated with a ureteral obstruction. In humans emergency decompression with ureteral stenting or nephrostomy tube placement is recommended. The authors describe the technical and clinical outcome of endoscopic and/or fluoroscopic-guided ureteropelvic lavage and ureteral stent placement for the treatment of obstructive pyonephrosis in dogs.

Thirteen dogs were retrospectively reviewed for presenting signs, pre/post-procedural clinical and imaging data, and shortand long-term outcomes. All were septic on presentation with obstructive ureterolithiasis in 11/13. Eleven had a positive culture from the bladder (n = 8) or renal pelvis (n = 3) (E. coli[7], Staph spp. [3] , and Klebsiella spp.[1]). Nine dogs were thrombocytopenic (median 63x10 3 , range 16-654k). Stents were placed endoscopically (n = 10) or with surgical-assistance (n = 3). Median hospitalization was 48 hours. Ten dogs were alive at the end of the study (median 150, mean 283 days). One dog was euthanized 2 days post-operatively for decompensating sepsis and 2 died of unrelated causes (5 months and 4 years post-stent). Two intraoperative complications included stent occlusion from shearing of a guidewire and wire penetration of the ureter at an embedded stone. Short-term complications included a bladder hematoma (n = 1) and transient dysuria (n = 3), which resolved without intervention. Long-term complications included stent encrustation requiring exchange (n = 1), recurrent UTIs (n = 2) and tissue proliferation at the ureterovesicular junction (n = 5). All patients lived with their stents in place and patent long-term.

This interventional procedure was a successful renal-sparing treatment for pyonephrosis. Patients had few complications and this technique could be considered over traditional surgery. Circumcaval (retrocaval) ureters are a rare embryological malformation resulting in ventral displacement of the caudal vena cava, which crosses the ureter, potentially causing a ureteral stric-ture. This retrospective study evaluates feline patients with circumcaval ureters that have associated ureteral obstructions. The objective is to report this condition in a series of cats and evaluate their short-and long-term outcome after interventional treatment.

There were 153 obstructed ureters in 114 cats. Twenty-four of 153 (15.6%) obstructed ureters, in 21 cats (18.6%), were circumcaval. Fifteen were right-sided, 3 left-sided, and 3 bilateral (75% right; 25% left). Causes of obstruction included: ureterolithiasis (n = 12); stricture (n = 7) or both (n = 5). The obstructive lesion was in the proximal ureter, at or just cranial to the caval-crossing in 23/24. The median pre-and post-op creatinine was 7.6 and 3.1 mg/dL, respectively. The initial treatment included: subcutaneous ureteral bypass device (SUB) (n = 12), ureteral stent (n = 12) and ureteronephrectomy (n = 1). Nineteen patients (90%) survived to discharge with a median hospitalization time of 6 days. Re-obstruction occurred in 4 ureters, all of which had stents. All SUBs remained patent long-term. The median survival time was >377 days (range, 2-1491 days) with 11 cats still alive. No cat died of recurrent ureteral obstruction and 7/10 deaths were due to kidney disease progression.

This study identified a group of ureterally obstructed cats with a circumcaval ureter at, or near, the obstructed site. In this series, a SUB was considered a better treatment option than a stent when a circumcaval ureter was identified. Cobalamin deficiency occurs concurrently with a variety of feline diseases and adversely affects cellular metabolism. An association between serum cobalamin or folate concentrations and chronic kidney disease (CKD) has not been reported in the feline literature. The purpose of this study was to compare serum cobalamin, folate, and methylmalonic acid concentrations in healthy cats and cats with CKD.

Serum concentrations of cobalamin and folate were determined by automated chemiluminescent competitive binding immunoassays and methylmalonic acid concentrations by gas chromatography-mass spectrometry. Twenty-seven healthy cats (determined by historical, physical, and clinicopathologic examinations) and 35 cats with CKD (defined as IRIS stages II through IV) were enrolled. Cats with CKD that had incomplete clinicopathologic data or that were diagnosed with gastrointestinal or pancreatic disease were excluded.

Cats with CKD had lower median serum cobalamin concentrations (920 vs. 1665 ng/L, p = 0.001) and higher median serum methylmalonic acid concentrations (481 vs. 289 nmol/L, p = 0.002) than healthy cats. No difference in serum folate concentrations was demonstrated between the two groups. For cats with CKD, a correlation between serum creatinine and serum cobalamin concentrations was not detected (Spearman's r = 0.185, p = 0.2883), while a trend towards a weak correlation between serum creatinine and methylmalonic acid concentrations was demonstrated (Spearman's r = 0.311, p = 0.0734).

Although nearly all cats with CKD had serum cobalamin and methylmalonic acid concentrations within the laboratory's established reference ranges, the differences between healthy cats and cats with CKD suggest impaired function of cobalamin-dependent cellular metabolism. Urinary tract infection (UTI) is common in dogs and early diagnosis is mandatory to limit complications. UTI is also a com-mon finding in chronic kidney disease (CKD) and can lead to acute kidney injury (AKI). The gold standard for the diagnosis of UTI is urine culture, but urinalysis is also frequently used for this purpose: it is cheap, reliable and easy to perform, but its sensitivity and specificity decrease in the presence of dilute urine or free catch sampling. Urinary NGAL (uNGAL) is a new biomarker used in the assessment of AKI and CKD, but the impact of UTI on uNGAL in dogs is unknown. The aim of this study was twofold: first, to determine if uNGAL is influenced by UTI and second, to determine a cut-off value that could identify UTI in non-azotemic adult dogs.

In this prospective blinded controlled study, 99 non-azotemic adult privately-owned dogs were recruited and divided in 3 groups: 25 dogs suspected of having a UTI and showing a positive urine culture (UTI group), 55 dogs suspected of having a UTI but with a negative urine culture (Negative goup) and 19 healthy control dogs with a negative urine culture (Healthy group). uNGAL and uNGAL/Cr were determined in all dogs using a canine specific ELISA kit (BioPorto Diagnostics A/S).

Median (IQR) uNGAL was significantly higher in the UTI group (14.22 ng/ml; 3.77-44.89) compared with the Negative (1.13 ng/ml; 0.56-4.18) and Healthy groups (0.24 ng/ml; 014-1.4) (p < 0.0001). Similarly, median uNGAL/Cr was also significantly higher in the UTI group (p < 0.0001) as compared to the Negative and Healthy groups. uNGAL was significantly but weakly correlated with age (p = 0.03, r² = 0.04), weight (p = 0.03, negative correlation, r² = 0.03) and pyuria (t-test, p < 0.0001). ROC curve analysis determined that a uNGAL cut off value of 3.38 ng/ml yielded a sensitivity and specificity of 76% and 70% respectively for UTI diagnosis. The positive and negative predictive values were 54% and 87%, respectively.

This study demonstrates that uNGAL is elevated in the presence of a UTI. This finding highlights that interpretation of uNGAL in CKD and AKI should take into account the presence of UTI. This study also shows that awaiting culture results, a uNGAL < 3.38 ng/ml could be useful, to rule out UTI in a nonazotemic dog. However, sensitivity and specificity of uNGAL to diagnose UTI in dogs does not appear to be superior to what is reported for routine urinalysis. Further investigations are needed to determine the influence of pyuria alone on uNGAL. The association between hypertension and feline chronic kidney disease (CKD) is well recognised and the prevalence of hypertension in CKD has been reported to be between 19.4% and 65%. The objectives of this study were to assess the prevalence of hypertension in cats with CKD seen in a first opinion clinical setting and to investigate subsequent changes in systolic blood pressure (SBP) over time in cats that were initially normotensive.

Cats with CKD ! 9 years old that were examined for the first time at two first opinion practices in London, UK between August 2000 and August 2012 and with a follow up period of ! 3 months and a minimum of 3 visits were retrospectively identified. CKD was diagnosed in cats with renal azotemia (plasma creatinine concentration >2.0 mg/dl either on two consecutive visits or in conjunction with USG <1.035). SBP was measured at all visits by a non-invasive Doppler technique. Cats with SBP ! 170 mmHg on two consecutive visits or SBP ! 160 mmHg with concurrent evidence of hypertensive retinopathy/choroidopathy were classified as hypertensive. Exclusion criteria were concurrent hyperthyroidism or treatment with medications known to alter SBP. Re-examination of cats was offered at a maximum of 8 week intervals. The slope of SBP over time curves were compared between cats that developed hypertension during follow-up (DH-group) and those that remained normotensive throughout (NT-group) using a mixed effects model. SBP at diagnosis of CKD was compared in the two groups using a Welch two sample t-test. In the DH-group the actual SBP measurements that resulted in the diagnosis of hypertension were not included in the analysis. In the NT-group all available data were included. Results are expressed as mean AE standard deviation.

Two hundred and sixty eight cats met the inclusion criteria. Of these, 102 cats (38.1%) were hypertensive at diagnosis of CKD or were diagnosed as hypertensive within 3 months of CKD diagnosis and 33 cats (12.3%) became hypertensive ! 3 months after diagnosis of CKD (DH-group). The remainder (133 cats; 49.6%) remained normotensive throughout follow-up (NTgroup). At diagnosis of CKD, mean SBP was significantly higher in the DH-group than in the NT-group [154.2 AE 14.1 vs. 135.0 AE 17.0 (P < 0.0001)]. Cats in the DH-group were diagnosed as hypertensive a median of 385 [range days after diagnosis of CKD. Cats in the NT-group had a median of 329 [range 91-2037] days of follow up. The slope of SBP over time was significantly greater in the DH-group than the NT-group [1.7 AE 1.8 mmHg per 100 days vs. 0.5 AE 1.6 mmHg per 100 days (P < 0.001)].

About 12% of cats that are normotensive at the time of CKD diagnosis will develop hypertension over time. Interestingly these cats have higher baseline, and a greater rate of increase, of SBP over time than their nomotensive counterparts. The high prevalence of hypertension in azotemic cats in this study shows the importance of monitoring of SBP. Symmetric dimethylarginine (SDMA) is a small molecule formed by methylation of arginine that is released from proteins into the blood during protein degradation. SDMA is primarily eliminated by renal excretion and might therefore serve as an endogenous marker of glomerular filtration rate (GFR). The objective of this study was to determine whether SDMA correlates with GFR in dogs with chronic progressive renal disease.

Seven male dogs affected with X-linked hereditary nephropathy (XLHN) and 3 unaffected male littermates were serially evaluated. sCr and SDMA were measured weekly; GFR was determined monthly. SDMA was determined by liquid chromatography-mass spectrometry operating in multiple reaction monitoring mode using a stable isotope dilution assay. GFR (estimated by iohexol clearance) was determined by injecting 300 mg I/kg followed by blood sampling at 5, 15, 30, 60, 120, 180, 240, and 360 minutes post-injection. Iohexol concentration was measured at Michigan State University, and GFR was calculated by non-compartmental analysis using commercial software (WinNonlin 6.3, Pharsight). Correlation of GFR with SDMA and sCr was determined using linear regression.

In unaffected dogs, SDMA remained stable (mean: 12 lg/dl; range: 9-16 lg/dl). SDMA increased in affected dogs (up to 100 lg/dl) during disease progression, corresponding to an increase in sCr and decrease in GFR. In affected dogs, correlations of sCr and SDMA with GFR were R 2 = 0.99 and R 2 = 0.95, respectively.

In conclusion, SDMA correlates well with GFR as estimated by iohexol clearance in dogs with progressive renal disease and might be a useful addition to sCr in the assessment of renal function. Dogs with chronic kidney disease (CKD) may progress with proteinuria according to the renal disease or in consequence of progressive loss of nephrons as glomerular hyperfiltration may develop in the remaining nephrons causing proteinuria. Glomerular proteinuria is characterized by urinary proteins loss of high molecular weight (HMW > 60 kDa~albumin), and tubular proteinuria of low molecular weight (LMW < 60 kDa). Several studies suggest that increase of proteinuria and albuminuria indicates the progression of CKD, and albuminuria may predominate when protein-to-creatinine ratio (UPC) is above 2.0. The aim of this study was to sequentially evaluate quantitatively (UPC and albumin-to-creatinine ratio UAC) and qualitatively (SDS-PAGE) the urinary proteins during the course of CKD, and to identify the segments of the nephrons that could be damaged. Eleven adult dogs with CKD, in stages 2 and 3, were divided in two groups (data recorded every 30 to 40 days; followed-up over 6 to 15 months). Group I was composed of 5 proteinuric dogs (8 to 10y-old) and Group II included 6 non-proteinuric (3 to 14yold); exclusion criteria were any concomitant diseases that could cause secondary renal disease as well as pre-or post-renal proteinuria. Control group was composed of 14 clinically normal adult dogs (punctual observation). In Group I, 3 out of 5 CKD dogs had UPC (Coomassie Blue) above 2.0 throughout the course (ranging from 2.1 to 12.8) and UAC (ELISA Immunology Consultants Laboratory) ranged from 0.017 to 0.15 (microalbuminuria = UAC 0.03 to 0.3); other 2 out of 5 CKD dogs had UPC ranging from 0.35 to 1.85 and UAC from 0.003 to 0.05 (microalbuminuria was detected only once in one dog). Thus in Group I, normal values of UAC predominated and no macroalbuminuria (UAC > 0.3) was detected, and SDS-PAGE revealed that those CKD dogs with UPC > 2.0 showed higher percentage of LMW proteinstubular pattern (84 to 100% of total proteins and 4 to 6 bands), as well as CKD dogs with UPC < 2.0 (40 to 100% of total proteins and 4 to 6 bands) when compared to control group (LMW proteins= 55.4 AE 12.5%; meanAESEM, and scarce or 3 to 4 bands of proteins) (unpaired t test; p < 0.05). All dogs of Group II had UPC < 0.6 and UAC < 0.006, however the qualitative proteins evaluation showed that the majority of dogs had tubular pattern (LMW proteins > 50% and increased number of proteins bands along the course of disease (4 to 5 bands), although no statistical difference was detected. Thus, the combined quantitatively and qualitatively as well as the serial evaluation of urinary proteins could provide more information regarding to the origin of proteinuria, suggesting that tubular involvement was the main renal alteration, including for those CKD dogs with UPC > 2.0, and thus adding more information for adequate diagnosis and therapy. Angiotensin receptor blockers (ARBs) may have an important place in feline medicine, particularly in cats with kidney or cardiovascular disease; however, their utility has not been systematically investigated in this species.

Six healthy intact male cats with telemetric arterial catheters were used to compare efficacy of once daily oral dosages of 3 ARBs (losartan [LOS, 3 mg/kg], irbesartan [IRB, 2, 6 and 10 mg/kg], and telmisartan (TEL, 0.5, 1 and 3 mg/kg]) compared to benazepril (BEN, 2.5 mg/cat) and placebo. Following 8 doses and~90 minutes after administration, cats were anesthetized and arterial BP was recorded before and after IV angiotensin I (Ang1) (20, 100, 500, 1000 ng/kg). This protocol was repeated 24 h post-dose for BEN and TEL (3 mg/kg). Two-way repeated measures ANOVA was used to test for differences in the change in SBP (DSBP) among groups. Significance was defined as P < 0.05.

Compared to placebo, at 90 min, all drugs except LOS produced significant attenuation of DSBP. The DSBP values for BEN, all doses of IRB and the 2 lowest doses of TEL were not significantly different. However, at 90 minutes 3 mg/kg TEL attenuated DSBP significantly more than BEN (P < 0.05 Iohexol clearance is commonly used to estimate glomerular filtration rate (GFR) in dogs, while serum creatinine (sCr) and symmetric dimethylarginine (SDMA) might be useful as endogenous markers of GFR. The variability of iohexol clearance, sCr, and SDMA assessments in dogs with chronic renal disease is unknown. The purpose of this study was to determine week-toweek variability in these measurements in female dogs with mild but stable chronic renal disease due to X-linked hereditary nephropathy (XLHN).

Twenty-four heterozygous (carrier) female dogs with XLHN were evaluated once per week for 3 weeks. Two dogs were excluded, one due to a urinary tract infection and another due to data quality issues. sCr was determined enzymatically and SDMA was determined by liquid chromatography-mass spectrometry operating in multiple reaction monitoring mode using a stable isotope dilution assay. Iohexol clearance was performed by injecting 270 mg I/kg followed by blood sampling at 2, 3, and 4 hours post-injection. Iohexol concentration was determined at Michigan State University, and iohexol clearance per dog was calculated using a one-compartment analysis and applying a quadratic correction. The variability of each analyte was determined using the power-of-the-mean model, and the reference change value percent (RCV%) was calculated.

Iohexol clearance, sCr, and SDMA were similar from week to week in each dog supporting stable renal disease in this cohort. The RCV% for iohexol clearance was 15% (95% confidence interval (CI): 10%, 20%) when iohexol clearance was low (1.5 ml/min/kg), and decreased as iohexol clearance increased (10%, 95% CI: 7%, 12%, at 3 ml/min/kg). The RCV% for sCr and SDMA was constant at 17% (95% CI: 14%, 21%) and 19% (95% CI: 15%, 24%), respectively.

In conclusion, the week-to-week variability of iohexol clearance, sCr, and SDMA in carrier females with XLHN is similar. Iohexol clearance must change by 12-20% (depending on its value) whereas sCr and SDMA must change by 21-24% in order to be 95% confident that a true increase or decrease in a subsequent measurement has occurred. Whereas glomerular diseases (GD) are considered pro-thrombotic, uremic renal diseases such as chronic kidney disease (CKD) and acute kidney injury (AKI) are mostly referred to as anti-thrombotic with increased risk of bleeding. Accurate assessment of the hemostatic status of animals with renal diseases is essential for diagnostic procedures and therapeutic interventions. However data on prevalence and types of hemostatic disorders and diagnostic value of available tests are sparse. The aim of this study was therefore to evaluate prospectively the hemostatic status of dogs with AKI (N = 187), CKD (N = 46), and GD (N = 23).

Assessment of hemostasis included platelet count, PT, aPTT, fibrinogen, and thromboelastometry (TEM). A subset of dogs (N = 73) was evaluated further with buccal mucosal bleeding time (BMBT), von Willebrand factor, protein C, protein S, factor VIII, D-dimer, and antithrombin.

Hypocoagulable status was evidenced by significantly prolonged BMBT in uremic dogs (CKD, AKI), mildly decreased platelet count (AKI), and slightly prolonged PT and PTT (AKI). Dogs with renal disease were however also characterized by prothrombotic features, including increased fibrinogen (AKI 58%, CKD 89%, GD 100%), decreased protein C (AKI 74%), decreased antithrombin (AKI 83%, CKD 50%), and hypercoagulable TEM profiles (AKI 40%, CKD 80%, GD 100%). Dogs with AKI had the widest spectrum of hemostatic disorders with 15% being hypercoagulable and 79% with mixed hypo-hypercoagulable features.

The wide range of hemostatic disorders observed in AKI and CKD emphasizes the need for a thorough individualized evaluation of affected animals. The magnitude of the changes further warrants a consideration for therapeutic interventions. The intestinal probiotic, Azodyl TM , has been advocated for the management of chronic kidney disease by enhancing the intestinal elimination of urea as a surrogate for nitrogenous toxins. However, the influence of Azodyl TM to alter intestinal urea clearance remains insufficiently validated to provide explicit therapeutic recommendation.

We evaluated the intestinal clearance of urea in four dogs (BW 30-46 kg) undergoing maintenance hemodialysis (HD) with formal urea kinetic modeling before (C) and following Azodyl TM administration (Tx) at 9 (1 dog) or 12 (3 dogs) capsules per day for 4-6 weeks. Dogs underwent twice weekly HD and were fed a constant protein and calorie intake. Urea kinetics were assessed by measurement of preHD serum urea nitrogen (pre HD SUN), weekly time-average urea concentration (TACurea), dialysis dose (eKt/V), and urea nitrogen appearance rate (G). Intestinal clearance (CI) of urea was determined as the difference between whole body urea clearance (G/TACurea) before and during Azodyl TM administration. Data in the follow-ing Conventional intermittent hemodialysis platforms designed for adult humans have extensive use in veterinary dialysis but may provide excessively rapid and unsafe treatment of severely uremic animals weighing <10 kg. To circumvent this limitation, we devised an external closed bypass loop adapted to the Gambro Phoenix that delivers product dialysate (Qd, 350-800 ml/min) at reduced flow (Qd, 1-15 ml/min) to and from the dialyzer. Dialysis intensity thus is controlled by the dialysate flow while maintaining all machine functionality at fast blood flow (Qb) to prevent clotting. Solute and fluid removal characteristics were tested on a 1.2 L in vitro simulation and in vivo in 4 cats and 1 dog with acute kidney injury using the SDBD. In vitro fluid removal was not statistically different between conventional IHD and IHD with SDBD at zero and 100 ml/hr.

Changes in blood and dialysate solute concentrations for a typical extended-slow dialysis session on a 7 kg uremic cat using SDBD at Qd = 5 ml/min and Qb = 40 ml/min are shown in the figure and Eicosanoids are potent lipid mediators involved in numerous physiologic and pathologic processes including orchestration of many aspects of the inflammatory response. Recent studies suggest that urinary bladder eicosanoid metabolism may be altered in chronic feline idiopathic cystitis (FIC). We hypothesize that changes in key eicosanoid biosynthetic enzymes (cyclooxygenase-1 and 2 [COX-1, COX-2], prostaglandin E synthases m and c [mPGES, cPGES], 5-lipoxygenase [5-LOX], and 5-lipoxygenase activating protein [FLAP]) may be involved in the pathogenesis of FIC. Of these genes, only the structure of COX-2 has been determined. Furthermore, no RT-qPCR system has been developed to examine changes in gene expression. The purpose of this study was to identify the feline homologues of these genes and to develop corresponding quantitative RT-PCR assays.

Fragments of feline target genes were identified from the partial cat genome sequence using dog and human reference sequences. Cat intron/exon structures were inferred and the predicted mRNA was translated to produce an open reading frame that correctly aligned with the proteins of experimentally verified sequences from other species. Inferred intron/exon structures were verified by amplification and sequencing of the whole cat cDNA sequences. In addition, RT-qPCR assays were developed to amplify a roughly 100 bp section within the cDNA for the six target genes and four reference genes.

The predicted amino acid sequences for all feline gene products showed considerable homology with other species. Direct sequencing confirmed RT-qPCR assay amplification products. These RT-qPCR assays are now available to examine relative changes in gene expression in FIC and other disorders. Chronic kidney disease (CKD) is one of the most common diseases affecting cats. The purpose of this study was to describe the dietary and medication patterns of cats with CKD. A web-based survey was advertised on CKD, pet, veterinary, and breed associated websites and list serves. Owners of 1090 cats with CKD participated in the study. Mean age of the cats was 13.7 AE 4.2 years and most cats (69%) lived in multi-cat households. Forty percent of cats in this study had concurrent diseases, with hyperthyroidism, heart disease, and inflammatory bowel disease being the most common.

Veterinarian recommendation remained the most common reason indicated for diet selection (66%). Many owners (41%) fed a mixture of canned and dry food, and 665 (61%) of owners fed a veterinary therapeutic diet as a component of the diet. Most owners (57%) reported that their cats did not have a normal appetite, and of these owners, 225 (52%) responded that their cats had a poor appetite or required coaxing to eat 5-7 days per week. Most cats (65%) were receiving subcutaneous fluids and 50% were administered oral medications; however, most cats were not receiving phosphorus binding medications. Forty-four percent and 41% of cats received commercial cat treats and dietary supplements, respectively. Hyporexia is a common problem for cats with CKD and may lead to cats being fed suboptimal diets for their disease. This information may be useful for treating or designing nutritional studies for cats with CKD. Variant alleles in NPSH1 and KIRREL2, the genes which encode the slit diaphragm proteins nephrin and filtrin/Neph3, respectively, were previously found associated with protein-losing nephropathy (PLN) in Soft Coated Wheaten Terriers (SCWT) by a genome-wide association study and subsequent gene sequencing of candidate genes in a statistically significant interval that differed among dogs with PLN compared with geriatric (14-18 year old) SCWT. Genotyping assays were developed for both of the single nucleotide polymorphisms (SNPs) in these genes that are in linkage disequilibrium in the breed. Homozygous positive dogs were shown to be at highest risk for the development of PLN, heterozygous dogs were at intermediate risk, and homozygous negative dogs were at low risk for the development of PLN. A prevalence study was performed to ascertain if breeders could safely remove carrier dogs in one generation. Cheek swab, blood, or semen samples were tested from 1208 SCWT dogs of all ages (median 4 yrs). Haplotypes are described as 1-1 (homozygous negative), 1-2 (heterozygous), and 2-2 (homozygous positive) for the PLN-associated variant alleles. The following table shows the frequencies found in various countries. Without genetic counseling with the knowledge of these haplotypes and assuming random breeding, the variant allele frequency would remain 43% in the USA. This high frequency indicates that it would be unwise to cull all carriers (1-2 or 2-2 dogs) of the variant alleles in one generation, thereby risking loss of genetic diversity, increased inbreeding, and the potential of increasing the incidence of other deleterious genetic traits. An approach to avoid producing high risk homozygous positive (2-2) dogs would be to preferably breed desirable heterozygous (1-2) or homozygous positive (2-2) dogs to homozygous negative (1-1) dogs.

A POTENTIAL CHRONIC KIDNEY DISEASE MARKER SYMMETRICAL DIMETHYL ARGININE. Mahalakshmi Padmanabhan, Edward Obare, Yerramilli Murthy. IDEXX Laboratories Inc., Westbrook, ME.

The methylated arginines [Symmetrical Dimethyl Arginine (SDMA), Asymmetrical Dimethyl Arginine (ADMA) and Monomethyl Arginine (MMA)] are derived from intranuclear methylation of L-arginine by protein-arginine methyltransferase (PRMT) and released into circulation after proteolysis. It is established that while ADMA is mostly cleared through hydrolysis by dimethylarginine dimethylaminohydrolase (DDAH), SDMA is mostly eliminated by renal excretion suggesting SDMA could be a potential marker for Chronic Kidney Disease (CKD). To evaluate SDMA as a marker for canine and feline CKD, a highly sensitive and specific analytical method based on LC /MS has been developed for the quantification of SDMA in biological samples and validated as per FDA and CLIA guidelines.

The LC separation was achieved using X-Bridge RP C-18 column and ion pairing agent. The API 4000 triple quadrupole mass spectrometer (Applied Biosystems/MDS Sciex) was operated in Multiple Reaction Monitoring (MRM) mode with positive electrospray interface. The MRM transition for SDMA was observed at m/z 203.2 -> 172.1. As part of the method validation, performance characteristics including sensitivity, carryover and interferences, matrix effect and recovery, linearity, accuracy and precision, ruggedness, stability, robustness and interfering substances were established. All performance metrics were within established FDA guidance for LCMS method validation.

Lower Limit of Quantitation (LLOQ) was found to be 1.56 ug/ dl; A nine point standard curve was established linearity with R 2 > 0.99; Intra and inter day CV were found to be <3% and no significant interference from hemolysis, lipemia or icterus was observed and related compounds such as arginine, MMA and ADMA had no significant impact on performance. Canine and feline serum and plasma were both shown to be acceptable sample types for the assay with no difference observed in performance.

In conclusion, we have developed and validated an LCMS analytical method for the measurement of a potential CKD marker in canine and feline samples. Urinary exogenous creatinine clearance (UECC) is considered as a good estimate of glomerular filtration rate (GFR) in cats, but is tedious and time-consuming. The objective of this study was to compare plasma exogenous creatinine clearance (PECC), previously reported as an alternative method, with UECC.

Six healthy cats (6-11 years, 3.2-4.5 kg) were placed in metabolism cages. All urine voided for 24 hours was collected. The bladder was emptied before and again 24 hours later. The emptied bladder and the cage were also rinsed with saline, and the rinsing water was collected for creatinine measurement. Exogenous creatinine (20 mg/kg) was administered by IV bolus. Blood was sampled before and then 5, 30 min, and 1, 2, 3, 5, 8 and 24 h after exogenous creatinine administration. Creatinine was assayed using an enzymatic method. Pharmacokinetic analysis was performed using a non compartmental approach. Data were compared by ANOVA. Results are expressed as meanAESD.

The proportion of creatinine recovered by rinsing the bladder and the cage was 6.5 AE 5.6% (range: 2-18%) of the total amount of creatinine excreted in urine. The UECC (2.2 AE 0.9 mL/min/ kg) was 15%-lower (P < 0.01) than PECC (2.6 AE 0.9 mL/min/ kg), but the two estimates were strongly correlated (r² = 0.97, P = 0.001).

In conclusion, PECC appears to be an acceptable and practicable alternative to UECC in cats. Dogs with severe acute kidney injury (AKI) typically display marked gastrointestinal signs, delaying nutritional support and compromising their recovery. This prospective study aimed at evaluating a new esophago-jejunal (EJ) feeding technique, in comparison to a standard esophageal (E) technique in dogs with AKI.

Twenty dogs with severe AKI treated with hemodialysis were randomly assigned to the two techniques. In the EJ group, an EJ-tube was introduced through a standard E-tube and advanced endoscopically to the jejunum. Feeding was administered continuously for at least 5 days. After removal of the EJ-tube, feeding was continued through the E-tube. In the control group E, dogs were fed 5 times daily through an E-tube. Placement technique, complications, and nutritional efficacy were compared between groups.

Feeding tubes were placed 1 day [1-1.8] after presentation. Median procedure time in the EJ group was 26.5 min, including 8.5 min [6.8-11.0] for the E-tube and 17.5 min [15-20.5] for the EJ-tube. EJ-tubes were left in place for 5 d [3.8-5.3 ]. Oral displacement of the tube was observed in 3/10 dogs. Efficiency of nutritional support was comparable between groups, with the goal to feed 66% of the requirements on d2 reached in 18/20 dogs (10 EJ, 8 E); and 100% on d3 in 16/20 dogs (9 EJ, 7 E). Relative loss of body weight during hospitalization was 6.1% [0.2-12.3] with no difference between groups.

This new technique of esophago-jejunal feeding therefore can be recommended as a convenient method for early nutritional support in dogs with severe uremia. Canine chronic kidney disease (CKD) is estimated to have a prevalence of 0.5-7%, and improved methods for the detection and monitoring of CKD are needed. Neutrophil gelatinase-associated lipocalin (NGAL), a protein detectable in blood and urine that increases secondary to renal dysfunction, is gaining utility as a renal disease biomarker in humans. The purpose of this study was to investigate serum and urine NGAL concentrations in normal dogs and dogs with naturally occurring CKD.

Forty-eight dogs assessed to be free of urinary tract disease (normal history, physical examination, clinicopathologic results, and blood pressure measurement) and 9 dogs with CKD IRIS stage I-III were enrolled in the study. Serum and urine NGAL concentrations were measured using a commercially available canine-specific ELISA kit (BioPorto Diagnostics). Creatinine and NGAL concentrations were compared using a Wilcoxon ranksum test.

The mean creatinine concentration of normal dogs [85.9 lmol/ L; standard deviation (SD) 21.1] was lower than that of CKD dogs [204.3 lmol/L; SD 76.6 (p < 0.001)]. Ranges of urine NGAL concentrations overlapped between normal (4-66 pg/ml) and CKD (4-400 pg/ml) dogs. However, mean urine NGAL concentration of normal dogs (11.9 pg/ml; SD 14] was lower than that of CKD dogs [172.4 pg/ml; SD 179.8 (p < 0.001)]. Similarly, the ranges of serum NGAL concentrations overlapped between normal (12-250 pg/ml) and CKD (31-400 pg/ml) dogs. However, mean serum NGAL concentration of normal dogs (97.2 pg/ml; SD 57.6) was lower than CKD dogs [223.8 pg/ml; SD 138.7 (p < 0.001)]. NGAL may be a useful biomarker for detecting and monitoring CKD in dogs. We are currently investigating Symmetrical Dimethyl Arginine (SDMA), a dimethylated derivative of arginine, as a potential marker for canine and feline Chronic Kidney Disease (CKD). Posttranslational modification of Protein arginine groups results in proteins with methylated-arginines. SDMA is one of the dimethylated derivatives of arginine and is released into cytoplasm after proteolysis. Circulating SDMA is mostly eliminated by renal excretion suggesting SDMA could be a potential marker for Chronic Kidney Disease (CKD). As part of our investigations, we have studied the stability of SDMA in canine and feline whole blood, serum and plasma in various types of sample collection tubes over a period of 7 days. SDMA was determined using a fully validated LC/MS method and API 4000 triple quadrupole mass spectrometer (Applied Biosystems/MDS Sciex) operating in Multiple Reaction Monitoring (MRM) mode with positive electrospray interface.

Serum was evaluated from red top tubes and serum separator tubes (SST). Plasma was evaluated from tubes containing EDTA, lithium heparin and sodium citrate whereas whole blood was evaluated from tubes containing EDTA. Three different temperatures (0 C, 4 C, room temperature) and 3 different concentrations (12, 35 and 100 u gr/dl) were studied to include the effect of freeze thaw on the SDMA.

The results suggested that SDMA is stable in whole blood, serum and plasma at 4C and room temperature up to 7 days. There are no noticeable differences between collection tubes, serum and plasma. While freezing either serum or plasma is acceptable, freezing whole blood is not recommended. 

phosphate in healthy cats, and in laboratory cats with induced renal failure. The objectives of this pilot study were to evaluate efficacy of LC in cats with naturally-occurring CKD, and describe any drug-associated adverse events.

Signalment, LC dose and administration frequency, use of other phosphate binders, biochemical analysis, and LC-associated adverse effects were recorded for 9 LC-treated cats from 3 referral hospitals. Cats were excluded if biochemical analyses had not been performed 0-7d prior to and 50-100d after start of LC therapy.

Median LC dosage (4 cats, AlOH+LC; 5 cats, LC alone) was 95 mg/kg/day (range, 32-126) Median (range) creatinine, phosphate and calcium-phosphate product (CaxP) were: Inappetance, the only adverse effect in >1 cat, could not be definitively associated with LC treatment versus progressive CKD.

This pilot study confirms that LC reduces serum phosphate when used alone or in conjunction with aluminum hydroxide. Given the absence of identifiable adverse effects, prospective clinical trials evaluating efficacy of LC monotherapy for control of CKD-associated hyperphosphatemia is warranted. Ingestion of plants of the genus Lilium is often associated with the onset of acute kidney injury (AKI) in cats, with reported mortality rates as high as 50-100%. Survival in cats with lily toxicity has been strongly correlated with early intervention, rapid GI decontamination and the initiation of aggressive renoprotective supportive care. The aim of the present study was to assess the survival of 30 cats, presented for AKI secondary to lily ingestion. Secondary objectives were to evaluate hydration status, plasma creatinine, BUN, serum calcium, phosphorus, potassium, Hct and urinary production according to patient outcome.

Medical records of 30 cats with AKI secondary to lily ingestion referred to the UC Veterinary Medical Center -San Diego and the Veterinary Specialty Hospital of San Diego between 2006 and 2012 were reviewed. Patients were divided into 2 groups; survived (Group 1, n = 22) and died (Group 2, n = 8). Eighteen of the 30 cats received aggressive medical management, while 12 cats were also managed with intermittent hemodialysis (IHD). At the time of presentation, cats in Group 2 had a significantly higher serum concentration of creatinine (p = 0.005), BUN (p = 0.002), potassium (p = 0.003) and phosphorus (p = 0.0006) and a significantly lower hematocrit (p = 0.001) and concentration of serum calcium (p = 0.04) compared to patients of Group 1. Hydration status and urine production at the time of presentation were not statistically associated with outcome. Contingency analysis (p < 0.05) revealed that cats with a creatinine greater than 2.0 mg/dl at the time of presentation were significantly (p = 0.009) more likely to die than those with a creatinine less than or equal to 2.0 mg/dl. The survival rate of patients receiving medical management was 88.8% while in patients requiring IHD patients was 50%. The overall survival rate was 73.3%.

The present study of AKI secondary to lily toxicosis showed an overall survival rate significantly higher than previously reported. Azotemia, hyperphosphatemia, hypocalcemia and anemia at the time of presentation were related to a worse prognosis and outcome. Cats presenting with AKI stages 1 and 2 were more likely to survive to discharge than those presenting in stages 3 through 5. The results of this study show that the overall prognosis for feline patients with AKI secondary to lily ingestion is good, especially in cases where early, aggressive medical management is initiated. In humans and dogs, serum Cystatin C (sCys C) seems superior to serum creatinine (sCr) to estimate glomerular filtration rate (GFR). Moreover, urinary Cys C (uCys C) is a tubular marker. A particle-enhanced nephelometric immunoassay (PENIA) is available to measure Cys C in humans. This study aimed to validate the human PENIA to measure feline sCys C and uCys C. Additionally, a pilot study was performed to compare sCys C and uCys C between healthy cats and cats with chronic kidney disease (CKD).

Western blot analysis was used to assess cross reactivity between the polyclonal rabbit anti-human antibody from the PENIA and feline Cys C. Sensitivity, imprecision and inaccuracy were determined for feline sCys C and uCys C. Finally, sCys C and uCys C were measured in 10 healthy and 12 CKD cats.

Cross reactivity was observed. The limit of detection (LOD) was 0.049 mg/L both for serum and urine. Intra-assay coefficients of variation (CV's) were 1.3% and 0.4% and inter-assay CV's 12.5% and 4.1% in serum and urine respectively. Cats with CKD had significantly higher sCys C (mean AESD 1.42 AE 0.68 versus 0.74 AE 0.20; P = 0.007) and uCys C/urinary Cr (645.1 AE 539.4 versus < LOD; P = 0.002) concentrations compared with healthy cats.

This study showed satisfactory validation results for feline Cys C. Cats with CKD showed significantly higher sCys C and uCys C concentration compared with healthy cats. Additional studies are worthwhile to evaluate Cys C as a marker of early kidney damage in cats. Acidifying diets are hypothesized to favor calcium oxalate (CaOx) urolithiasis in cats, by increasing urinary calcium excretion. Previous studies have refuted the effect of urine acidification on CaOx relative supersaturation (RSS), but compared diets differing in nutrient contents, and in small numbers of animals. The goal of this study was to test the effect of urinary pH on urinary calcium excretion and calcium oxalate RSS in healthy cats.

Thirteen adult cats (7 males, 6 females) were fed sequentially four maintenance dry expanded diets for 9 days, followed by 5 days of individual urine collection. The diets contained identical ingredients and nutrient content except for sulfur, as sodium bisulfate substituted sodium chloride for gradual acidification. PH and Ca, Mg, Na, ammonium, phosphate, citrate, sulfate, oxalate and urate concentrations were measured in each urine pool. 24-hour excretions (lmol/kg/day) were calculated from urine volumes. CaOx RSS was calculated using SUPERSAT TM .

The mixed procedure of SAS was used to assess the effect of diet (fixed effect in 4 levels), including cat as a random term. For each parameter, residuals of the model were normally distributed. Data are expressed as Least Square MeansAESE. Significance level was set at 0.05.

Diets induced urine pH of 5.93 AE 0.03, 6.02 AE 0.03, 6.23 AE 0.03 and 6.37 AE 0.04. Calcium excretion increased linearly with decreasing pH (P < 0.0001), while oxalate and citrate excretions decreased (P < 0.001). CaOx RSS was not affected by urinary pH (P = 0.63).

This study confirms the absence of effect of urinary pH on CaOx RSS for pH ranges commonly encountered in feline maintenance diets. Efficacy of nitrofurantoin (NFN) for treatment of uncomplicated urinary tract infections (UTI) in people is >70%. In dogs and cats, however, NFN is preferentially used for treatment of resistant UTI, with an anecdotally high prevalence of adverse effects (AE). Study objectives were to determine NFN efficacy for treatment of UTI, and describe NFN-associated AE.

Animals prescribed NFN were retrospectively identified. Inclusion criteria for determining NFN efficacy included cultureconfirmed susceptible UTI prior to NFN start, and repeat culture within 45d of NFN discontinuation. Inclusion criteria for AE description included treatment of suspected or confirmed UTI (culture confirmation not required) or for UTI prophylaxis, and ability to determine NFN dosage and days at AE onset and resolution.

NFN efficacy: 18 dogs and 2 cats were prescribed 2.9-5.2 mg/ kg NFN (median, 4.0) for 5-75d (median, 14d], q8 h (n = 18) or q12-24 h (n = 4) for E. coli (n = 15), Enterococcus, (n = 4), or concurrent E. coli/Enterococcus (n = 2) UTI. Resolution was confirmed for 8/21 (40%) UTI (6/15 E. coli; 2/4 Enterococcus). NFN AE: Peripheral neuropathy (n = 3) or gastrointestinal disturbances (n = 4) developed 3-52d (median, 7.5d) after start of NFN; total daily mg/kg dose was similar in dogs with (13.2; range, 4.1-17.1) or without (12.1; range, 3.3-18.4) AE. Effects resolved in 6 dogs 2-6d (median, 4d) after NFN reduction or discontinuation; 1 dog was euthanized 3d post onset of neuropathy.

Low NFN efficacy is likely due to preferential use for treatment of complicated UTI, and inappropriately long dosing intervals in some animals. AE consistently resolved with NFN discontinuation or dose reduction. Cystourethroscopy has enhanced the diagnosis and treatment of numerous conditions in veterinary medicine. Rigid telescopes used in female cytoscopy provide well-illuminated highly detailed images using rod lens technology. The flexible ureteroscope has been used for male urethrocytoscopy but is limited in image quality and procedural abilities due to the fiberoptic system, diminished illumination, and smaller working channel. The purpose of this study was to describe the technique, efficacy, and complications using a percutaneous perineal approach to the male urethra in order to gain access for rigid cystoscopy.

The perineal approach was performed ten times in nine dogs for ectopic ureter laser ablation of idiopathic renal hematuria sclerotherapy. The dogs were placed in dorsal recumbency. Using fluoroscopic guidance, an 18 gauge renal access needle was advanced trans-perineally into the pelvic urethra. Guidewire access was obtained and the access site dilated to accept a 16Fr peel-away sheath. A rigid cystoscope was then placed through the sheath to perform the procedures. Urinary catheters were placed following three of ten procedures for three to eighteen hours. The only identified peri-operative minor complication included urination form the perineal site approximately six hours post-operatively once in a single dog. No signs of stranguria or pollakiuria or incisional complications were identified in any of the 9 dogs post-operatively at follow-up examination or contact (range 4 to 1248 days).

The percutaneous perineal approach in male dogs for rigid cystoscopy appears to be a safe and effective means of facilitating endoscopic procedures. The study's purpose was to determine diagnostic accuracy of a compartmented culture and susceptibility plate (CCS) for detection of urinary tract infection and antimicrobial susceptibility in dogs. The CCS is an agar plate with one compartment for quantitative analysis using a chromogenic substrate allowing for bacterial identification and 5 antibiotic impregnated compartments: ampicillin, amoxicillin plus clavulanate, cephalothin, enrofloxacin, and trimethoprim-sulfamethoxazole. For assay validation (phase 1), frozen canine isolates of previously characterized bacteria (n = 62) were revitalized and tested with CCS and standard aerobic microbiological culture (SAMC). For the clinical study (Phase 2), urine samples collected for routine diagnostics from clinical canine patients (n = 147) were tested in parallel and blinded fashion with CCS and SAMC.

For phase 1 at 24 hours, 46/62 (74%) samples had growth on CCS and 100% on SAMC. For samples with growth, CCS correctly identified 45/46 (97.8%) bacteria. Bacteria with no growth were Gram-positive cocci (Staphylococcus sp. (n = 7, 100%), Enterococcus sp. (n = 7, 58%)). In phase 2, the overall sensitivity of CCS was 81% and specificity was 99%. There was 94% accuracy with 98% positive predictive value and 92% negative predictive value. Enrofloxacin and trimethoprim-sulfa susceptibilities had greatest concordance with those determined by SAMC (71% and 95.8% respectively).

In conclusion, CCS accurately excluded urinary tract infection but was less reliable for diagnosing infection, especially with Gram-positive cocci. The biggest study limitation was small sample size.

NU-33 DEVELOPMENT OF FELINE CYSTATIN C ENZYME-LINKED IMMUNOSORBENT ASSAY KIT IN PLASMA AND URINE, AND ITS CLINICAL SIGNIFICANCE. F Hoshi 1 , J Nakata 1 , C Takahashi 1 , S Chikazawa 1 , Y Hori 1 , K Kanai 1 , N Ito 1 , S Higuchi 1 , Y Kinoshita 2 , H Urita 2 , Y Yamagata 2 . 1 Kitasato University School of Veterinary Medicine, Aomori, Japan, 2 Nipro Co., Osaka, Japan.

Cystatin C (CysC) is a low molecular weight protein belonging to the cystatin superfamily. Human CysC is mainly used as an index of the glomerular filtration rate as a biomarker of kidney function. By using a human CysC measurement kit, we could measure the serum levels of CysC in dogs but not in cats. Further, the complete cDNA coding for CysC of several mammalian species has been identified, but the feline CysC (FeCysC) mRNA remains unknown. The aim of this study is to analyse the complete cDNA sequence of FeCysC, produce a recombinant FeCysC (rFeCysC), develop anti-rFeCysC monoclonal antibodies (mAbs) subsequently, construct an assay system for feline CysC, and investigate its clinical significance.

The full-length cDNA that encodes FeCysC has been cloned from feline white blood cells. The cDNA consists of 796 nucleotides and includes an open reading frame encoding a polypeptide of 146 amino acids. Next, we developed the anti-rFeCysC mAbs. The rFeCysC was expressed in Escherichia coli BL21 and used as an antigen to immunise mice. The reactivity of the mAbs to native FeCysC was examined against the urinary protein from cats with chronic kidney disease (CKD) by using western blot analysis. The 3 anti-rFeCysC mAbs, namely, 3-9G, 7-7C, and 9-12F were able to recognise native FeCysC. A sandwich enzymelinked immunosorbent assay kit was developed using a combination of 3-9G and the horseradish peroxidase labelled 7-7C.

The sensitivity of the assay kit that we developed was 0.098-50 ng/ml. The accuracy of the assay was measured by evaluating the analytical recovery of rFeCysC, which was added at a concentration of 0.001-1 ng. We succeeded in the development of the feline CysC assay kit in plasma and urine. The plasma and urine FeCysC concentration were significantly higher for the CKD cats than for the healthy cats. Therefore, we suggest the use of FeCysC as a marker protein to facilitate the evaluation of feline CKD. Urethral sphincter mechanism incompetence (USMI) is reportedly the most common cause of acquired urinary incontinence in dogs. Estrogen replacement therapy is one of the medical treatment options however estradiol (E2) esters and synthetic estrogens, which have a long nuclear receptor occupancy time, have been associated with estrogenic effects, such as mammary gland neoplasia, reproductive organ neoplasia and bone marrow suppression, in dogs. Estriol (E3), a naturally-occurring metabolite of E2, has short nuclear estrogen receptor occupancy. E3 has been shown to improve continence, through increasing urethral resistance and closure pressure, in incontinent female dogs and has a favorable safety profile at therapeutic doses in dogs (0.5 to 2 mg/day) for years and at high doses (10 mg/day) for prolonged periods (90 days).

To demonstrate the safety of E3 in ovariohysterectomized female dogs a study was conducted in accordance with requirements set forth by the US Food and Drug Administration (FDA), Center for Veterinary Medicine. The study evaluated the long-term safety of E3 administered orally at 1X, 3X, and 5X the maximum label dose, of 2 mg per dog per day, to healthy, ovariohysterectomized beagle dogs over 6 months.

Twenty-four purpose-bred, ovariohysterectomized female beagle dogs were included in a placebo-controlled study using a randomized complete block design by body weight. The dogs were administered E3 once daily at a dose of 0 mg (placebo), 2 mg, 6 mg, or 10 mg per dog per day. Physical examinations were conducted during acclimation and in weeks 3, 9, 15, 21 and 25. Laboratory examinations (hematology, clinical chemistry, and urinalysis) were assessed during acclimation and in weeks 3, 6, 9, 12, 15, 18, 21 and 25 . After euthanasia, a gross post mortem (including organ weights) was carried out. Relevant tissue samples, including bone marrow, were taken for histopathology and examined by a board-certified pathologist (AVCP). Tissue samples for the placebo and 5X groups were examined and, if indicated based on differences in microscopic lesions between these groups, samples from the 3X and 1X groups.

The most common effects seen following 6 months of administration of E3 at doses of up to 5X the recommended daily label dose were short term estrogenic effects, such as redness and swelling of the vulva, vulvar discharge, teat enlargement. Gross and microscopic changes in the vulva (beginning in week 1 at 6 mg or 10 mg/day or week 2 at 2 mg/day) were also noted. Vulvar discharge and teat enlargement were observed mainly during the second half of the study. No signs of stump pyometra or neoplasia or abnormalities in myeloid cells were reported.

The most common observations were short term estrogenic effects most likely associated with the pharmacologic effects of E3. E3 (Incurin tablets) is safe when administered orally on a daily basis to ovariohysterectomized female dogs at doses up to 5X the maximum recommended label dose for the treatment of estrogen-responsive USMI of 2 mg/day. Eicosanoids are potent lipid mediators involved in numerous physiologic and pathologic processes including orchestrating many aspects of the inflammatory response. Eicosanoids are synthesized from the metabolism of polyunsaturated fatty acids through the cyclooxygenase (COX) and lipoxygenase (LOX) pathways to produce prostaglandins (PG), thromboxanes (TX), leukotrienes (LT), lipoxins (LX), maresins, resolvins and protectins. Recent studies suggest that urinary bladder eicosanoid metabolism may be altered in cats with chronic feline idiopathic cystitis (FIC). The purpose of this study was to describe eicosanoid metabolites in urinary bladder tissue of healthy cats to establish normal baseline profiles.

Bladder tissues were collected from 4 male and 3 female healthy cats, aged 0.5 to 8 years, euthanized as part of unrelated studies. Only cats with a stable dietary history and normal urinalysis, urine culture, and bladder histopathology results were evaluated. Tissues were collected immediately after euthanasia and frozen in liquid nitrogen. Concentrations of lipid mediators in bladder wall tissue specimens were determined by liquid chromatography-mass spectroscopy (LC/MS).

Mean (AESD) tissue concentrations (pg metabolite/mg tissue) of mediators in whole bladder were as follows: PGD 2 = 2.9 AE 1.3; PGE 2 = 2.7 AE 3.3; PGF 1a = 35.0 AE 19.2; PGF 2a = 3.0 AE 1.7; 8-iso-PGF 2 = 9.3 AE 8.5; TXB 2 = 9.1 AE 6.0; HETE-5(S)= 2.5 AE 1.2; HETE-11(S)= 3.5 AE 2.9; HETE-12(S)= 3.4 AE 3.3; HETE-15(S)= 10.7 AE 6.8; 5-OxoETE= 1.7 AE 2.3; 15-OxoETE= 2.6 AE 2.0; LTB 4 = 10.9 AE 24.3; LTD 4 = 35.3 AE 22.4; LXA 4 = 12.9 AE 7.7; HODE-9(S)= 41.4 AE 19.9; HODE-13(S)= 33.6 AE 16.4; 9-OxoODE= 97.0 AE 73.3; 13-OxoODE= 43.5 AE 20.1; maresin-1 = 3.8 AE 6.7; protectin= 77.6 AE 82.0; resolvin-1 = 2.6 AE 2.4; resolvin-2 = none detected. Lipid mediator distribution did not differ significantly by tissue microenvironment (mucosa/submucosa vs. detrusor vs. whole bladder).

Both pro-and anti-inflammatory lipid mediators, and mediators associated with lipid peroxidation and oxidative stress were readily detected and quantified in urinary bladder tissues of healthy cats by LC/MS. This is the first report documenting expression of eicosanoids with known anti-inflammatory properties in feline bladder tissues. The influences of age, sex, disease states, and dietary or pharmacologic interventions on bladder eicosanoid expression require further investigations. Eicosanoids are potent lipid mediators involved in numerous physiologic and pathologic processes. Eicosanoids are synthesized from polyunsaturated fatty acids through the cyclooxygenase (COX) and lipoxygenase (LOX) pathways. Recent studies suggest that urothelial eicosanoid metabolism may be altered in cats with chronic idiopathic cystitis (FIC) and humans with interstitial cystitis. The objective of this study was to characterize urothelial expression of key components of eicosanoid biosynthesis in healthy cats and cats with FIC or urolithiasis.

Urinary bladder tissues were obtained by cystotomy from 8 healthy cats, 6 symptomatic non-obstructed untreated cats with chronic FIC, and 3 cats with urolithiasis. Specimens were fixed with 10% formalin, embedded in paraffin, and immunohistochemically stained for COX-1, COX-2, and 5-LOX.

Strong COX-1 and 5-LOX immunoreactivity was observed in cell nuclei in all urothelial cell layers in specimens from normal cats. The most uniform COX-1 and 5-LOX immunoreactivity tended to localize to apical urothelial cells. Similar patterns of COX-1 and 5-LOX immunoreactivity were observed in specimens from affected cats. No appreciable urothelial COX-2 immunoreactivity was observed in healthy or affected cats. Suburothelial infiltrates of COX-2-positive cells associated with urothelial ulceration were observed in one FIC and one urolithiasis cat. This is the first report describing COX-1, COX-2, and 5-LOX immunoreactivity in urothelium of healthy cats and cats with FIC or urolithiasis. Contrary to some models of cystitis in other species, we did not observe urothelial COX-2 expression in FIC or urolith-induced cystitis. Urothelial eicosanoid metabolism in feline health and disease requires further investigations. Cystocentesis is considered as the most appropriate procedure for collecting urine in cats. The aim of this study was to perform a survey to evaluate this professional practice among general practitioners (GPs).

A multiple-choice questionnaire was sent by mail to all the French small animal practitioners (n = 8198). Forty-two experts (academics, board-certified industries and clinical laboratories) were also asked to give their opinion about the ability of GPs to perform cystocentesis.

The percentage of replies received from GPs was 27.2% (n = 2228) and from experts 85.7% (n = 36). The frequency of sampling by cystocentesis (without ultrasonography) in the awake cat was: never (30.7%), sometimes (42.4%), often (22.8%), always (2.8%), no answer (1.3%). Cystocentesis was considered as the reference method of urine sampling by 67% of GPs. According to GPs, the indications for cystocentesis were urinalysis (44.3%), bacteriology (72.9%), and decompression of the bladder (62.1%). Cystocentesis without ultrasonography was considered as very easy, easy, rather difficult, and impracticable by 12.3, 48.2, 25.6 and 2.5% of GPs, respectively. In contrast, the experts considered that cystocentesis was difficult or impracticable for all GPs. 41.9% of GPs indicated that they would be interested in attending a specific course on cystocentesis.

In conclusion, there is a current need for education in cystocentesis. Moreover, this study illustrates the importance of carrying out large surveys to evaluate professional practices. Cats are considered to concentrate urine most of the time. Consequently, many internists recommend that cats with Urine Specific Gravity (USG) 1.035 be evaluated for potentially inappropriate concentrating ability. Additionally, factors such as diet are thought to affect USG. However, no data exist examining the prevalence of USG 1.035, or causes of USG 1.035 in the general cat population.

We enrolled 595 clinicians in general practice to prospectively measure USG from apparently cats presenting for annual evaluations or elective procedures. 111 practitioners provided data on 656 cats. Patient data included age, breed, visit reason, fasting status, sampling time, gender, diet, lifestyle and perceived water intake. These variables, and selected interactions, were included in a general linear model and a logistic model to examine effects on USG. Descriptive summary statistics were examined.

Cats ranged in age from 12 weeks to 21 years. USG ranged from 1.015 to 1.091. 78/657 cats had USG 1.035; a cause was identified in 20/78 cats. USG declined with age. USG 1.035 was more likely in cats presenting for elective procedures, and for females fed mostly or exclusively canned food. No associations with USG were found for fasting, sampling time or lifestyle.

Our data support the commonly held wisdom that most apparently healthy cats concentrate their urine to a USG>1.035. Approximately 25% of apparently healthy cats failing to concentrate their urine have identifiable disease, suggesting that USG 1.035 warrants additional evaluation. Common modifications to diet and lifestyle that attempt to decrease USG in cats require re-evaluation, as they appear not to affect USG. A diagnostic test for feline idiopathic cystitis (FIC) is lacking. This study evaluated the diagnostic use of a test based on infrared microspectroscopy (IRMS) of blood collected using bloodspot cards. Blood from 22 healthy cats (age 7.0 [6.0-7.0] (median [range]) years, bodyweight (BW) 6.2 [3.7-9.0] kg) and 20 cats with FIC (age 9.9 [2.9-12.2] years, BW 5.3 [3.4-8.6] kg) from the same colony, was applied to bloodspot cards (Whatman TM 903) and left to dry. A 3 mm punched circle was placed in a buffer solution and centrifuged, after which the filtrate was concentrated, applied onto a microarray slide and duplicate spectra obtained using IRMS. 1 Spectra were analyzed with principle component analysis (PCA).

Healthy cats were significantly younger (P = 0.0003) though did not differ in BW (P = 0.15) compared to FIC cats. PCA revealed statistically significant (P < 0.05) difference between groups. The separation was predominantly attributable to the band at 1546 cm À1 , which is related to metabolites of tryptophan.

This study confirmed differentiation between healthy and FIC cats based on specific infrared spectra using samples collected on bloodspot cards. Further evaluation is needed to understand the potential role of tryptophan metabolites in cats with FIC. Urethral obstruction (UO) is a common emergency of male cats. Some clinicians advocate bladder flushing at the time of urinary catheter placement, while others do not. The purpose of this prospective randomized study was to evaluate the effect of bladder flushing on short-term outcome.

Male cats presenting to the emergency service with a UO were eligible for study inclusion. Partial biochemical profile, venous blood gas, PCV and TS were performed at presentation. After relief of the UO, and with a urinary catheter in place, the bladder was either flushed with up to 500 mL of sterile saline (flush), or the catheter was directly attached to a closed collection system without flushing (no flush). Cats with urinary stones were excluded from further evaluation. Duration of catheterization, incidence of in-hospital re-obstruction and length of stay (LOS) were compared between groups.

One hundred and forty-seven cats were enrolled (n = 70 flush, n = 77 no flush). Twenty were excluded from analysis due to documented urolithiasis. Azotemia was identified in 82/127 and hyperkalemia in 56/127 at presentation. Crystalluria was present in 59/106 cats that had urinalysis performed; all but one had magnesium triple phosphate crystals. The median duration of catheterization was 36 hours (range, 0 -172 hours). UO recurred in 27/127 cats (21%) during hospitalization (11/62 flush vs. 16/65 no-flush P = 0.466 95% CI). The median LOS was 3 days and did not differ between groups.

Urinary bladder flushing at time of initial urinary catheterization does not appear to influence duration of catheterization, inhospital recurrence of UO or LOS. Up to 50% of the cats between 5 and 10 years of age in developed countries are either overweight or obese. Traditional intervention to treat obesity in cats is to put the cats on chronic caloric restriction (CCR). Cats respond to CCR by reducing their resting energy expenditure, which can slow down weight loss and predispose the cats to rebound after weight loss. The objective of this study is to compare the effects of CCR and intermittent caloric restriction (ICR) on weight loss, body fat loss, and lean body mass loss in overweight cats.

After the baseline maintenance energy requirement (MER) and body composition for each cat were determined, twenty eight overweight cats were randomized into two groups based on baseline MERs, body weight, and% body fat. The cats in the CCR group were fed 75% of their MERs for 6 months, while the cats in the ICR were fed 75% of their MERs during the first 2 weeks and 100% of their MERs during the 2 nd 2 weeks of the months for 6 months. Daily food intake, weekly body weight, and monthly quantitative magnetic resonance for body composition were performed during the trial. The results show that the ICR was as effective as the CCR in promoting weight loss and body fat loss except that the cats in the ICR group lost significantly (p = 0.0002) more body fat than the cats in the CCR group (4.33 AE 0.36% vs 2.49 AE 0.32%) at the end of the first month of weight loss. Body condition scores correlate well with body fat mass measured by dual-energy X-ray absorptiometry (DEXA), but reliability varies amongst observers. If BCS could be indirectly estimated from photographs (iBCS), an online system could be developed, circumventing problems with observer reliability. The current study determined the feasibility of using photographs to estimate iBCS.

Pre-and post-weight loss photographs from overweight and obese dogs referred to the Royal Canin Weight Management Clinic, University of Liverpool UK, were used. Twelve observers, with a range of experience, examined the photographs and estimated body condition (iBCS) using three methods: (1) a zoometric assessment of a dorsal photograph to determine the ratio of thoracic width to abdominal width (iBCS meas ); (2) a subjective estimate of body condition from a photograph with no standard-ized positioning (iBCS sub ): and an amalgamation of the two, whereby the score assigned from the T:A ratio was modified, if necessary, after subjectively reviewing standardized dorsal and lateral photographs (iBCS adj ).

Moderate associations with body fat, measured by dual-energy X-ray absorptiometry, were seen with all methods (median R s of the 3 methods: 0.64-0.74; P < 0.001), and all methods agreed moderately well with actual BCS (median kappa of the 3 methods: 0.55-0.70; P < 0.001). Age, sex, breed, coat length and coat color had no significant effect on iBCS calculated by any method. However, scores assigned by lay operators were less reliable than those with veterinary training (P = 0.035), whereas reliability was best for dogs with an actual BCS of 5/5, and worst for dogs with an actual BCS of 4/5 (P < 0.001).

These findings suggest that an estimate of body condition can be obtained from an indirect assessment of photographs, although reliability depends upon experience, and is worse for dogs in overweight condition. Obesity is the most common nutritional disorder of pet dogs (prevalence of 23-59%). This 31-week study investigated a nutraceutical containing 1 g leucine and 13 mg pyridoxine designed to maintain lean muscle mass while decreasing adiposity when compared with positive and negative controls. After determining individual maintenance calorie requirements over 4 weeks, eighteen healthy adult Beagles were fed a canned adult diet (CAD) in calories sufficient to maintain a lean body weight with excess calories supplied by oil and kitten food to induce a body fat of at least 30% over 15 weeks. During a weight loss phase, excess calories were removed, and dogs were followed for 12 weeks on nutraceutical with CAD (N), placebo with CAD (P), or a therapeutic weight loss diet (WLD). During the weight loss phase, ANOVA revealed no significant difference in food intake (kcal/ BW kg ) between groups. The N group lost a higher percentage of their body weight (21.0 vs. 6.9%, p = 0.085) and lost more body fat compared to P (12.1 vs. 2.6%, p < 0.001), but did not differ from WLD. Change in lean mass did not differ between N and WLD (11.4 vs. 15.0%), but both lost more lean mass compared to P (2.4%, p < 0.001). These data show dogs eating maintenance levels of CAD performed better when receiving N when compared with P. Without changing to WLD or restricting calories below lean maintenance levels, N dogs were able to lose similar amounts of weight and body fat as WLD dogs while maintaining similar lean mass. Primary hyperlipidemia in Miniature Schnauzers (MS) is a common metabolic disorder of unknown etiology. Feeding a lowfat diet is commonly recommended as the initial approach for the management of this condition. There are several commercial diets that are being marketed as low-fat, but their effect on lipoprotein profiles in dogs with hyperlipidemia has not been evaluated. Therefore, the aim of the present study was to evaluate the effect of a commercially available low-fat diet on serum lipoprotein profiles in MS with primary hyperlipidemia.

Sixteen MS with primary hyperlipidemia and 28 healthy MS were enrolled. Each of the MS with hyperlipidemia had a total of 4 blood samples collected. Two samples were collected while the dog was on its regular diet; these samples were used to diagnose and confirm the presence of hypertriglyceridemia. After the collection of the 2 initial samples, the dogs' diet was changed to a dry low-fat diet (Royal Canin Gastrointestinal TM Low Fat TM ) and two additional samples were collected at approximately 8 and 12 weeks after the diet change. Lipoprotein profiling was carried out using a bismuth sodium ethylenediaminetetraacetic acid (NaBi(EDTA)) density gradient ultracentrifugation method as previously described. Sliced inverse regression (SIR) and logistic regression analyses were used to test whether there is a relationship between group and lipoprotein profile. Significance was set at p < 0.05 for all analyses.

Before the diet change, 15/16 (94%) of hyperlipidemic MS were classified as hyperlipidemic based on their lipoprotein profile alone. After the diet change, significantly fewer Miniature Schnauzers (7/16; 44%; odds ratio: 19.3; 95% CI: 2.0-184.0; p = 0.006) remained classified as hyperlipidemic by lipoprotein profile analysis, while the majority (56%) were classified as normal. The most important changes in lipoprotein fractions in response to diet change as determined by SIR analysis were decreases in triglyceride-rich lipoproteins and low-density lipoprotein (LDL)-1 in combination with increases in LDL-4 and high-density lipoprotein (HDL)-3c. Dogs that eventually responded to the diet change could be separated with 88% accuracy from the ones that did not respond based on lipoprotein profile analysis before the diet change (chi-square=8.99; p = 0.003). Dogs that did not respond to the diet change tended to have initially lower LDL fractions and higher HDL fractions than the ones that responded.

In conclusion, the study diet was found to significantly shift lipoprotein profiles of hyperlipidemic MS towards normality. A subgroup of dogs did not fully respond to the diet change. Dogs that eventually responded to the diet could be separated from those that did not based on lipoprotein profile analysis before the diet change. Dietary factors have been implicated in the etiology of feline urolithiasis. Main dietary changes to reduce the recurrence of calcium oxalate (CaOx) and struvite calculi are based on increasing urine volume (high sodium diet, addition of water to the diet, canned food) or decreasing precipitation risk of the lithogenic components in the urine (modifying pH of the urine, decreasing lithogenic components or including crystallization inhibitors). Water composition can be very variable depending on the origin and quality. The purpose of this work was to study whether water mineral composition and hardness can affect cat urine composition.

Twelve adult healthy cats were used for this study. In a crossover design, during 2 weeks, cats received 2 different commercial natural spring waters: Water A (Ca:4 mg/dl, Mg:1.8 mg/dl, Na:1.3 mg/dl, Cl:0.67 mg/dl, HCO 3 :18 mg/dl) or Water B (Ca:100 mg/dl, Mg:38 mg/dl, Na:35 mg/dl, Cl:58 mg/dl, HCO 3 :306 mg/dl). During the whole study, all the cats received a commercial balanced diet during. Daily food and water intake were recorded. Urine samples were taken at day 11 after the beginning of the study and density, volume, pH of the urine were analyzed for each cat. Urine concentration of minerals (Ca, Mg, P, Na, K, SO 4 2-, NH4 + , Cl -), crystal inhibitors (citrate, pyrophosphate (PP), phytate, glycosaminoglycans), oxalate, uric acid and creatinine were analyzed in pools of 2 cat urines. Urine relative supersaturation was determined for CaOx (RSS-COM) and struvite (RSS-STR) using Equil-92 program. A general linear model (GLM) with repeated measures was performed to assess the effect of water type in all variables analyzed. Values were considered as statistically significant for P value < 0.05.

Water consumption was slightly higher for water A compared to water B (102 AE 4 ml vs 97 AE 4 ml) but no differences were observed on pH, density and urine volume between both water groups. Urinary Ca, Na, K and SO 4 2content was higher with water B consumption (in mg/L: Ca:36 AE 8 vs 57 AE 11, Na:3502 AE 162 vs 3875 AE 238, K:5289 AE 193 vs 5674 AE 203, SO 4 2-:7977 AE 193 vs 8623 AE 420) while PP was higher with water A (14 AE 1 mg/L vs 12 AE 0.8 mg/L). RSS-COM showed higher urinary values with water B cats when compared to water A (7 AE 1 vs 14 AE 4), while no differences on RSS-STR were showed.

In conclusion, mineral composition of water can have a significant effect on urine composition of cats and should be considered as a dietary factor that could modify the risk of urolithiasis in cats. The objective of this study was to document the changes in body composition of overweight/obese dogs fed a new food moderate in calories, fat and protein, designed for weight loss and weight maintenance. Twenty beagle dogs were determined to be at least 20% over their individual calculated ideal body weight as determined by dual x-ray absorptiometry (DXA). During the weight loss phase of the study, dogs were offered a once daily ration of the test food based on each individual's energy requirements set to achieve weight loss of 1-2% body weight/week. Dogs were placed on the weight maintenance protocol once they achieved target body weight (15-20% of their body composition as fat) or at the end of 4 months. The weight maintenance phase lasted an additional 4 months. Food offerings were adjusted as needed to maintain the body weight achieved at the end of the weight loss period. Monthly DXA was performed to determine body composition changes during weight loss and weight maintenance. Changes in body weight, percent body fat and lean mass at each time point were compared to baseline (month 0 for weight loss phase and month 4 for weight maintenance phase) using a paired t-test. During the weight loss phase, average body weight decreased from 17.6 AE 0.71 to 13.6 AE 0.52 kg (P < 0.01). Average percent body fat decreased from 41.0 AE 1.01 to 26.0 AE 1.5 (P < 0.01). Lean body mass decreased from 9.82 AE 0.33 to 9.53 AE 0.30 kg (P = 0.04). During the weight maintenance phase, average total body weight decreased slightly from 13.6 AE 0.52 to 13.4 AE 0.50 kg; this change was not significant. Average percent body fat decreased from 26.0 AE 1.5 to 23.0 AE 1.6 (P = 0.03). Average lean body mass increased from 9.53 AE 0.30 to 9.78 AE 0.35 kg (P = 0.04). Serum biochemical values remained normal during weight loss and maintenance. Based on this data, the test food provided safe and effective weight loss and maintenance. Lean body mass increased during the weight maintenance phase in the face of continued loss of body fat. The objective of this study was to document body composition changes during weight loss followed by weight maintenance in cats fed a new food moderate in fiber, fat and protein. Twenty domestic shorthair cats were determined to be at least 20% over their individual calculated ideal body weight as determined by dual x-ray absorptiometry (DXA). During the weight loss phase of the study, cats were offered a ration of the test food based on each individual's energy requirements set to achieve weight loss of 1-2% body weight/week. Cats were placed on the weight maintenance protocol once they achieved target body weight (15-20% of their body composition as fat) or at the end of 4 months. The weight maintenance phase lasted an additional 4 months. Food offerings were adjusted as needed to maintain the body weight achieved at the end of the weight loss period. Monthly DXA was performed to determine body composition changes during weight loss and weight maintenance. Changes in body weight, percent body fat and lean mass at each time point were compared to baseline (month 0 for weight loss phase and month 4 for weight maintenance phase) using a paired t-test. During the weight loss phase, average body weight decreased from 6.60 AE 0.35 to 5.04 AE 0.27 kg (P < 0.01). Average percent body fat decreased from 40.1 AE 1.5 to 26.1 AE 1.5 (P < 0.01). Lean body mass decreased from 3.75 AE 0.16 to 3.54 AE 0.16 kg (P < 0.01). During the weight maintenance phase, average total body weight decreased slightly from 5.04 AE 0.27 to 4.88 AE 0.25 kg, (P = 0.04). Average percent body fat decreased from 25.5 AE 1.4 to 20.7 AE 1.7 (P < 0.01). Average lean body mass increased from 3.55 AE 0.17 to 3.70 AE 0.17 kg (P < 0.01). Serum biochemistry values remained in the normal range during both weight loss and maintenance. Based on this data, the test food provided safe and effective weight loss and maintenance. Lean body mass increased during the weight maintenance phase in the face of continued loss of body fat. The purpose of this study was to test the hypothesis that dog plasma concentrations of selected nutrients decrease after undergoing caloric restriction for weight loss. Thirty-one overweight dogs that had successfully lost at least 15% of initial body weight between 2005 and 2011 were included in the study. Dogs remained healthy throughout the study, and no signs attributable to nutrient deficiency were noted. Percentage weight loss was 28.3% (16.0-40.1%) starting body weight, over a period of 250 days (91-674 days). Median energy intake during the weight loss period was 62 (44 to 74) Kcal/kg 0.75 target weight per day. Nutrients that had been previously identified to be at potential risk of deficiency during caloric restriction were measured in plasma (choline, amino acids) and urine (selenium) at the initiation and completion of a standardized weight loss regimen in dogs. Choline (P = 0.046) and threonine (P = 0.02) decreased after weight loss. Glycine (P = 0.041), and urinary selenium:creatinine ratio (P = 0.006) both increased after weight loss. There were no other significant differences in plasma nutrient concentrations.

Since concentrations of most measured nutrients did not change significantly, the data are not consistent with a widespread nutrient deficiency in dogs undergoing caloric restriction using a diet formulated for weight loss. However, the change in plasma choline concentration might indicate an increased requirement during weight loss. Further prospective studies are required to establish choline reference ranges and the effect of weight loss on choline requirements in the dog. Body composition, glomerular filtration rate (GFR) and circulating metabolites were measured in 70 dogs from 5 to 15 years of age fed one of five protein concentrations (15, 16, 22, 25, and 30% ) in a one year study. The study protocol was reviewed and approved by the Institutional Animal Care and Use Committee, Hill's Pet Nutrition, Inc., Topeka, KS, USA. All dogs were provided with regular opportunities for socialization and environmental enrichment. Dogs experienced behavioral enrichment through daily interaction and play time with caretakers, and by daily opportunities to run and exercise with access to toys. Dietary protein was sufficient in all foods as shown by no response to treatment, and maintenance of albumin, hemoglobin, hematocrit and lean body mass over the one year study. Increasing dietary protein was associated with increasing GFR (P < 0.01) and circulating urea (P < 0.01) as well as decreasing creatinine and symmetrical di-methyl arginine (SDMA) concentration (P < 0.01). Total lean body mass was positively associated with circulating creatinine concentration (P < 0.01). GFR was influenced by circulating concentrations of creatinine, SDMA, the creatinine and SDMA interaction, lean body mass, and fat mass. These data show that GFR in the dog responds to protein load as influenced by dietary protein intake, is influenced by body composition, and that creatinine, SDMA, body lean, and body fat all have a predictive benefit in estimating GFR. Body composition and circulating metabolites were measured in 39 dogs from 3 to 15 years of age fed one of three foods in a six month study (Control food -Prescription Diet k/d â , Control with added fish oil, and control with soybean and coconut oil as well as no added animal fat. The study protocol was reviewed and approved by the Institutional Animal Care and Use Committee, Hill's Pet Nutrition, Inc., Topeka, KS, USA. All dogs were provided with regular opportunities for socialization and environmental enrichment. Dogs experienced behavioral enrichment through daily interaction and play time with caretakers, and by daily opportunities to run and exercise with access to toys. Dietary protein was sufficient in all foods as shown by no response to treatment, and maintenance of albumin, and lean body mass over the six month study. The amount of lean body mass was positively correlated to serum concentrations of creatinine, negatively correlated to inosine and unrelated to SDMA and urea (P < 0.05). During the feeding period all foods had reduced SDMA and urea (urea was reduced at one month on study while SDMA was reduced at 3 months through the end of the study). Inosine was increased at one month and remained elevated through the end of the study period. Kynurenic acid was not influenced by diet or time. The objective of this study was to evaluate the antiemetic properties of maropitant, acepromazine and electroacupuncture (EA) in dogs receiving morphine as an anesthetic premedication.

The study population included 222 male and female dogs of various breeds, aged from 0.60 to 10.0 years of age, and weigh-ing from 1.90 to 55.0 kg scheduled for elective surgical procedures. Dogs were randomly assigned to one of six treatment groups (37 dogs per group). Group I, II and II received saline (placebo) 0.10 mL/kg (SC), maropitant citrate 1.0 mg/kg (SC), and acepromazine maleate 0.05 mg/kg (IM), respectively, 20 minutes before morphine administration. Group IV underwent EA at acupoint PC-6 and Group V received EA at acupoints PC-6, ST-36, GB-34, BL-20 and BL-21 for 20 minutes before morphine administration. Group VI (sham) received EA at a non-acupoint for 20 minutes before morphine administration. All dogs received 0.5 mg/kg morphine (IM) after treatment. Vomiting or retching (V/R) was recorded for 20 minutes after morphine administration; signs of nausea and sedation scores were also assessed before treatment as a baseline, before morphine administration, and 10, 15 and 20 minutes after morphine administration.

The incidence of V/R and the number of V/R events were significantly lower in Group II (37.8%; 21 events) and Group III (45.9%; 38 events) compared to Group I (75.7%; 88 events) and Group VI (86.5%; 109 events) (p < 0.05). Groups IV and V had a V/R incidence of 64.8% and 70.3%, respectively, and the number of V/R events were significantly lower in Groups IV (35 events) and V (34 events) than Groups I and VI (p < 0.05). A significantly shorter duration of V/R was found in Groups II (0.06 AE 0.19 min), III (0.48 AE 1.10 min), IV (0.20 AE 0.46 min), and V (0.15 AE 0.33 min) groups compared to Groups I (1.32 AE 1.41 min) and VI (1.87 AE 2.62 min) groups (P < 0.01).

The incidence of signs of nausea was significantly lower in Groups III (8.1%), IV (18.9%) and V (10.8%) compared to the Groups I (29.7%), II (32.4%), and VI (40%) following morphine administration (p < 0.05). The severity of signs of nausea was significantly less in Groups III, IV and V compared to Groups I, II and VI (p < 0.05).

Group III had the highest sedation scores. Group IV and V also showed a marked sedative effect compared to Group I, II and VI.

In conclusion, maropitant (Group II) and acepromazine (Group III) were effective in preventing morphine-induced V/R. EA (Group IV and V) did not influence the incidence of V/R but was effective in reducing V/R events per dog. Acepromazine and EA were effective in preventing signs of nausea induced by morphine. EA appeared to have marked sedation effect after treatment. Point-of-care (POC) glucometers have been reported to provide inaccurate measurements for anemic and hemoconcentrated samples. The purpose of this study was to determine the effect of a range of hematocrits on a veterinary-marketed POC glucometer and develop a formula to correct the POC reading given a known hematocrit.

Sixty mLs of heparinized blood were obtained from 6 healthy dogs; each was processed into packed red blood cells and plasma. Packed red cells were resuspended using varying quantities of plasma to achieve packed cell volumes (PCV) from 0-94%. Duplicate readings were obtained using the POC glucometer (POCgluc); samples were then centrifuged, decanted, and frozen. Plasma samples were batch analyzed on a clinical laboratory biochemical analyzer (CPgluc).

Mean PCV and mean POCgluc readings were calculated for each dilution. POC failed to read four samples with PCV >80%. A repeated measures ANOVA was used to test for differences between POCgluc and CPgluc. Delta glucose (Dgluc) was calculated by subtracting POCgluc from CPgluc. A linear regression model was used to describe the relation between Dgluc versus PCV; a correction formula for POCgluc (CorrPOCgluc) was developed.

There was a significant difference between POCgluc and CPgluc (p < 0.0001). Mean Dgluc was 41 mg/dl with a range of -99 to +62 mg/dl. Applying the correction formula to POCgluc, the average difference between CorrPOCgluc and measured CPgluc was 5.5 mg/dl with a maximum difference of 19 mg/dl; CorrPOCgluc was more accurate than the measured value (POCgluc). The correction formula is being validated in a prospective clinical trial. Needle stick injuries (NSIs) are common among veterinary staff, and carry the risk of trauma, drug reaction or infection. Most NSIs can be prevented using basic safe handling practices. The objectives of this study were to describe sharps handling practices and the incidence of NSIs associated with routine companion animal appointments. This was done as part of a study evaluating hand hygiene behavior.

Observation of sharps handling behaviour was performed in 51 clinics for approximately 3 weeks each using 2 small wireless surveillance cameras: one in an exam room, one in a nearby backroom area. Descriptive data from 33 clinics are presented, including 1623 appointments. An approved sharps container was present and readily accessible (e.g. not in a drawer or cupboard) in the exam room in 12 (36%) clinics. Sharps use (needle or scalpel) was observed in 831 (51%) appointments, 828 of which were needles. A needle was uncapped using the mouth in 242/828 (29%), recapped by hand in 695/828 (84%) or left uncapped on a table or counter in 134/831 (16%) appointments. All visible sharps were properly disposed prior to the end of the appointment in 306/831 (37%) cases. One apparent NSI was observed, during recapping of a needle used for vaccination.

Although there is significant room for improvement in sharps handling behaviours in companion animal clinics, the incidence of observed NSIs in this study was low. Veterinary staff must be diligent regarding safe handling and disposal of sharps in order to protect themselves and others. Both dog owners and veterinarians recognize cloudy eye (nuclear sclerosis, NS) as a common problem in aging dogs. NS is similar to "senile cataracts" in humans, which may result in similar visual impairment in dogs. Previously, a 5 point lens cloudiness score (LCS) (Tobias cloudiness scale) was shown to correlate with age in dogs (Tobias AJVR 1995) . The objective of this study was to determine if this same scale could be used to monitor NS changes over time associated with aging.

Eye exams were completed by a veterinary ophthalmologist and LCS assessed by two veterinarians independently, on each eye on 18 healthy Beagles (Age Mean 6.48 AE 0.30 year; 7 neutered males and 11 spayed females); and 27 Labradors (Age Mean 8.29 AE 0.82 year; 14 neutered males and 13 spayed females). LCS was repeated every year for 5 years. Statistics were done using a repeated measures analysis of covariance with the age of the dogs when they started the study as a covariate.

Dogs showed a significant increase in LCS over time (P < 0.05). There was no breed difference for progression rate with an average of 0.33 AE 0.14 for Labradors and 0.28 AE 0.15 for Beagles per year, respectively. However, the onset of lens cloudiness was later in Beagles compared to Labradors, with an approximate two year delay.

Use of this lens cloudiness grading scale can provide a practical measure to assess and monitor NS development and progression in dogs in clinical practice.

OT-5 SERUM C-REACTIVE PROTEIN CONCENTRATIONS IN DOGS WITH IDIOPATHIC EPILEPSY. DB Calvo, SI Miyashiro, KK Kanayama, JS Garcia, SRR Lucas. School of Veterinary Medicine and Animal Science, University of São Paulo, Brazil.

C-reactive protein (CRP) is one of the most important acute phase proteins in both human and veterinary medicine and an important inflammation and infection biomarker, also assisting in the diagnosis of neurological disorders. This study aimed to measure the serum CRP concentrations in patients with idiopathic epilepsy (IE), without structural or infectious alterations, presenting generalized seizures episodes in order to verify if the seizures interfere in CRP concentrations. Therefore, 56 dogs were evaluated in three study groups. Control group, composed by 23 healthy dogs, group B with 17 dogs presenting IE which had seizures within 24 hours, and group C with 16 dogs presenting IE, which had seizures in a period of time between 24 hs and 5 days. The mean of CRP concentration was 0.98 (range: indetectable -6,36 lg/mL) for the control group, 2.14 lg/mL for group B (range: indetectable -5,03) and 0.68 lg/mL (range: indetectable -1,98 lg/mL) for group C. Statistical analysis was performed by Kruskal-Wallis followed by Dunn's test and no significant difference on CRP concentration was observed between the control group and those animals that had seizure episodes for more than 24 hours. A significant difference was observed when compared groups control and C with those that had seizures within 24 hours (p = 0.0002). Although the values of CRP concentration were within the accepted range for many authors, it was concluded that seizures generate in dogs with IE an acute phase response characterized by mild increase of serum CRP, probably caused by muscle contraction. Maximum CRP concentration occurs at approximately 24 hours after seizures, and was not observed detectable increase after this time, Inappropriate urination is a problem with serious implications for both cats and their human families. As part of a larger research project aimed at investigating behavioral and physical differences among cats presented with inappropriate urination, owners of 18 spraying and 23 toileting cats with normal control subjects from the same multi-cat households (n = 39) were recruited. Whilst large amounts of urine deposited on horizontal surfaces are typically found in cases of "toileting", "spraying" cats tend to deposit urine on vertical areas for chemical communication purposes. The case-control dyads were brought together by the owners to the veterinary hospital of the University of São Paulo, for a medical work-up (i.e. physical examination, complete blood count, biochemical profile, urinalysis and urine culture, abdominal ultrasound of the urinary system and cystoscopy where possible in female participant cats only). Medical concerns were identified with similar frequency in both the otherwise healthy recruited "sprayers" (i.e. 38.9% -cases versus 5.6%controls) and "toileters" (32.1% -cases versus 21.4%-controls) (Pearson Chi 2 test, p = 0.639). Medical alterations such as renal calculi, higher creatinine levels, although within normal reference range and "glomerulations" (detected during cystoscopy) in both "sprayers" and "toileters" were common. The link between early renal disease and different types of urinary act, particularly when urinary function does not seem to be diminished remains unknown. Likewise, the potential for renal formations (e.g. calculi within the kidney) to cause pain and as a consequence bring about a change in urination behaviour has been completely overlooked. If such conditions act either as causes or contributors to inappropriate urination and have not been detected due to medical exams being focused on the lower urinary tract or urinalysis this might explain, at least in part, the refractoriness of some cases. Since the veterinary health goal should be to provide complete care for the feline patient with a urinary behaviour problem a detailed medical examination including imaging evaluation of the upper urinary tract should be encouraged in refractory cases.

OT-7 EVALUATION OF THREE DIFFERENT POINT-OF-CARE TESTS FOR QUANTITATIVE DETECTION OF CANINE C-REACTIVE PROTEIN. AK Jasensky 1,2 , R Einspanier 1 , B Kohn 2 . 1 Institute of Veterinary-Biochemistry, Faculty of Veterinary Medicine; Freie Universit€ at Berlin, 2 Clinic of Small Animals, Faculty of Veterinary Medicine, Freie Universit€ at Berlin, Germany.

The C-reactive protein (CRP) is a major acute phase protein in human and canine patients. Recently, a number of different assays for the detection of canine CRP (cCRP) have been evaluated; a few of them are suitable for fast quantitative in-house diagnostic testing.

The objective of this study was the evaluation of three different point-of-care (POC) systems for the measurement of cCRP.

Blood samples of 23 healthy and 102 sick dogs were included in the study. Serum CRP measurements were performed using three different POC systems (TECOmedical AG; EUROLyser Diagnostica; LifeAssays). An ELISA (TECOmedical AG) validated for canine samples was used as reference method. Statistical analysis included descriptive statistics, Spearman's correlation coefficient, and comparisons of POC systems with ELISA by Bland-Altman plot. Inter-assay reliability was determined by calculation of Cronbach's Alpha.

ELISA cCRP values ranged from 0.1 to 4.7 mg/L (median 1.3) and from 0 to 282 mg/L (median 17.3) for healthy and sick dogs, respectively. The Spearman's correlation coefficient of cCRP values between the ELISA and the POC tests were 0.89 for TECOmedical, 0.85 for EUROLyser, and 0.97 for LifeAssays. Confidence intervals for means of differences between ELISA and POC systems (Bland-Altman plots) were -13.84 AE 74.58 for TECOmedical, 5.75 AE 36.48 for EUROLyser, and 12.02 AE 40.26 for LifeAssays. Inter-assay reliability was above 0.9 for all POC assays.

All POC tests were able to detect cCRP. The correlation coefficient differed between individual POC systems. Due to deviating results, serial measurements should be performed with the same POC system. While the benefits of routine wellness screening are suspected and supported by veterinarians, to the authors' knowledge, there exist no large studies evaluating this claim. The aims of this study were to evaluate biochemical and hematological profiles from clinically healthy dogs for abnormalities, investigate for trends with respect to age, breed, sex, and specific organ dysfunction, and to formulate recommendations based on the results for the most appropriate frequency of wellness testing. The authors hypothesized that the study's results would support the use of routine wellness blood testing and would be beneficial to veterinarians when counseling pet owners with regards to the utility of these tests.

Dogs presenting to 28 primary care veterinarians in Southern Ontario for routine blood screening, that were assessed as clinically healthy by both the owner and veterinarian, were included in this prospective study from January to July 2011. Data was collected from 1421 canine patients, including signalment and 16 biochemical and hematological parameters. All analyzed results were from a single major external laboratory. Statistical analyses employed t-test to assess continuous variables and proportion ratio test to assess dichotomous variables.

The study population consisted of 125 breeds, patients were 0.3 to 16 years of age (mean of 6.1 years), and no sex bias was identified. Abnormalities that were considered significant were identified within 39.5% of the study population, including hypoalbuminemia, hyperglobulinemia, elevated hepatic enzyme activity (ALT, ALP), azotemia, hyperglycemia, hyperkalemia, hypo-or hyper-calcemia, anemia, erythrocytosis, thrombocytopenia, thrombocytosis, lymphocytosis, neutropenia, and neutrophilia. Hypoalbuminemia, elevated ALT and ALP enzyme activities, elevated serum urea and creatinine concentrations, alterations in hematocrit, alterations in platelet concentration, and alterations in neutrophil concentration were statistically correlated with increasing age. In addition, the severity of the increase was statistically correlated with increasing age for ALP and ALT enzyme activity and serum creatinine concentration. Elevated ALP enzyme activity was statistically more likely to be present in females compared to males. Additionally, if elevated ALP enzyme activity was present, elevated ALT enzyme activity or thrombocytosis were, independently, likely to be present. Female patients in the study population were also more likely to have marked neutrophilia or alterations in hematocrit. The majority of statistically significant biochemical and hematological abnormalities were present in patients of a mean age of six years or greater.

This study supports the utility of wellness blood screening in the general canine population. Routine wellness testing is of particular importance in patients six years or greater to aid in the detection of occult disease. Reproduction is a risky affair; a lifespan cost of maintaining reproductive capacity, and of reproduction itself, has been demonstrated in a wide range of animal species. However, little is understood about the mechanisms underlying this relationship. Most cost of reproduction studies simply ask how reproductive capacity influences age at death, but are blind to the subjects' actual causes of death. Lifespan is a composite variable of myriad causes of death and it has not been clear whether reproductive capacity influences all causes of death equally. We compared causes of death among 41,045 sterilized and reproductively intact domestic dogs, using a previously-described pathophysiologic classification system a and stratified by age using a Cochran-Mantel-Haenzel (CMH) test. We found that sterilization increased life expectancy by 13.8% in males (v 2 = 446, P < 10 -6 ) and 26.3% in females (v 2 = 1372, P < 10 -6 ). Sterilized dogs were dramatically less likely to die of infectious disease (v 2 = 184.4, P < 10 -6 ), trauma (v 2 = 268.7, P < 10 -6 ), infectious disease (v 2 = 184.4, P < 10 -6 ), vascular disease (v 2 = 8.25, P = 0.004), and degenerative disease (v 2 = 7.7, P = 0.006). In contrast, sterilized dogs died more commonly from neoplasia (v 2 = 300.4, P < 10 -6 ) and immune-mediated disease (v 2 = 167.2, P < 10 -6 ). These findings suggest that to better understand how reproduction affects lifespan, a shift in research focus is needed. Beyond the impact of reproductive capacity on when individuals die, we must investigate its impact on why individuals die.

a The ongoing, systematic collection, analysis, and interpretation of disease-related data is essential to the planning and evaluation of small animal practice. Similar studies are available in western countries, whereas this is the first report to document such an analysis in the Middle East. The aim of this study was to analyze the demography of presented cases and owner's attitudes towards veterinary care in the area.

Records of cases admitted to the Small Animal Clinic (SAC), Veterinary Teaching Hospital (VTH) at Assiut University, between 2007 and 2009 were analyzed for the following: total number of feline and canine cases in each year, sex, age and breed. Case fatalities / year and most common presenting causes were also analyzed.

Compared to the very limited number of cases presented in 2007 (n = 14), there was a triple and double increase in the number of both feline and canine cases in the following two years (n = 48 & 138, n = 40 & 72) respectively. Case fatalities were higher in the first year compared to the following two years (7.14%, 5.68% and 2.02% respectively). The most popular cat breed was the Persian and the Angora (80%, 63% and 62%) and that for canines was the Griffon (75%, 28% and 24%) over the study period. The most common cause for presentation was general health check (GHC) for cats (21.43% -29.55%) followed by gastrointestinal problems (19%-21%) and dermatological problems (7%-32%). The most common cause for presenting dogs at the clinic was similar, however in the first year, 67% of cases were presented with dermatological problems and 33% for reproductive problems and in the following two years, the third most common cause for presentation was trauma (9%-13%). The age range over the three years for cats was 1-132 months (average= 16, median = 10), that for dogs was 0.5-132 months (average = 41, median = 8).

The increase in number of cats and dogs presented for GHC indicates an increase of owner-awareness of the importance of GHC and routine vaccinations to the health of their pets. The frequency of cases presented with gastrointestinal and dermatological causes prompted us to invest in developing investigative tools and equipment in these two specialties. Analyzing the case records yearly will help make important decisions to develop clinics across the country. While the benefits of routine wellness screening are suspected and supported by veterinarians, to the authors' knowledge, there exist no large studies evaluating this claim. The aims of this study were to evaluate biochemical and hematological profiles from clinically healthy cats for abnormalities, investigate for trends with respect to age, breed, sex, and specific organ dysfunction, and to formulate recommendations based on the results for the most appropriate frequency of wellness testing. The authors hypothesized that the study's results would support the use of routine wellness blood testing and would be beneficial to veterinarians when counseling pet owners with regards to the utility of these tests.

Cats presenting to 28 primary care veterinarians in Southern Ontario for routine blood screening, that were assessed as clinically healthy by both the owner and veterinarian, were included in this prospective study from January to July 2011. Data was collected from 277 feline patients, including signalment and 16 biochemical and hematological parameters. All analyzed results were from a single major external laboratory (Idexx Laboratories Inc, Markham, Ontario). Statistical analyses employed t-test to assess continuous variables and proportion ratio test to assess dichotomous variables.

The study population consisted of patients aged 0.3 to 19 years of age (mean of 7.8 years) and no sex bias was identified. Within the study population, 65.7% of patients had abnormalities that were considered significant, including hypoal-buminemia, hyperglobulinemia, elevated hepatic enzyme activity (ALT, ALP), azotemia, hyperglycemia, hypo-or hyper-kalemia, hypercalcemia, elevated serum thyroxine (T 4 ) concentration, anemia, erythrocytosis, thrombocytopenia, thrombocytosis, lymphopenia, lymphocytosis, neutropenia, and neutrophilia. Elevated ALT, ALP enzyme activities, elevated serum urea concentration, elevated serum creatinine concentration, elevated serum T 4 concentration, and alterations in neutrophil concentration were statistically correlated with increasing age. In addition, the severity of neutrophilia or neutropenia were statistically correlated with increasing age. If a patient had an elevated serum creatinine concentration, they were statistically more likely to have an elevated serum urea concentration. In the study population, 75% of patients with an elevation in one or both renal parameters were 11 years or older. All patients with an elevated serum T 4 concentration were 14 years or older.

This study supports the utility of wellness blood screening in the general feline population. Routine evaluation for renal disease is most important in feline patients 11 years and older. Similarly, routine evaluation of serum T 4 concentration is most important once cats reach 14 years of age and older. The Canines and Childhood Cancer (CCC) Study seeks to examine the ability of animal-assisted therapy (AAT) to impact the well-being and distress levels of pediatric patients with acute lymphoblastic leukemia (ALL) and their families, as well as the therapy dogs who visit them. Through collaboration with Pfizer Animal Health, this study addresses two crucial research gaps within the Human-Animal Interactions field:

Is the incorporation of animals into clinical settings and systems scientifically efficacious? What are the effects of these interactions on therapy dogs? This presentation will examine the findings and lessons learned from the study's pilot, ending in April 2013. The pilot study utilized a three-site, randomized control design over a three month data collection period to begin addressing the following hypotheses:

Pediatric cancer patients with ALL who receive AAT will experience less distress throughout the course of their treatment sessions than patients who do not receive AAT, as measured by the Observational Scale of Behavioral Distress, blood pressure, and heart rate variability. Parent(s)/primary caregivers of pediatric cancer patients with ALL who receive AAT will experience less distress throughout the course of their child's treatment sessions than parent(s)/primary caregivers of patients who do not receive AAT, as measured by the State Trait Anxiety Inventory, the Pediatric Inventory for Parents, and heart rate variability. Therapy dogs will exhibit minimal distress over the course of the study, as measured by handler-self reports, canine salivary cortisol and observer ratings of videotaped behavior via an AAT ethogram. The domestic cat research community has developed an illumina Infinium iSelect Feline 63K DNA array that was financially supported by Hill's Pet Food, Inc and distributed by competitive awards from the Cat Health Network. These DNA arrays are useful for case-control studies of cat diseases and traits, limiting the need for extended pedigrees for causative gene mapping to chromosomal regions. However, the successful use of DNA arrays is dependent on the density of the single nucleotide polymorphisms (SNPs) on the array, genetic make-up of the case-control samples, and the dynamics of the cat population under examination. Our research efforts have focused on the genetic structuring of cat populations, indicating that different genetic markers can define eight worldwide "races" of cats and that the most common 29 breeds form only~21 cat breed populations. In additional, our examination of genetic linkage disequilibrium demonstrates that certain breeds, such as Burmese and Birman, have high linkage disequilibrium and inbreeding, requiring less cats for powerful case-control analyses. However, breeds that have low inbreeding, low linkage disequilibrium and therefore are very similar to random bred cats, require higher samples sizes for a sufficient analysis. These low power cat breeds, such as Siberians and random bred cats, will likely need a higher density DNA array for successful studies. Our research group has examined over 2,000 cats for various diseases and traits to identify locations for these conditions on cat chromosomes and the subsequent analysis of regional candidate genes for causative mutations. We have previously reported success with the identification of Burmese hypokalaemic polymyopathy to a mutation in the WNK4 gene on cat chromosome E1. The studies have also been successful with the localization of a progressive retinal atrophy in Persians, a major gene causing brachycephaly and a craniofacial dysmorphology in Burmese cats, and several dominant and recessive coat colors and fur types, such as the Selkirk Rex mutation. This report will present several cat case-control studies and the genetic analyses of cat populations, highlighting the potential reasons for the successes and failures of case-control studies. In addition, the genome sequence of the domestic cat has reached 14x coverage. Small groups of cats, such as a cat with a disease or abnormal condition and its parents and or normal sibs, can be genetically sequenced as a means to discover causative mutations. Direct sequencing of human patients with abnormalities is rapidly becoming a standard of care. The need and feasibility of direct sequencing and higher density DNA arrays for companion animal health care will be presented, highlighting genetics of the domestic cat. Itraconazole is available in two FDA-approved formulations (Sporanox â and generic) and in compounded formulations. The purpose of this study was to determine the pharmacokinetics and relative bioavailability of these formulations in 9 healthy Beagle dogs.

After a 12 hour fast, each dog received 100 mg itraconazole (average dose: 10.5 mg/kg) of each formulation with small meal in a randomized, 3-way, 3-period, crossover design with an 8-day washout period. The 3 formulations studied were Sporanox â , an approved human generic capsule, and compounded itraconazole prepared in the college's pharmacy. Plasma was collected at predetermined intervals for analysis using high pressure liquid chromatography (HPLC). Concentration data were analyzed using non-compartmental pharmacokinetics to determine: area-underthe-curve (AUC), peak concentration (C MAX ), and terminal halflife (t1/2), and coefficients of variation (CV%). AUC, geometric mean (lgÁhr/mL) The compounded formulation produced an AUC and C MAX that were only 8.0% and 4.1% of the reference formulation (Sporanox â ), respectively. In contrast, the generic formulation produced an AUC and C MAX that were 110% and 86.3% of the reference formulation (Sporanox â ), respectively.

Using our acceptance criteria, generic itraconazole is bioequivalent to the innovator Sporanox â in dogs, but compounded itraconazole is not bioequivalent and produced such low concentrations in dogs that it is unlikely to be effective. Based on this study, compounded formulations of itraconazole cannot be recommended for dogs. The FDA-approved label for cefovecin (Convenia) warns that administration with another highly-protein bound drug, such as a non-steroidal anti-inflammatory drug (NSAID) could produce a protein displacement drug interaction. Carprofen and cefovecin are two highly protein-bound (>99% and 98.5%, respectively) drugs frequently administered to dogs. A protein displacement interaction may produce higher unbound drug concentrations, changes in pharmacokinetics, and ultimately adverse effects or lack of efficacy. The purpose of this study was to determine whether cefovecin alters the plasma pharmacokinetics of carprofen in dogs. Carprofen was administered intravenously (4 mg/kg) alone and in combination with subcutaneously administered cefovecin in a two-period crossover study utilizing six adult dogs. Plasma concentrations of carprofen enantiomers were analyzed by HPLC and pharmacokinetics were determined using compartmental methods. Cefovecin did not influence the pharmacokinetics of S(+) carprofen, but did affect the R(-) enantiomer. Area under the curve (AUC), clearance, and terminal half-life (T 1/2 ) were significantly (p < 0.05) less following concurrent carprofen and cefovecin administration when compared with carprofen alone. The R(-) enantiomer geometric means for AUC were 200.6 and 148.2 lgÁh/mL, means for Cl were 10.0 and 13.5 mL/ min/kg and means for the terminal half-life (T½) were 8.5 and 5.5 h for carprofen alone and in combination with cefovecin, respectively. Volume of distribution at steady-state was unchanged and was 0.10 L/kg for both enantiomers. Although the PK profile of the R(-) enantiomer was significantly influenced by concurrent cefovecin administration, the pharmacologically active S(+) enantiomer was not. The results of this study indicate that a clinically significant drug interaction is unlikely from the concurrent administration of carprofen and cefovecin in dogs. Cyclosporine and prednisone may be prescribed in combination for treatment of immune-mediated disorders. These medications have both been shown to have effects on insulin and glucose homeostasis. The purpose of this study was to describe the development of diabetes mellitus (DM) in dogs receiving both cyclosporine and prednisone.

Cases were obtained by searching the Texas A&M University veterinary medical database for dogs with a diagnosis of immunemediated hemolytic anemia, immune-mediated thrombocytopenia, or granulomatous meningoencephalitis from 2010-2012. Cases were included if serum glucose values were available following initial diagnosis and initiation of therapy, and if hyperglycemia developed after administration of cyclosporine and prednisone. Signalment, weight, and doses of cyclosporine and prednisone were recorded. Cases were divided into two groups-dogs diagnosed with DM, and dogs with hyperglycemia only and not DM. Dogs were diagnosed with DM if they had evidence of persistent hyperglycemia, glucosuria, and compatible clinical signs. All data was check for normality using a D'Agostino & Pearson omnibus normality test and a Mann-Whitney test was calculated to compare median drug doses and weights between dogs that developed DM and hyperglycemia only dogs.

Eighteen dogs met inclusion criteria. Of these, 4/18 (22.2%) were diagnosed with DM and required insulin to control clinical signs, while 14/18 (77.8%) were hyperglycemic only. DM dogs and dogs with hyperglycemia only had no significant different in median dose of cyclosporine (11.4 mg/kg/day vs. 10.3 mg/kg/day, respectively; P = 0.9576) or prednisone dose (3.5 mg/kg/day vs. 3.3 mg/kg/day, respectively; P = 0.7499). Dogs also had no significant difference in weight between DM and hyperglycemia only groups (6.6 kg vs. 8.3 kg, respectively; P = 0.4570). For the four dogs with DM, insulin was administered for > 1 month in all dogs, and > 6 months in 2/4 (50.0%). Only 1/4 DM dogs ultimately became non-insulin dependent.

Dogs may develop hyperglycemia when receiving combined cyclosporine and prednisone. A relatively high percentage (22.2%) of these dogs may develop DM requiring insulin therapy on either a short or long-term basis. Alimentary lymphoma comprises one of the most common malignancy in cats and hematological and biochemical profile are important to detect alterations and to select the treatment protocol. The goal of this study was to retrospectively analyse the hematologic (CBC) and biochemical (total protein, albumin, urea, creatinine, alanine aminotrasnferase, aspartate aminotransferase, alkaline phosphatase and gamma glutamyl transferase) records of 27 cats with alimentary lymphoma. A clinical history was acquired and a complete physical examination was performed for each patient at the time of the first presentation. Presenting signs included weight loss, lethargy, anorexia and diarrhea. The diagnosis was determined by cytology or histopathology. Large cell or lymphoblastic lymphomas were the most common cell type comprising 63% of the cats (17/27) and 37% presented small cell or lymphocytic lymphomas (10/17). Cats had a median age of 11 years (range 3-16 years). Regarding retroviral status, 7,4% (2/ 27) were infected by FIV and 3,7% (1/27) by FeLV. In relation to CBC, nonregenerative normocytic normochromic anemia (PCV < 30%) was observed in 26% of the cases (7/27) and leukocytosis (>19.000 leukocytes/mm 3 ) with neutrophilia (>12.000 neutrophil/mm 3 ) in 22% (6/27), associated with lymphocytosis (>7000 lymphocytes /mm 3 ) in 7,4% (2/27) and monocytosis (>1000/mm 3 ) in 15% (4/27). Anemia was present in two small and five large cell lymphomas, including the cat FeLV positive, and not related to FIV status. The main biochemical alterations were hypoalbuminemia (<2,3 g/dL) in 15% of the cats (two large cell and two small cell lymphomas -4/27) and mild increase in levels of aspartate aminotransferase (>100 U/L) in 11% (3/27). One of the cats with hypoalbumineia also showed anemia. No alterations were observed in creatinine, alkaline phosphatase and gamma glutamyl transferase. As conclusion, the most common alterations in cats with alimentary lymphoma were anemia and leukocytosis. Ondansetron is a 5-HT 3 receptor antagonist that is an effective anti-emetic in cats. It is used in the treatment of nausea and vomiting associated with chemotherapy and chronic illness such as chronic kidney disease. However, pharmacokinetic studies in cats have not previously been performed. The purpose of this study was to evaluate the pharmacokinetics of ondansetron in healthy cats.

Six healthy research cats with unremarkable complete blood count, serum biochemistry and urinalyses were utilized. Jugular catheters were placed under ketamine/butorphanol sedation 12 hours prior to sampling for ease of sample collection. Each cat received 2 mg oral (mean 0.43 mg/kg), subcutaneous (mean 0.4 mg/kg) and intravenous (mean 0.4 mg/kg) ondansetron in a cross-over manner with at least a 5 day wash out period. Serum samples were obtained prior to, and at 0. 25, 0.5, 1, 2, 4, 8, 12, 18 , and 24 hours after administration of ondansetron. Serum was separated and frozen prior to analysis. Ondansetron concentrations were measured using liquid chromatography coupled to tandem mass spectrometry. Non-compartmental pharmacokinetic modeling was performed. Repeated measures ANOVA was used to compare parameters between administration routes.

Pharmacokinetic parameters are summarized in the table below. Parameters that are significantly different (p < 0.05) are denoted with (* IV vs. PO; ** IV vs. SQ;^PO vs. SQ).

Intravenous Subcutaneous administration of ondansetron to healthy cats is more bioavailable and has a longer half-life than oral administration. This information may be helpful for management of patients with chronic disease. D-penicillamine is a chelating agent commonly used for treatment of canine copper-associated hepatitis. However, therapeutic responses are variable, and there is little information available characterizing d-penicillamine pharmacokinetics in dogs. Coadministering the drug with food to prevent vomiting is often recommended for dogs, which contradicts recommendations for people. The purpose of this study was to describe pharmacokinetics of orally administered d-penicillamine in fasted and nonfasted healthy dogs. We hypothesized that co-administration of d-penicillamine with food reduces bioavailability and maximum plasma drug concentrations (C max ).

Nine dogs received oral d-penicillamine (12.5 mg/kg) fasted and with food in a randomized, crossover design. Blood samples were collected before and 0.25, 0.5, 1, 2, 3, 4, 8, 12, and 24 hours after dosing. Plasma d-penicillamine concentrations were measured using liquid chromatography coupled with tandem quadrupole mass spectrometry. Non-compartmental pharmacokinetic modeling was performed.

Two fasted dogs (22%) vomited after receiving d-penicillamine. Mean C max +/standard deviation (SD) was 8.7 + /-3.1 lg/ml (fasted) and 1.7 + /-1.6 lg/ml (fed). Mean area under the curve +/-SD was 17.6 + /-6.0 lg/mlÁhr (fasted) and 5.5 + /-3.5 lg/mlÁhr (fed). There were significant reductions in relative bioavailability and C max in fed dogs compared to fasted dogs (P < 0.001).

Our observations indicate that co-administration of d-penicillamine with food significantly reduces plasma drug concentrations in dogs. Decreased drug exposure could result in reduced copper chelation efficacy, prolonged therapy, additional cost, and greater disease morbidity. Consequently, we recommend that d-penicillamine be administered on an empty stomach whenever possible. P-7 DETERMINATION OF SERUM SYMMETRIC DIMETHY-LARGININE REFERENCE LIMIT IN CLINICALLY HEALTHY DOGS. V Rentko 1 , M Nabity 2 , M Yerramilli 3 , E Obare 3 , M Yerramilli 3 , J Aguiar 3 , R Relford 3 . 1 Tufts University, N. Grafton MA, 2 Texas A&M University, College Station, TX, 3 IDEXX Laboratories, Inc, Westbrook, ME.

Symmetric dimethylarginine (SDMA) is a methylated amino acid derivative released during protein degradation. SDMA is almost exclusively eliminated by renal excretion. Therefore, serum SDMA may serve as an endogenous marker of glomerular filtration rate.

The objective of this study was to establish a reference limit for serum SDMA in dogs. Serum samples were collected from 151 clinically healthy dogs (based on physical examination) of varying gender, age, and breed. Data from 29 dogs were excluded on the basis of age (< 1 year old, n = 11), health status (n = 17), or immeasurable serum creatinine concentration (sCr) (n = 1). Serum SDMA concentrations were determined by LCMS (liquid chromatography mass spectroscopy) operating in MRM (Multiple Reaction Monitoring) mode using a stable isotope dilution assay.

Reference intervals were determined for both SDMA and sCr based on a non-parametric model using 2-sided 95% confidence interval. Outliers in these data were established as being outside of mean AE3 standard deviations. Two outliers were identified and removed from the reference interval calculations. The reference interval for SDMA was 6-13 ug/dL and for sCr was 0.5-1.6 mg/ dL (Figures 1, 2 , respectively). Comparisons of SDMA and sCr were made, and agreement was 98% (118 dogs) using the exclusive upper reference limits.

In conclusion, these data suggest that a reference limit for SDMA in >1 year old healthy dogs can be established at <14 ug/ dL. These data can be used in further evaluations of SDMA as a marker of renal disease in dogs. The new product 4CYTE TM (Interpath P/L, Australia) is a formulation of New Zealand Green Lipped Mussel, marine cartilage, abalone, and a proprietary extract Epiitalis â , from the plant Biota orientalis. This 4CYTE TM is intended to prevent and treat pain in dogs with osteoarthritis.

The objective of this study was to compare the efficacy of 4CYTE TM with a positive control NSAID carprofen, for the treatment of naturally occurring osteoarthritis in the dog. The primary hypothesis was that improvements in lameness scores were not different between treatments.

A randomised controlled clinical trial was used to assess response to therapy. The sample size for CI90, Power 80%, 80% responders was estimated at 40 animals per group. Eighty two dogs with clinical signs of osteoarthritis were enrolled in the study of which 16/82 (19.5%) were later excluded due to lack of radiographic evidence of osteoarthritis. The remaining 66 dogs completed the study. The main breeds represented were cross breeds 20/82 (24%), Labradors, Retrievers and Border Collies as the main pure bred cases 22/82(27%) and the remaining dogs were a variety of pure breeds. All dogs were over 6 months of age, from 10 to 50 kg and were enrolled from May 2011 to May 2012 in Victoria, Australia. The primary outcome was defined as the binomial "improvement versus no improvement" of Owner Lameness Score at Day 28 compared to Day 0 (OLS). Other outcomes were the improvement in OLS, Veterinary Lameness Score (VLS) and the Owner Mobility Scores (OMS) at Days 14 and 28. All outcomes used ordinal observation scales with masked assessment. Outcomes were evaluated for non-inferiority using the 90%CI of the difference between two independent proportions. The Hodges-Lehmann estimate of the median of the pairwise differences was also calculated from the ordinal ratings.

For dogs dosed with 4CYTE TM versus carprofen, OLS at Day 14 improved for 10/29 (34%) dogs versus 13/37 (35%) dogs, and at Day 28 had improved for 14/29 (48%) versus 13/37 (35%), respectively. The product 4CYTE TM was found to be non-inferior to carprofen at Day 14 for the OMS, and non-inferior at Day 28 for all three outcomes. For all outcomes at all time-points, the median pairwise differences were not different.

These results support the conclusion that 4CYTE TM was at least as effective as carprofen in relief of pain and lameness resulting from osteoarthritis in adult dogs, i.e. clinical end-point bioequivalence was demonstrated. Pioglitazone is a peroxisome-proliferator activated receptor gamma agonist used in humans for the management of type-II diabetes as well as in metronomic chemotherapy protocols. The primary aim of this study was to develop a simple method of measuring pioglitazone in canine plasma and then to determine the plasma concentration of orally administered pioglitazone in dogs. Blood glucose curves were also performed.

A simple and extractionless high-performance liquid chromatography method using ultraviolet detection in tandem with mass spectrometry was developed for the determination of pioglitazone in canine plasma. The method was validated with excellent sensitivity, accuracy, precision, and recovery and enabled unambiguous evaluation and quantitation of pioglitazone in canine plasma.

The method was used to quantitate the plasma levels of pioglitazone in dogs after administration of a single oral dose of approximately 0.5 mg/m 2 . Whole blood was collected immediately prior to and after oral administration pioglitazone at 0 hours, 0.5, 1, 2, 3, 4, 6, 8, 10, 12, and 24 hours.

Pharmacokinetic results revealed both significant inter-patient absorption variability and significant intra-patient absorption variability when fasted versus non-fasted plasma levels were measured. Peak serum levels were reached between 1-3 hours. Pioglitazone was completely eliminated from the plasma within 24 hours. Based on the kinetics of elimination, more frequent oral dosing may be necessary to maintain plasma concentrations. Blood glucose values remained within normal limits in these nondiabetic dogs.

In conclusion, a sensitive and selective HPLC-tandem mass spectrometry method for determination of pioglitazone in canine plasma has been developed. Pioglitazone is absorbed after oral administration in dogs and appears to be well tolerated after oral administration. The aim of this study was to compare the efficacy of three different medications in treatment of ear mite infestation accompanied by otitis externa in cats Three medications were tested for efficacy on ear mite infestation in cats using a randomized trial. Cases presented to the Small Animal Clinic, Faculty of Veterinary Medicine, Assiut University for signs of ear mite infestation and/or otitis externa were recruited and randomly assigned to one of the 3 groups for evaluation of response to treatment over a period of 30 days. Cases which did not respond to treatment within 2 weeks of treatment were changed to other medication. A simple questionnaire was designed to evaluate owner satisfaction of the medication price, ease of administration, cat tolerance and improvement of signs. Ivermectin (Noromectin â , Norbrook) was given at a dose rate of 0.2 mg/Kg of body weight in 4 weekly injections. A single topical dose of Fipronil (Frontline Combo, Merial) was applied to cat's scruff area. The third group of cats were given Canaural ear drops (Canaural â , VetXX) at a dose rate of 4-7 drops / ear BID for 21 consecutive days. All cases were either brought to the clinic re-evaluation or the owner was contacted by phone to evaluate progress of each case.

The ivermectin-treated group (n = 7), 71.43% of cases recovered from ear mite infestation based on signs resolving and disappearance of presenting signs. Only 60% of cases treated with frontline showed an improvement of signs (n = 5). All cases treated with Canaural showed complete recovery with complete absence presenting signs and signs of otitis externa (n = 5). Average age of cats in all groups ranged from two months to six years (mean= 17.33, median=6.5),.

These findings suggest that Canaural is most suitable when otitis externa is present along with ear mite infestation. Frontline Combo is not as effective as the two other medications used in this study. Golden Retriever muscular dystrophy (GRMD) is the best animal model of Duchenne's muscular dystrophy (DMD), which is the most common muscular dystrophy in humans. Up to 80% of people with DMD die of respiratory failure. Evaluation of the pulmonary system in dogs with GRMD has been limited to clinical and radiographic assessments. Further investigation into the respiratory impairment in GRMD dogs is necessary to better characterize this valuable model of DMD, and to provide practical tools for the serial monitoring of responses to novel treatments. We hypothesized that arterial blood gas analysis and tidal breathing spirometry could be reliably performed in GRMD and control dogs without sedation; and, that dogs with GRMD would have evidence of hypoventilation as evidenced by an increase in partial pressure of arterial carbon dioxide (PaCO 2 ) and decreased tidal volume (TV).

Arterial blood gas analysis and tidal breathing spirometry were performed on 11 dogs with a mild phenotype of GRMD and 11 age-matched, phenotypically normal, carriers (controls). Arterial blood was collected from the dorsal pedal artery and analyzed immediately (VetScan i-STAT, Abaxis). Spirometry was performed using an anesthetic face-mask with rubber gasket and a calibrated pneumotachograph (model 3700, Hans Rudolph, Inc.). The signal was collected with the Ponemah Physiology Platform 4.90-SP2 (Data Sciences International). Data were compared between groups using Students t-test or Mann-Whitney rank sum test, with P < 0.05 considered significant. Results are reported as median (25 th /75 th percentile).

Arterial blood gas data were successfully obtained from all 22 dogs. Spirometric data were successfully obtained from 21 of 22 dogs, resulting in the elimination of one pair of dogs from analysis. Compared with controls, dogs with GRMD had higher PaCO 2 [38.8 (35.8/41.2) mmHg; controls 35.1 (32.1/36.1) mmHg; P = 0.014], and bicarbonate concentrations [23.6 (21.3/23.9) mEq/L; controls, 19.9 (18.4/20.8) mEq/L; P < 0.001]. Dogs with GRMD also had marked elevations in peak tidal expiratory flow [PTEF; 867 (573/1086) ml/sec; controls 477 (373/691) ml/sec; P = 0.002], and PTEF:peak tidal inspiratory flow [2.01 (1.53/ 2.48); controls 1.15 (1.00/1.16); P < 0.001]. Differences were not detected for TV or inspiratory flow measurements.

Arterial blood gas analysis and spirometry were successfully performed without sedation, supporting the feasibility of using these techniques for serial monitoring of respiratory function in dogs with GRMD; and, a relative increase in PaCO2 was found in GRMD dogs. Increased PTEF has not been reported in people with DMD. This novel finding could be a consequence of hyperinflation, commonly seen on radiographs of dogs with GRMD, and deserves further study. Feline bronchial disease (FBD) is a common inflammatory disease of the lower airways. Human medicine researchers have found that obesity may play a significant role in the pathogenesis of pulmonary diseases through mechanisms that may involve proinflammatory mediators. Barometric whole-body plethysmography (BWBP) is a noninvasive pulmonary function test that allows a dynamic study of breathing patterns. The objective of this preliminary study was to evaluate if there were significant differences in tidal volume [TV] and bronchoconstriction indexes enhanced pause [Penh] and Pause [PAU] between normal weight FBD (NW-FBD) and obese FBD (O-FBD) cats by using BWBP. Forty-five client-owned cats with natural FBD were included. Cats did not have a previous history of upper airway, cardiac or systemic diseases and had negative results when tested for heartworm, FeLV and FIV diseases. Thirty-two of them were NW-FBD cats and thirteen were O-FBD cats. Obesity was defined as being 120% or more of the optimal body weight. The study was approved by the ethical committee of Veterinary Medicine Service of Las Palmas de Gran Canaria University (Spain) and it was carried out in accordance with the current European legislation on animal protection. Measured results evidenced that no significant differences were detected for any of the pulmonary function variables analyzed between NW-FBD and O-FBD cats. Statistically significant differences with P-value < 0.05

We must keep in mind some limits of this study as the high variability in bronchial involvement and to have the chance of doing bronchoreactivity tests. Although in human and canine respiratory medicine obesity is clearly associated with increased asthma severity, measured results suggest that obesity do not worse TV and bronchoconstriction indexes in natural FBD cats. Asthma is characterized by airway eosinophilia, hyperresponsiveness (AHR) and remodeling. Bronchoprovocation with methacholine (MCh) and lung biopsy remain the gold standard to assess AHR and remodeling. We hypothesized that MCh challenge during computed tomography (CT) would document AHR and remodeling in asthmatic but not healthy cats.

Healthy and experimentally asthmatic cats (n = 6/group) were anesthetized for ventilator-acquired pulmonary mechanics and CT. Baseline parameters were acquired by nebulizing saline for 30 seconds followed by four minutes of ventilator data collection. Breathholds were used to obtain inspiratory and expiratory CT scans after nebulization. Aforementioned studies were repeated using nebulized MCh (0.0625, 2 and 32 mg/ml) instead of saline. The study terminated at the dose of MCh doubling baseline resistance (EC200Raw). Lung attenuation (in Hounsfield Unit (HU)) was measured at baseline and EC200Raw using OsiriX v. 3.8.

While not significantly (p = 0.18) different, three asthmatics reached EC200Raw at low (<2 mg/ml) MCh doses; all healthy cats required the highest MCh dose to reach EC200Raw. There was a significant (p = 0.008) increase in soft tissue attenuation in the left lung field in asthmatic versus healthy cats at baseline (-826.6 AE 11.1 vs. -873.6 AE 8.6 HU, respectively) and throughout MCh nebulization. In expiration, dorsal lung field attenuation significantly increased at EC200Raw in asthmatic cats while it remained similar to baseline in healthy cats. No significant difference between the left lung volume inspiratory:expiratory ratio between groups was noted at baseline, but it significantly (p = 0.034) increased at EC200Raw in asthmatic compared to healthy cats.

Pilot findings suggest tandem pulmonary mechanics and CT imaging may detect AHR and remodeling in asthmatic cats. Pilot data suggested adipose-derived mesenchymal stem cells (aMSCs) diminished some aspects of the immune response in experimental feline asthma. We hypothesized that aMSCs would improve Bermuda grass allergen (BGA)-induced airflow limitation.

Nine "high-responder" cats (severe responses to BGA challenge) were selected from 24 cats with experimental asthma. Five cats received six i.v. infusions of aMSCs (range: 0.42-2.5X10E7 aMSCs/infusion) every two weeks; four cats received placebo. Ventilator-acquired pulmonary mechanics were performed at baseline, day 3 and week 6 using BGA bronchoprovocation (1.25-40 ug/ml); the endpoint was the BGA concentration increasing baseline airway resistance by 150% (EC150Raw). A Visual Assessment Score (VAS; 0-10 cm with 10 being severe), a visual analog scale used to grade severity of clinical signs after allergen challenge, was performed on days 0, 2, 38, 72, and 110. Data were analyzed using a two-way ANOVA with p < 0.05 considered significant. The EC150Raw was not significantly different between groups over time (p = 0.311); however, a few cats had marked variability. Although the VAS did not change significantly in either group (p = 0.163), over time it decreased in the aMSC group (VAS baseline 5.94 cm, VAS time-weighted average 4.83 cm) and increased in the placebo group (VAS baseline 4.53 cm, VAS time-weighted average 6.26 cm).

In conclusion, aMSCs do not appear to alter BGA-induced airflow limitation or clinical signs in high-responder asthmatic cats in the short term. This could be due to (1) BGA not being the ideal bronchoprovocant, (2) high-responder asthmatics being more resistant to treatment or (3) lack of efficacy. Airway hyperresponsiveness (AHR) is a key feature of asthma and can be measured using bronchoprovocation. Direct (methacholine; MCh) or indirect (adenosine-5-monophosphate; AMP or mannitol) bonchoprovocants are used in humans, the latter inducing AHR only with pre-existing airway inflammation. We hypothesized that indirect bronchoprovocants would be better at discriminating healthy from asthmatic cats than MCh.

Healthy and experimentally asthmatic cats (n = 6/group) were randomized to receive increasing doses of MCh, AMP or mannitol. Cats received all bronchoprovocants with a 1 month washout before crossover to the next bronchoprovocant. Pulmonary mechanics were measured in anesthetized and mechanically ventilated cats using an Engstrom Carestation ventilator. Saline for baseline and increasing doses of each bronchoprovocant were aerosolized for 30 seconds followed by 4 minutes of data collection between doses. The study endpoint was reached when airway resistance exceeded 200% of baseline values (EC200Raw).

There was a significant difference (p < 0.001) in the airway response of asthmatic versus healthy cats over the range of MCh concentrations despite no significant difference in the EC200Raw between groups. Response to MCh was significantly greater (p < 0.05) than in normal cats at MCh concentrations as low as 0.0625 mg/ml. For AMP, a small subset of asthmatics (2/6) responded at low concentrations; 4 asthmatics and all healthy cats failed to respond even to the highest concentrations. Likewise, mannitol was unreliable as a bronchoprovocant, with only 1 asthmatic cat and no healthy cats responding.

In conclusion, MCh discriminated asthmatic from healthy cats but neither AMP nor mannitol was an effective brochoprovocant in our model. The purpose of this study was to compare the diagnostic quality of bronchoalveolar lavage fluid (BALF) cytology acquired by manual aspiration (MA) using a handheld syringe to suction pump aspiration (SPA) via suction trap connection techniques in healthy dogs.

Bronchoalveolar lavage was performed under general anesthesia using endoscopic guidance in twelve healthy purpose bred beagles. MA was performed with a 35-mL syringe attached to the bronchoscope's biopsy channel. SPA was performed using 37.5 mmHg negative pressure applied directly to the bronchoscope's suction valve via a disposable aspiration tube. MA and SPA techniques were performed in randomized order in opposite caudal lung lobes of each dog. Two aliquots were infused at each lavage site using a weight-adjusted aliquot volume (1 mL/kg). A total nucleated cell count and differential cell count were performed on all BALF samples. A semi-quantitative scale for quality, surfactant score, and percent of retrieved fluid were also evaluated. Results were compared using a paired Student's T-test and Wilcoxon sign rank test.

The proportions of BALF retrieved (mean difference = 23.0%, p < 0.0001) and total nucleated cell counts (median difference = 100 cells/lL, p = 0.029) were significantly higher for SPA than MA.

The results suggest that SPA provided BALF samples of higher diagnostic quality than MA in healthy dogs. The SPA technique may improve the diagnostic accuracy of BAL in dogs. The incidence and significance of subsegmental pulmonary (SSP) thromboembolism is currently under investigation. In addition, clinical significance of SSP thromboembolism is also uncertain. We aimed to evaluate the clinical and diagnostic features of SSP thromboembolism in an experimental canine model.

Obstruction of pulmonary arterial branches was experimentally induced by utilizing barium-coated autologous blood clot in 3 dogs via intravenous injection using external jugular vein. The clinical signs, hemodynamic changes (blood pressue, electrocardiogram, echocardiography), coagulation (aPTT, PT, FDPs and D-dimer test) and cytokine variation (TNF-a, IL-4, IL-6, and IL-10) were evaluate over a 24-hour period (baseline, 3, 6, 12 and 24 hours after the autologous blood clot injection). Multidetector computed tomography (MDCT) was conducted to evaluate the pulmonary obstruction and histopathologic confirmation was made.

Pulmonary artery pressure gradient (PAPG) were increased 12 hours after the autologous blood clot injection (14.2 AE 2.8 mmHg to 23.6 AE 1.7 mmHg, p = 0.003) and normalized 24 hours later (P < 0.01). However, no abnormalities other than a PAPG were detected including D-dimer value. Among the cytokines, proinflammatory cytokine IL-6 expression was elevated and others were decreased. Infused radio-opaque clots were confirmed with MDCT and histopathologic examination. Pulmonary parenchymal changes such as arterial dilation and inflammatory reactions were also confirmed in histopathologic examinations, and were barely observable in MDCT.

In this experimental study, we made radio-opaque small emboli and induced SSP thromboembolism. Thus, we infer that obstruction of the small segmental and subsegmental arteries can induce pulmonary thromboembolism (PTE) and PTE-related pulmonary parenchymal changes. An uncommon but potentially serious side effect of tracheal intubation for general anesthesia is an animal severing the endotracheal tube (ET) upon regaining consciousness but prior to extubation. There is very little information in the veterinary literature regarding treatment options and outcome of this potential side effect of intubation.

The purpose of this study was to characterize the signalment, procedure performed prior to ET amputation, removal method used and success of removal, appearance of the airways after removal of the ET, anesthetic complications, and additional treatment administered in cases diagnosed with severed endotracheal tube tracheal foreign bodies. Medical records from the Ontario Veterinary College and the Mississauga-Oakville Veterinary Emergency Hospital were searched for cases with endotracheal tube tracheal foreign body. Two cats and three dogs were presented in the period from 1990 to 2011, ranging from 1 to 7 years of age and 4 to 40 kg. Procedures performed prior to endotracheal tube amputation included ovariohysterectomy, cryptorchidectomy, onychectomy, endoscopic esophageal foreign body removal, and surgical correction of gastric dilation and volvulus. ET size and type were not recorded in 4 of 5 cases. Amputation of the ET occurred in general veterinary practice for the first three cases, and in a specialty referral practice in the last two cases. In cases where amputation occurred in general veterinary practice, the animals were sedated and transported to a specialty referral practice. The severed ET was successfully removed in all cases using grasping endoscopic forceps with bronchoscopic visualization, with no recorded anesthetic complications. Flexible bronchoscopy was utilized in 2 cases, and rigid bronchoscopy was utilized in 1 case; in the remaining 2 cases, the type of endo-scope used was not recorded. After removal of the ET, the trachea was noted to have mild to moderate erythema in 3 cases, and there was mild laryngeal erythema and swelling in an additional case. One cat had a mild cough on recovery that resolved within 24 hours. One dog developed pyrexia 24 hours postremoval, which was attributed to complications from the surgery to correct gastric dilation and volvulus. Medications administered secondary to removal of the ET included a single intravenous, anti-inflammatory dose of dexamethasone in two cases; one of these two cases also received a 5 day course of antibiotic. All cases were discharged from hospital with complete resolution of all respiratory signs within 3 days of presentation.

While the incidence appears rare, severed ET tracheal foreign bodies can occur during recovery from general anesthesia and require immediate intervention. Both rigid and flexible bronchoscopy with grasping forceps were shown to be effective methods to remove an ET tracheal foreign body.

Surveillance for exposure to tick-borne pathogens provides important information necessary for protecting local human and animal health, deployed military servicemembers and working dogs in various parts of the world. The purpose of this study was to determine the molecular and seroprevalence of tick-borne pathogens in military working dogs, shelter animals and pet populations in Barranquilla, Colombia. EDTA-anticoagulated whole blood and serum samples were collected from 70 military working dogs, 101 dogs from regional and community shelters and 47 client-owned animals presenting to veterinary hospitals. Serum samples were tested for antibodies to Ehrlichia ewingii, E. canis, E. chaffeensis, Anaplasma phagocytophilum, A. platys and Borrelia burgdorferi using species-specific peptides in an ELISA format. DNA was extracted from each sample of whole blood and tested for target DNA from Ehrlichia spp. and Anaplasma spp. by real-time PCR analysis. Of the 218 serum samples tested, 62 (28.4%) had antibodies only to E. canis and 15 (6.9%) had antibodies only to A. platys. However, 101 (46.3%) samples had antibodies to both E

Leptospirosis is a zoonotic disease of world wide occurrence. It affects more than 150 mammalian species including humans, dogs and cats, among others. Leptospires spp. belong to a group of motile, aerobic or microaerophilic gram-negative spirochetes, with a hook-shaped end. Pathogenic and non-pathogenic species co-exist in nature.Between 2009 and 2011, blood samples collected from 3907 dogs in Germany were sent to our laboratory and screened for Leptospiral antibodies by means of microscopic agglutination tests (MATs) that involved a total of 35 116 assays. The serovars included in the tests were L. Icterohaemorrhagiae, L. Canicola, L. Pomona, L. Bratislava, L. Grippotyphosa, L. Saxkoebing, L. Autumnalis, L. Australis and L. Serjoe. Results were considered positive if the antibody titre was ! 1:400.L. Grippotyphosa and L. Saxkoebing have been reported to be the most common serovars causing Leptospiral infections in dogs. In this study, the prevalence of positive antibody titres for the two particular serovars was found to be relatively low. Whereas the prevalence of L. Saxkoebing showed a transient increase in 2010, the overall pattern of positive antibody responses to L. Grippotyphosa showed a slight decrease during the course of the study.In 2009, 4% of all positive dogs were sero-positive against a single serovar. This proportion remained stable in the following years, albeit with a slightly decreasing tendency. The number of dogs developing dual seropositivity against two serovars increased steadily, as did the number of dogs seropositive against four different serovars simultaneously. None of the tested samples turned out to be seropositive against L. Serjoe.In this study, L. Bratislava, L. Icterohaemorrhagiae and L. Australis were found to be the most frequent serovars eliciting high-titre antibody responses. However, during the course of the study, some significant dynamic alterations in seropositivity frequency were observed with the aforementioned serovars. In 2009 and 2010, the highest antibody titres (1:3200) were most often found in the order of L. Bratislava > L. Autumnalis > L. Australis > L. Icterohaemorrhagiae. In 2011, the order changed to L. Bratislava > L. Australis > L. Icterohaemorrhagiae > L Autumnalis.Our data suggest that, considering the high magnitude of antibody responses, cross reactivities of the wild serovars (L. Bratislava, L. Australis and L. Autumnalis) with vaccination serovars (L. Canicola, L. Icterohaemorrhagiae/ Copenhageni) were rather unlikely. This study shows that the serovars hitherto considered "rare" played a greater role than previously thought in eliciting Leptospiral antibody responses in dogs. Identification of fungal organisms in urine sediment may allow rapid and cost-effective diagnosis of systemic infections. While, B. dermatitidis has been sporadically noted in urine sediment from infected dogs; however, the frequency at which organisms may be identified has not been systematically evaluated. Aims of this prospective study were to determine frequency of identifying B. dermatitidis organisms in urine sediment from dogs with confirmed systemic infections, and to determine whether presence of fungal organisms is more likely in intact males.

Urine was prospectively collected from dogs with cytologically-confirmed blastomycosis prior to initiation of antifungal therapy. Urine sample volume and method of collection were not standardized. Fungal culture, enzyme immunoassay (EIA) for B. dermatitidis antigen, and examination of urine sediment cytospin preparations were performed on each sample.Results were available for 21 dogs (10 MC, 3 MI, 6 FS, 2 FI); B. dermatitidis urine antigen titers were consistent with active infection in all dogs (median, 18.73 EIA units; range, . B. dermatitidis were identified in urine sediment in 2 of 3 MI, 1 of 2 FI, and 0 of 16 MC and FS dogs; the single dog with a positive urine fungal culture was 1 of the 2 MI sedimentpositive dogs.