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The diagnosis of sepsis in calves is challenging. Blood culture and clinical signs combined with a complete blood count have been used for the diagnosis of sepsis. Recent literature in humans and animal species has been focused on sepsis-specific biomarkers, such as procalcitonin (PCT), that may more accurately and efficiently diagnose sepsis. The aim of this study was to evaluate plasma PCT concentrations in healthy and septic calves. Twenty healthy control calves and 58 sick calves with septic systemic inflammatory response syndrome (SIRS) based on SIRS score and clinical findings were included. Calves with septic SIRS were further divided in septic SIRS survivors (SSS) and non-survivors (SSNS). Plasma PCT concentrations were measured with a commercial ELISA assay for cattle. A receiver operating characteristic curve was used to determine cut-off values and corresponding sensitivity and specificity for the diagnosis of sepsis. Differences in plasma PCT concentration between groups (control vs. SSS vs. SSNS) were evaluated. Plasma PCT concentrations in healthy calves and those with septic SIRS were 33.3pg/mL (0-44.3pg/mL) and 166.5pg/mL (85.9-233.0pg/mL), respectively (P<0.001). The optimal cut-off value to predict septic SIRS was 67.39pg/mL (81.0% sensitivity, 95.0% specificity). Plasma PCT concentrations were 127.4pg/mL (72.2-216.0pg/mL) and 234.3pg/mL (204.5-309.4pg/mL) in the SSS and SSNS subgroups, respectively. Statistically significant differences were found among groups (control vs. SSS and SSNS, P<0.0001; SSS vs. SSNS, P>0.05). These results confirmed an increase in plasma PCT concentrations in calves with septic SIRS, as previously reported in humans and other species., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

While certainly not a novel concept, faecal microbiota transplant (FMT) has recently garnered renewed interest in veterinary medicine due to its remarkable success in treating recurrent Clostridium difficile infection (CDI) in man. There is a dearth of information on indications and efficacy of FMT for the treatment of gastrointestinal disorders in the horse; however, based on evidence in man and other veterinary species, and anecdotal reports in horses, FMT may be a useful treatment for selected cases of acute and chronic diarrhoea and inflammatory bowel disease (IBD) in the horse. In the absence of evidence, expert opinion is offered on case selection and FMT procedure. More research is needed to explore the efficacy, indications and optimal preparation, storage and delivery of FMT to horses., (© 2016 EVJ Ltd.)

Equine serum hepatitis (i.e., Theiler's disease) is a serious and often life-threatening disease of unknown etiology that affects horses. A horse in Nebraska, USA, with serum hepatitis died 65 days after treatment with equine-origin tetanus antitoxin. We identified an unknown parvovirus in serum and liver of the dead horse and in the administered antitoxin. The equine parvovirus-hepatitis (EqPV-H) shares <50% protein identity with its phylogenetic relatives of the genus Copiparvovirus. Next, we experimentally infected 2 horses using a tetanus antitoxin contaminated with EqPV-H. Viremia developed, the horses seroconverted, and acute hepatitis developed that was confirmed by clinical, biochemical, and histopathologic testing. We also determined that EqPV-H is an endemic infection because, in a cohort of 100 clinically normal adult horses, 13 were viremic and 15 were seropositive. We identified a new virus associated with equine serum hepatitis and confirmed its pathogenicity and transmissibility through contaminated biological products.

A new enteric virus of adult horses, equine coronavirus (ECoV), has recently been recognized. It is associated with fever, lethargy, anorexia, and less frequently, colic and diarrhea. This enteric virus is transmitted via the feco-oral route and horses become infected by ingesting fecally contaminated feed and water. Various outbreaks have been reported since 2010 from Japan, Europe and the USA. While the clinical signs are fairly non-specific, lymphopenia and neutropenia are often seen. Specific diagnosis is made by the detection of ECoV in feces by either quantitative real-time PCR, electron microscopy or antigen-capture ELISA. Supportive treatment is usually required, as most infections are self-limiting. However, rare complications, such as endotoxemia, septicemia and hyperammonemia-associated encephalopathy, have been reported, and have been related to the loss of barrier function at the intestinal mucosa. This review article will focus on the latest information pertaining to the virus, epidemiology, clinical signs, diagnosis, pathology, treatment and prevention of ECoV infection in adult horses., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Hepatitis C virus (HCV) requires the liver specific micro-RNA (miRNA), miR-122, to replicate. This was considered unique among RNA viruses until recent discoveries of HCV-related hepaciviruses prompting the question of a more general miR-122 dependence. Among hepaciviruses, the closest known HCV relative is the equine non-primate hepacivirus (NPHV). Here, we used Argonaute cross-linking immunoprecipitation (AGO-CLIP) to confirm AGO binding to the single predicted miR-122 site in the NPHV 5'UTR in vivo. To study miR-122 requirements in the absence of NPHV-permissive cell culture systems, we generated infectious NPHV/HCV chimeric viruses with the 5' end of NPHV replacing orthologous HCV sequences. These chimeras were viable even in cells lacking miR-122, although miR-122 presence enhanced virus production. No other miRNAs bound this region. By random mutagenesis, we isolated HCV variants partially dependent on miR-122 as well as robustly replicating NPHV/HCV variants completely independent of any miRNAs. These miRNA independent variants even replicate and produce infectious particles in non-hepatic cells after exogenous delivery of apolipoprotein E (ApoE). Our findings suggest that miR-122 independent HCV and NPHV variants have arisen and been sampled during evolution, yet miR-122 dependence has prevailed. We propose that hepaciviruses may use this mechanism to guarantee liver tropism and exploit the tolerogenic liver environment to avoid clearance and promote chronicity.

The current study aimed at the investigating the potential use of phosphorylated neurofilament H (pNF-H) as a diagnostic biomarker for neurologic disorders in the horse. Paired serum and cerebrospinal fluid (CSF) samples (n=88) and serum only (n=30) were obtained from horses diagnosed with neurologic disorders and clinically healthy horses as control. The neurologic horses consisted of equine protozoal myeloencephalitis (EPM) (38 cases) and cervical vertebral malformation (CVM) (23 cases). Levels of pNF-H were determined using an ELISA. The correlation between CSF and serum concentrations of pNF-H was evaluated using Spearman's Rank test and the significance of the difference among the groups was assessed using a nonparametric test. Horses had higher pNF-H levels in the CSF than serum. Horses afflicted with EPM had significantly higher serum pNF-H levels in comparison to controls or CVM cases. The correlation between CSF and serum pNF-H levels was poor in both the whole study population and among subgroups of horses included in the study. There was significant association between the likelihood of EPM and the concentrations of pNF-H in either the serum or CSF. These data suggest that pNF-H could be detected in serum and CSF samples from neurologic and control horses. This study demonstrated that pNF-H levels in serum and CSF have the potential to provide objective information to help in the early diagnosis of horses afflicted with neurologic disorders., (Published by Elsevier Ltd.)

OBJECTIVE To determine effects of oral administration of Yunnan Baiyao on platelet activation, coagulation, and fibrinolysis in healthy horses. ANIMALS 12 healthy adult horses. PROCEDURES In a randomized blinded crossover study that included a 4-week washout period between treatments, horses were orally administered a paste containing Yunnan Baiyao (15 mg/kg) or placebo at 12-hour intervals for 3 days. Blood samples were collected before start of treatment (time 0) and at 24 and 72 hours for a CBC, measurement of fibrinogen concentration, coagulation screening tests, and a panel of assays to assess platelet activation (including ADP- and collagen-induced aggregation and closure times, flow-cytometric variables of platelet-leukocyte aggregates, platelet membrane P-selectin and phosphatidylserine expression, and microparticle release), von Willebrand factor (vWF) concentration, and cofactor activity. In addition, thrombelastography was used to evaluate fibrin formation in tissue factor-activated whole blood and plasma and to assess tissue plasminogen activator-induced plasma fibrinolysis. For each treatment, values obtained before and 72 hours after start of administration were compared by use of Wilcoxon signed rank tests. RESULTS Yunnan Baiyao treatment had no significant effect on any hemostatic variable, compared with results for the placebo treatment. CONCLUSIONS AND CLINICAL RELEVANCE Administration of Yunnan Baiyao at a dosage typically used in clinical practice had no effect on in vitro measures of platelet or vWF function and no enhancement of fibrin-clot formation or stability. Any hemostatic actions of Yunnan Baiyao may require higher dosages or result from cell-surface interactions at sites of vascular and tissue injury not examined in this study.

Acute leukemia is rare in horses. Herein we describe historical, clinicopathologic, and postmortem findings in 6 horses with acute leukemia. Medical records of horses with >20% bone marrow blasts and cytochemical or immunophenotyping results were reviewed. Affected horses were 2-8 y of age and of different breeds and sex. Horses were presented acutely with nonspecific signs (e.g., fever, lethargy). Characteristic hemogram findings were bi- or pancytopenia with low blast numbers. Histologic examination revealed extramedullary infiltrates, especially in lymph nodes, spleen, kidney, liver, and lungs. Leukemias were classified as B-cell ( n = 3) and acute myeloid leukemia (AML) ( n = 3). Tumors in 4 cases expressed multiple lineage markers, which complicated classification. Acute leukemia should be suspected in horses with moderate-to-severe bi- or pancytopenia. Blood smears should be reviewed for neoplastic cells, and bone marrow examination is required for diagnosis. Leukemia classification is best achieved using combined morphologic, cytochemical, and immunophenotyping results.

Endotoxemia represents a significant clinical and economic problem for the equine industry. This study assesses the kinetics of soluble CD14 (sCD14), chemokine (CC motif) ligand 2 (CCL2), interleukin 10 (IL-10) and plasma procalcitonin (PCT) in healthy horses after the intravenous infusion of lipopolysaccharide (LPS). The aim was to contribute to the basic understanding of the equine species-specific kinetics of these molecules in response to LPS exposure, which could support further findings in clinical studies and identify valuable inflammatory biomarkers for equine practice. Eleven healthy horses were involved in this experimental in vivo study. Horses were classified as healthy before the LPS infusion. After the pre-infusion blood collection (T0), all horses received an infusion of E. coli endotoxin (30ng/kg over 30min). Data and samples were collected 1h (T1), 2 (T2), 3 (T3) and 24h (T24) after infusion. Plasma sCD14, CCL2 and IL-10 were evaluated with a fluorescent bead-based assay, while PCT was evaluated with an equine PCT ELISA assay. A one-way ANOVA test was performed between each blood-sampling time for PCT, sCD14 and IL-10, and a Friedman test was performed for CCL2. Plasma PCT, IL-10 and CCL2 concentrations increased statistically significantly at T1, T2 and T3 compared to T0. No statistically significant differences were found between plasma IL-10 and CCL2 concentrations between T0 vs T24, although plasma PCT values remained high 24h after LPS infusion. Plasma sCD14 concentration showed no statistically significant differences for any of sampling times. Our results demonstrate that LPS injection into healthy horses results in PCT, CCL2 and IL-10 increases in plasma without an increase in sCD14. The increases in PCT, CCL2 and IL-10 are related to the inflammatory response induced by circulating lipopolysaccharide., (Copyright © 2016 Elsevier B.V. All rights reserved.)

OBJECTIVE To evaluate the effects of treatment of horses with standard platelet inhibitors on ex vivo inhibition of platelet activation by equine herpesvirus type I (EHV-I). ANIMALS II healthy adult horses. PROCEDURES In a double-blinded, placebo-controlled crossover study, horses were treated orally for 5 days with theophylline (5 mg/kg, q 12 h), pentoxifylline (10 mg/kg, q 12 h), clopidogrel bisulfate (4 mg/kg, q 24 h), acetylsalicylic acid (20 mg/kg, q 24 h), or placebo. Horses received all treatments, each separated by a 3-week washout period. Platelet-rich plasma was prepared from citrated blood samples obtained before each treatment session and 4 hours after each final drug dose. Platelets were exposed to 2 EHV-I strains (at I plaque forming units/cell) or positive (thrombin-convulxin) and negative control substances for 10 minutes, then platelet activation was assessed by determining the percentages of P-selectin-positive platelets and platelet-derived microparticles (PDMPs; small events positive for annexin V) with flow cytometry. Platelet aggregation in response to 10μM ADP was also assessed. RESULTS No significant differences in median percentages of P-selectin-positive platelets and PDMPs in EHV-I-exposed platelets were identified between measurement points (before and after treatment) for all drugs, nor were differences identified among drugs at each measurement point. Only clopidogrel significantly inhibited platelet aggregation in response to ADP in platelet-rich plasma samples obtained after that treatment session. CONCLUSIONS AND CLINICAL RELEVANCE Treatment of horses with standard platelet inhibitors had no effect on EHV-I-induced platelet α-granule exteriorization or microvesiculation and release of PDMPs ex vivo, suggesting these drugs will not prevent platelet activation induced directly by EHV-I in vivo.

Reasons for Performing Study: A recent survey of European Colleges (European College of Equine Internal Medicine [ECEIM] and European College of Veterinary Surgeons [ECVS]) revealed the different strategies implemented by, and some of the challenges facing, European clinicians presented with cases of post operative ileus (POI). It was concluded that further comparative analysis of opinions, canvassed from additional colleges of equine veterinary specialism worldwide, would provide valuable additional insight into current POI knowledge on a more global scale., Objectives: To report and compare the current strategies favoured by American veterinary specialists when managing POI in horses that underwent emergency colic surgery., Study Design: Cross-sectional survey., Methods: Electronic invitations were sent to 814 Large Animal specialists, including 3 colleges: the American College of Veterinary Internal Medicine (ACVIM), American College of Veterinary Surgeons (ACVS) and the American College of Veterinary Emergency and Critical Care (ACVECC)., Results: The response rate was 14% (115/814). The majority of respondents (68%) reported an estimated prevalence range of POI of 0-20%. The presence of reflux on nasogastric intubation was the main criterion used to define POI. A lesion involving the small intestine was considered the main risk factor for POI. Anti-inflammatory drugs, intravenous (i.v.) fluids and antimicrobial drugs were the primary strategies used when managing POI. Flunixin meglumine and i.v. lidocaine were the drugs most commonly used in the treatment of horses with POI. Supplementary management strategies targeted mainly the prevention of post operative adhesions, infection and inflammation., Conclusions: There is a lack of consensus on the clinical definition of POI. Prospective and objective clinical assessment of the effectiveness of the different strategies contained within this and the European survey is necessary in order to identify a standardised approach to the management of equine POI., (© 2015 EVJ Ltd.)

Objective: To report a transnasal, endoscopically guided ventral surgical approach for accessing the cranial and caudal segments of the sphenopalatine sinus for mass removal in a horse., Study Design: Case report., Animal: Adult horse with acute onset blindness referable to a soft tissue mass within the sphenopalatine sinus., Clinical Report: A 7-year-old Warmblood gelding presented with a history of running into a fence and falling. No neurologic signs were identified at initial examination but acute blindness was noted 3 weeks later. On computed tomography (CT) the sphenopalatine sinus was filled with a large homogeneous mass with poor contrast enhancement that extended dorsally with thinning to the dorsal cortex of the sphenoid bone, just rostral to the entrance of the optic canals into the cranial cavity. Surgical access to the sphenopalatine sinus was achieved using a transnasal, endoscopically guided ventral pharyngotomy approach and the mass lesion was removed. A presumptive diagnosis of chondroma was made based on histopathology. The horse recovered well from surgery, and although it has not regained vision as of 6.5 years postoperatively, the disease has not progressed., Conclusion: Transnasal, endoscopically-guided ventral surgical access to the sphenopalatine sinus is possible in horses and may improve access in horses with disease extending caudally beyond the palatine portion of the sinus. Use of smaller diameter or specialized instruments, such as various endoscopic bone cutting instruments, and CT image guidance may improve sinus access by this route., (© Copyright 2016 by The American College of Veterinary Surgeons.)

Adult equine hepatocytes have proven challenging to culture long term in vitro as they rapidly lose their morphology and functionality, thus limiting studies on liver function and response to disease. In this study, we describe for the first time the differentiation of equine mesenchymal stromal cells (MSC) from a variety of sources into functional hepatocyte-like cells (HLC). First, we differentiated equine umbilical cord blood (UCB)-derived MSC into HLC and found that these cells exhibited a distinct polygonal morphology, stored glycogen as visualized by periodic acid Schiff's reagent staining, and were positive for albumin and other hepatocyte-specific genes. Second, we demonstrated that UCB-HLC could be revived following cryopreservation and retained their phenotype for at least 10 days. Third, we differentiated three sets of MSC from bone marrow (BM), adipose tissue (AT), and peripheral blood (PB), matched within the same horse. We achieved a 100% differentiation success rate with BM, 0% with AT, and 66% with PB. An additional set of nine PB-MSC samples resulted in an overall success rate of 42% (n = 12), and age or gender did not seem to have an effect on the success of hepatic differentiation from that source. In a final set of experiments, we evaluated the use of these HLC as tools in different fields of biomedical research like virology, to study viral growth, and toxicology, to study chemicals with hepatic toxicity. Equine HLC were found susceptible for infection with the equine herpesviruses type 1 (EHV-1), -2, and -5, and exhibited a more sensitive dose-dependent response to arsenic toxicity than the commonly used human hepatocellular cell line HepG2. Taken together, these data indicate that equine MSC can be efficiently differentiated into HLC and these equine HLC could be a useful tool for in vitro studies.

Early diagnosis of high intracranial pressure (ICP) is critical for minimizing progressive brain injury due to reduced cerebral perfusion. In people, detecting enlargement of the optic nerve sheath diameter (ONSD) by transpalpebral ultrasonography has been found to be an accurate test for high ICP. Aims of this prospective, observational, cross-sectional study were to test hypotheses that (1) ultrasonographic measurement of ONSD would be repeatable in horses, (2) have acceptable interobserver agreement, and (3) would be correlated with age and body weight. The sample population included 48 horses without clinical signs of high ICP and with varying ages and body weights. Two observers independently performed ONSD measurements in both eyes. All measurements ranged from 2.6 to 6.5 mm. The mean difference of repeated measures within observers was ≤0.1 mm and the coefficients of variation ranged from 5.0% to 8.8%. The mean difference of measures between observers was ≤0.2 mm. After correcting for performing multiple tests, no significant rank correlation (all r < 0.4 [absolute value]) was detected between ONSD and age or body weight. However, we observed smaller ONSD in foals versus adults (all P ≤ 0.002). In the foals, all observed measures of rostrocaudal and dorsoventral ONSD were <5 mm. In the adults, all observed measures of rostrocaudal and dorsoventral ONSD were ≤6.5 mm. Findings indicated that ultrasonographic ONSD measurement is a feasible test for use in horses of varying ages and sizes. Further investigation of this ultrasonographic measure as a clinical test for horse with suspected high ICP is warranted., (© 2015 American College of Veterinary Radiology.)

The objective of this study was to evaluate the effect of a standardized botanical extract of Croton lechleri, named crofelemer extract, on fecal dry matter and fecal scores on diarrheic newborn Holstein bull calves induced by enterotoxigenic Escherichia coli. A double-blinded randomized clinical trial was performed in which 60 newborn Holstein bull calves were clean caught and transported to an isolation facility where calves were individually housed and randomly assigned to 1 of 3 treatment groups: placebo (control), enteric-coated formulation of crofelemer extract (ECROF), and nonenteric-coated formulation of crofelemer extract (CROF). Diarrhea was induced at first feeding with an inoculum of the enterotoxigenic Escherichia coli (ATCC 31616) administered with a third of the recommended dose of a colostrum replacer. All calves enrolled in this study received treatments starting on the second feeding (diarrhea onset) and treatments were administered before feeding time (0600 and 1600h) for 6 feedings consecutively. All calves in this study had failure of passive transfer. The only cause of death in this study was due to septicemia, accounting for 1 death out of each treatment group. All the calves were examined twice daily, within 2h after feeding, from d 1 (prechallenge) until 10, on d 15, and a last examination on d 25 of life. Five parameters were evaluated during each examination; rectal temperature, clinical assessment of dehydration status, fecal scores, attitude, and appetite. No differences were observed between treatment groups for rectal temperature, attitude, and appetite. Fecal dry matter was analyzed as prechallenge fecal dry matter, dry matter during treatment, and fecal dry matter after treatment cessation. No difference in prechallenge fecal dry matter was observed and prechallenge fecal dry matter was used as a covariate in the models. Fecal dry matter during treatment was significantly higher for ECROF calves when compared with control calves and CROF calves. Additionally, ECROF fecal dry matter after treatment cessation had a statistical tendency to be higher when compared with control calves. Together, these results suggest that enteric-coated formulation of the standardized crofelemer extract, a natural-product with antisecretory properties, can significantly increase fecal dry matter of neonatal calves with experimentally induced enterotoxigenic Escherichia coli diarrhea. More research is needed to test the efficacy of enteric-coated crofelemer on incidence and severity of secretory diarrhea on calves under natural challenge conditions., (Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Background: The diseases most frequent associated with SIRS in adult horses are those involving the gastrointestinal tract. An early diagnosis should be the goal in the management of horses with SIRS., Objective: The objective of this study was to evaluate the plasma procalcitonin (PCT) concentration in healthy and SIRS horses to assess differences between the two groups., Animals: Seventy-eight horses (30 healthy and 48 SIRS)., Methods: Prospective in vivo multicentric study. Horses were classified as SIRS if at least 2 of the following criteria were met: abnormal leukocyte count or distribution, hyperthermia or hypothermia, tachycardia, tachypnea. Healthy horses showed no clinical or laboratory signs of SIRS. Plasma PCT concentrations were measured with a commercial ELISA assay for equine species. Results were expressed as mean±standard deviation. T-test for unpaired data was performed between healthy and SIRS group. SIRS group was divided in 4 subgroups and t-test was performed between healthy versus each subgroup., Results: PCT concentrations in healthy and SIRS horses were 18.28 ± 20.32 and 197.0 ± 117.0 pg/mL, respectively. T-test showed statistical differences between healthy versus SIRS group and between healthy versus all subgroups., Conclusions and Clinical Importance: Results showed an increase in PCT concentration in SIRS horses as previously reported in humans and dogs. PCT could be used as a single assay in equine practice for detection of SIRS., (Copyright © 2015 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Unlabelled: Equine hepacivirus (EHCV; nonprimate hepacivirus) is a hepatotropic member of the Flaviviridae family that infects horses. Although EHCV is the closest known relative to hepatitis C virus (HCV), its complete replication kinetics in vivo have not been described, and direct evidence that it causes hepatitis has been lacking. In this study, we detected EHCV in 2 horses that developed post-transfusion hepatitis. Plasma and serum from these horses were used to experimentally transmit EHCV to 4 young adult Arabian horses, two 1-month-old foals (1 Arabian and 1 Arabian-pony cross), and 2 foals (1 Arabian and 1 Arabian-pony cross) with severe combined immunodeficiency (SCID). Our results demonstrated that EHCV had infection kinetics similar to HCV and that infection was associated with acute and chronic liver disease as measured by elevations of liver-specific enzymes and/or by histopathology. Although most of these animals were coinfected with equine pegivirus (EPgV), also a flavivirus, EPgV viral loads were much lower and often undetectable in both liver and blood. Three additional young adult Arabian-pony crosses and 1 SCID foal were then inoculated with plasma containing only EHCV, and evidence of mild hepatocellular damage was observed. The different levels of liver-specific enzyme elevation, hepatic inflammation, and duration of viremia observed during EHCV infection suggested that the magnitude and course of liver disease was mediated by the virus inoculum and/or by host factors, including breed, age, and adaptive immune status., Conclusion: This work documents the complete infection kinetics and liver pathology associated with acute and chronic EHCV infection in horses and further justifies it as a large animal model for HCV., (© 2015 by the American Association for the Study of Liver Diseases.)

Nonprimate hepacivirus (NPHV) is the closest known relative of hepatitis C virus (HCV) and its study could enrich our understanding of HCV evolution, immunity, and pathogenesis. High seropositivity is found in horses worldwide with ∼ 3% viremic. NPHV natural history and molecular virology remain largely unexplored, however. Here, we show that NPHV, like HCV, can cause persistent infection for over a decade, with high titers and negative strand RNA in the liver. NPHV is a near-universal contaminant of commercial horse sera for cell culture. The complete NPHV 3'-UTR was determined and consists of interspersed homopolymer tracts and an HCV-like 3'-terminal poly(U)-X-tail. NPHV translation is stimulated by miR-122 and the 3'-UTR and, similar to HCV, the NPHV NS3-4A protease can cleave mitochondrial antiviral-signaling protein to inactivate the retinoic acid-inducible gene I pathway. Using an NPHV consensus cDNA clone, replication was not observed in primary equine fetal liver cultures or after electroporation of selectable replicons. However, intrahepatic RNA inoculation of a horse initiated infection, yielding high RNA titers in the serum and liver. Delayed seroconversion, slightly elevated circulating liver enzymes and mild hepatitis was observed, followed by viral clearance. This establishes the molecular components of a functional NPHV genome. Thus, NPHV appears to resemble HCV not only in genome structure but also in its ability to establish chronic infection with delayed seroconversion and hepatitis. This NPHV infectious clone and resulting acute phase sera will facilitate more detailed studies on the natural history, pathogenesis, and immunity of this novel hepacivirus in its natural host.

Objective: To summarize the current medical literature and provide a clinical perspective of whole blood point-of-care (POC) biomarkers in large animal emergency and critical care practice., Data Sources: Original studies, reviews, and textbook chapters in the human and veterinary medical fields., Summary: POC biomarkers are tests used to monitor normal or disease processes at or near the patient. In both human and veterinary medicine these tools are playing an increasingly important role in the management of critical diseases. The most important whole blood POC biomarkers available for veterinary practitioners include l-lactate, cardiac troponin I, serum amyloid A, triglyceride, creatinine, and glucose, although many other tests are available or on the horizon., Conclusion: Whole blood POC biomarkers enable clinicians to provide improved management of critical diseases in large animals. These tools are especially useful for establishing a diagnosis, guiding therapy, and estimating disease risk and prognosis., (© Veterinary Emergency and Critical Care Society 2015.)

Besnoitiosis is an emerging infectious disease of donkeys (Equus asinus) in the United States for which there are currently no serologic methods of diagnosis. A study was performed to evaluate physical examination findings and 3 serologic assays for the detection of Besnoitia bennetti infection in donkeys. A prospective study of 416 donkeys from 6 privately owned herds across 5 U.S. states (New York, Pennsylvania, Vermont, Oregon, and Washington) was performed. Donkeys were examined for clinical lesions suggestive of besnoitiosis and evaluated for antibodies against B. bennetti using a fluorescent antibody test (FAT) and 2 immunoblot assays specific for bradyzoite and tachyzoite antigens, respectively. Donkeys were confirmed to be infected with B. bennetti by histology (cases; n = 32) and 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 83% of the time. Donkeys with besnoitiosis had significantly higher FAT titers (P < 0.001) and numbers of bradyzoite (P < 0.001) and tachyzoite (P < 0.001) immunoblot bands than control donkeys. The sensitivity and specificity of the serologic assays for detecting besnoitiosis was 88% and 96% for FAT, 81% and 91% for bradyzoite immunoblot, and 91% and 92% for tachyzoite immunoblot, respectively. Fluorescent antibody and immunoblot assays are effective at identifying donkeys with besnoitiosis and provide a more efficient and less invasive diagnostic alternative to histology., (© 2014 The Author(s).)

Objective: To evaluate the effects of clopidogrel on clinical and clinicopathologic variables in healthy horses with experimentally induced endotoxemia., Animals: 12 adult mares. Procedures-Horses were assigned with a randomization procedure to receive clopidogrel (4 mg/kg, once, then 2 mg/kg, q 24 h; n = 6) or a placebo (6) through a nasogastric tube. After 72 hours of treatment, horses received lipopolysaccharide (LPS; 30 ng/kg, IV). Heart rate, respiratory rate, rectal temperature, CBC variables, plasma fibrinogen concentration, serum tumor necrosis factor-α concentration, plasma von Willebrand factor concentration, and measures of platelet activation (including ADP- and collagen-induced platelet aggregation and closure times, thrombelastography variables, and results of flow cytometric detection of platelet membrane P-selectin, phosphatidylserine, and microparticles) were determined at various times before and after LPS administration by investigators unaware of the treatment groups. Statistical analyses were performed with repeated-measures ANOVA., Results: 4 of 6 clopidogrel-treated horses had significant decreases in ADP-induced platelet aggregation before and after LPS administration. Heart rate increased significantly after LPS administration only for the placebo group. No significant differences were detected between groups for CBC variables, closure time, and plasma concentration of fibrinogen or serum concentration of tumor necrosis factor-α, and no clinically relevant differences were detected for other hemostatic variables., Conclusions and Clinical Relevance: In this study, administration of LPS did not induce platelet hyperreactivity in horses on the basis of measures of platelet adhesion, aggregation, degranulation, and procoagulant activity. Administration of clopidogrel was associated with variable platelet antiaggregatory activity and attenuated some clinical signs of endotoxemia.

Leptospirosis, caused by Leptospira spp., is one of the most common zoonotic diseases in the world. We tested four recombinant proteins of Leptospira interrogans, namely, rLipL21, rLoa22, rLipL32, and rLigACon4-8, to evaluate their potential for use as antigens for the diagnosis of equine leptospirosis. We employed equine sera (n = 130) that were microscopic agglutination test (MAT) negative and sera (n = 176) that were MAT positive for the 5 serovars that most commonly cause equine leptospirosis. The sensitivity and specificity of ELISA compared to MAT were 82.39% and 86.15%, respectively, for LigACon4-8, 77.84% and 92.31%, respectively, for Loa22, 77.84% and 86.15%, respectively, for LipL32, and 84.66% and 83.85%, respectively, for LipL21. When one of the two antigens was test positive, the sensitivity and specificity of ELISA were 93.75% and 78.46%, respectively, for rLigACon4-8 and LipL32, 93.18% and 76.15%, respectively, for rLigACon4-8 and LipL21, 89.77% and 80.77%, respectively, for rLigACon4-8 and Loa22, 91.48% and 78.46%, respectively, for LipL21 and Loa22, 93.75% and 76.92%, respectively, for LipL21 and LipL32, and 90.34% and 80.77%, respectively, for Loa22 and LipL32. In conclusion, we have developed an indirect ELISA utilizing rLigACon4-8, rLoa22, rLipL32, and rLipL21 as diagnostic antigens for equine leptospirosis. The use of four antigens in the ELISA was found to be sensitive and specific, the assay was easy to perform, and the results concurred with the results of the standard Leptospira MAT.

Reasons for Study: To determine whether low-dose, low-frequency doxycycline administration is capable of achieving chondroprotective concentrations within synovial fluid (SF) while remaining below minimum inhibitory concentration 90 (MIC90 ) of most equine pathogens and would be an option in the management of osteoarthritis., Objectives: To determine whether low-dose, low-frequency oral administration of doxycycline can attain in vivo SF concentrations capable of chondroprotective effects through reduction of matrix metalloproteinase (MMP)-13 activity, while remaining below MIC90 of most equine pathogens., Study Design: Descriptive pharmacokinetic study with crossover design., Methods: Two groups of 6 horses received oral doxycycline. Plasma and SF doxycycline concentrations were measured using high performance liquid chromatography. Group 1 received 5 mg/kg bwt q. 24 h with 21 blood and 8 SF samples collected over 120 h; Group 2 received 5 mg/kg bwt q. 48 h with 27 blood and 11 SF samples collected over 192 h. Cultured synoviocytes were treated with interleukin-1α (1 ng/ml) for 24 h to stimulate MMP synthesis, and then SF was added to the culture medium for 96 h. MMP-13 protein and mRNA were measured in synoviocyte culture medium and synoviocytes, respectively., Results: Mean doxycycline concentration ≥0.043 μg/ml (previously demonstrated to inhibit MMP-13) was achieved in plasma by t = 0.25 h and SF by t = 48 h in Group 1, and in plasma by t = 0.17 h and SF by t = 1 h in Group 2. Synoviocyte culture medium containing doxycycline from Groups 1 and 2 had significantly decreased active MMP-13 protein concentration, and synoviocytes cultured in this medium had significantly decreased MMP-13 gene expression compared to controls. Plasma doxycycline concentration in both groups and SF doxycycline concentration in Group 2 demonstrated a cumulative effect., Conclusions: Low-dose orally administered doxycycline achieves SF concentrations in vivo capable of diminishing MMP-13 expression. This study supports the use of doxycycline as a disease modifying osteoarthritic drug., (© 2013 EVJ Ltd.)

Donkeys (Equus asinus) are used as both companion and working animals throughout the world and in some countries, their meat and milk are used for human consumption. Here we report the first serological survey of Toxoplasma gondii in donkeys in the United States. Serum samples from 373 donkeys from eight farms in five states were tested for T. gondii antibodies by the modified agglutination test (MAT). Twenty-four of 373 (6.4%) of donkeys were seropositive, with MAT titers ranging from 25 to ≥ 200. All seropositive donkeys were Miniature breed. Seropositivity prevalence was 7.0% in female donkeys (20/282) and 4.1% in male donkeys (4/91). No donkeys less than 24 months of age (129) were seropositive, suggesting postnatal transmission of infection. Domestic cats were present on six of the eight farms. Three cats from one farm had MAT titers of 200. Viable T. gondii was isolated from the hearts of two cats, but not from brain tissues. Genotyping of isolate DNA extracted from culture-derived tachyzoites using 10 PCR-restriction fragment length polymorphism (RFLP) markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, PK1, L358 and Apico loci) revealed that both isolates were clonal Type II (ToxoDB PCR-RFLP genotype #1). This is the first serological survey for T. gondii in donkeys in the United States, and suggests that donkey milk and meat should be considered as a potential source for human infection. The role of barn cats in the transmission of T. gondii to donkeys on farms warrents further investigation., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Objective: To evaluate the platelet activation response before and after treatment with clopidogrel in horses., Animals: 12 healthy adult mares., Procedures: In a masked study, horses (6/group) were randomly allocated to alternately receive placebo or clopidogrel via nasogastric tube at a loading dose of 4 mg/kg followed by 2 mg/kg every 24 hours. Blood samples were collected before and 72 hours after initiation of treatment for ADP- and collagen-induced light transmission aggregometry; determination of closure time in collagen-ADP cartridges; modified thrombelastography for comparison of maximal amplitudes generated by kaolin, reptilase, and reptilase plus ADP activation; and flow cytometric tests to detect platelet fibrinogen binding, P-selectin expression, and phosphatidylserine externalization before and after ex vivo stimulation with thrombin, convulxin, thrombin with convulxin, and calcium ionophore., Results: Clopidogrel administration induced a significant decrease in mean aggregation response to 5 μM and 10 μM ADP stimulation; however, 2 horses had resistance to clopidogrel's inhibitory action. Significant differences after clopidogrel treatment were not found in any other tests of platelet function., Conclusions and Clinical Relevance: Assays using commercially available reagents were configured to measure different variables of the platelet activation response; however, clopidogrel's platelet inhibitory action was only detected by ADP-induced light transmission aggregometry. Results also suggested that horses, like humans, have interindividual variability in response to clopidogrel that may influence the drug's clinical efficacy as an antiplatelet agent.

Reasons for Performing Study: Placentitis is a prevalent cause of abortion, premature delivery and neonatal death in mares. Early diagnosis is paramount for the survival of the fetus and delivery of a live foal., Objectives: To determine: 1) Serum amyloid A (SAA) profile in healthy mares during late gestation; 2) if placentitis affects SAA concentrations and 3) the effects of therapy on SAA concentrations and pregnancy outcome in mares with placentitis., Methods: In Experiment I, 15 healthy pregnant mares were evaluated from 280 days of gestation to 60 h post partum. In Experiment II, pregnant mares were inoculated intra-cervically with Streptococcus zooepidemicus (Day 280-295) and assigned to control (n = 5) and treatment (n = 9) groups. Treatment was initiated at the onset of clinical signs. Serum amyloid A concentrations were determined prior to inoculation and then weekly until abortion or delivery., Results: Serum amyloid A remained at low concentrations (95% confidence interval [CI]: 3.2-8.1 mg/l) during late gestation followed by a significant increase within 36 h post partum; SAA returned to basal concentrations by 60 h post partum. In Experiment II, SAA significantly increased within 96 ± 56 h of inoculation in control mares followed by abortion. Therapy was effective (P<0.05) in preventing the rise in SAA in 66% (6/9) of mares and only one out of 3 mares with increased SAA aborted. Overall, the incidence of abortion was higher in mares with increased SAA concentrations (75%; 6/8) compared with mares in which SAA remained at baseline concentrations (0/6)., Conclusions: Mares with placentitis had significant increased SAA within 96 h post inoculation and concentrations remained increased until abortion in untreated mares. Successful treatment either prevented the rise of SAA concentration or decreased its concentration to baseline concentrations, followed by delivery of a live foal., Potential Relevance: Serum amyloid A may be used as a prognostic indicator in cases of ascending placentitis in the mare., (© 2012 EVJ Ltd.)

Leptospira immunoglobulin (Ig)-like (Lig) proteins are a novel family of surface-associated proteins in which the N-terminal 630 amino acids are conserved. In this study, we truncated the LigA conserved region into 7 fragments comprising the 1st to 3rd (LigACon1-3), 4th to 7.5th (LigACon4-7.5), 4th (LigACon4), 4.5th to 5.5th (LigACon4.5-5.5), 5.5th to 6.5th (LigACon5.5-6.5), 4th to 5th (LigACon4-5), and 6th to 7.5th (LigACon6-7.5) repeat domains. All 7 recombinant Lig proteins were screened using a slot-shaped dot blot assay for the diagnosis of equine leptospirosis. Our results showed that LigACon4-7.5 is the best candidate diagnostic antigen in a slot-shaped dot blot assay. LigACon4-7.5 was further evaluated as an indirect enzyme-linked immunosorbent assay (ELISA) antigen for the detection of Leptospira antibodies in equine sera. This assay was evaluated with equine sera (n = 60) that were microscopic agglutination test (MAT) negative and sera (n = 220) that were MAT positive to the 5 serovars that most commonly cause equine leptospirosis. The indirect ELISA results showed that at a single serum dilution of 1:250, the sensitivity and specificity of ELISA were 80.0% and 87.2%, respectively, compared to those of MAT. In conclusion, an indirect ELISA was developed utilizing a recombinant LigA fragment comprising the 4th to 7.5th repeat domain (LigACon4-7.5) as a diagnostic antigen for equine leptospirosis. This ELISA was found to be sensitive and specific, and it yielded results that concurred with those of the standard MAT.

Objective: To assess pharmacokinetics of pregabalin in horses after a single intragastric or IV dose., Animals: 5 healthy adult mares., Procedures: Horses received 1 dose of pregabalin (approx 4 mg/kg) via nasogastric tube in a crossover-design study; after a 3-week washout period, the same dose was administered IV. Food was not withheld. Plasma pregabalin concentrations in samples obtained 0 to 36 hours after administration were measured by use of ultra-performance liquid chromatography with triple quadrupole tandem mass spectrometry. Pharmacokinetic variables were estimated by means of noncompartmental analysis., Results: Mild sedation was observed in 2 horses following intragastric and IV pregabalin administration. Signs of mild, transient colic or behavioral abnormalities were observed in all horses following IV administration. After intragastric administration, median (range) maximal plasma concentration was 5.0 μg/mL (4.4 to 6.7 μg/mL), time to maximal plasma concentration was 1. 0 hour (0.5 to 2.0 hours), elimination half-life was 8.0 hours (6.2 to 9.4 hours), and area under the curve from time 0 to infinity (AUC0-∞) was 47.2 μg·h/mL (36.4 to 58.4 μg·h/mL). After IV administration, initial concentration was 22.2 μg/mL (19.8 to 27.7 μg/mL), elimination half-life was 7.74 hours (6.94 to 8.17 hours), and AUC0-∞ was 48.3 μg·h/mL (44.8 to 57.2 μg·h/mL). Bioavailability was 97.7% (80.7% to 109.8%). Median predicted values for minimal, mean, and maximal steady-state plasma concentrations after intragastric administration assuming an 8-hour dosing interval were 3.9, 5.3, and 6.3 μg/mL, respectively., Conclusions and Clinical Relevance: At a simulated intragastric dosage of approximately 4 mg/kg every 8 hours, median pregabalin steady-state plasma concentration in healthy horses was within the therapeutic range reported for humans. Therapeutic concentrations and safety of this dosage have not been established in horses.

Theiler's disease is an acute hepatitis in horses that is associated with the administration of equine blood products; its etiologic agent has remained unknown for nearly a century. Here, we used massively parallel sequencing to explore samples from a recent Theiler's disease outbreak. Metatranscriptomic analysis of the short sequence reads identified a 10.5-kb sequence from a previously undescribed virus of the Flaviviridae family, which we designate "Theiler's disease-associated virus" (TDAV). Phylogenetic analysis clusters TDAV with GB viruses of the recently proposed Pegivirus genus, although it shares only 35.3% amino acid identity with its closest relative, GB virus D. An epidemiological survey of additional horses from three separate locations supports an association between TDAV infection and acute serum hepatitis. Experimental inoculation of horses with TDAV-positive plasma provides evidence that several weeks of viremia preceded liver injury and that liver disease may not be directly related to the level of viremia. Like hepatitis C virus, the best characterized Flaviviridae species known to cause hepatitis, we find TDAV is capable of efficient parenteral transmission, engendering acute and chronic infections associated with a diversity of clinical presentations ranging from subclinical infection to clinical hepatitis.

Reasons for Performing Study: To date, no information is available on the true biological elimination half-life (T(1/2) ) of cardiac troponin I (cTnI) in the equine species. Such data are required to better evaluate the optimal time to acquire the cTnI sample following acute myocardial injury., Objective: To determine the T(1/2) of equine cTnI., Methods: Four healthy ponies received i.v. injections of recombinant equine cTnI. Plasma cTnI concentrations were measured with a point-of-care cTnI analyser at multiple time points after injection. Standard pharmacokinetic analysis was performed to establish the T(1/2) of cTnI., Results: The average T(1/2) of cTnI was determined to be 0.47 h using a single rate elimination model., Conclusion: The elimination of recombinant equine cTnI following i.v. administration is very rapid. Establishing the T(1/2 ) of troponin provides critical information in understanding the clinical application of this cardiac biomarker in equine practice., (© 2012 EVJ Ltd.)

Equine motor neuron disease (EMND) is a neurodegenerative disorder of unknown etiology affecting horses worldwide. Trans-Active Response DNA Binding Protein of 43 kDa (TDP-43) has been reported in the central nervous system (CNS) of several neurodegenerative conditions in humans including Amyotrophic Lateral Sclerosis (ALS) and assumed to play role in the disease. We examined whether horses afflicted with EMND express the TDP-43 in CNS. Ten horses with EMND and 6 controls of different ages and breed we enrolled. Detection of presence of TDP-43 protein in the CNS was analyzed by immunohistochemical staining using rabbit anti-human TARDBP (TDP-43) polyclonal antibody. Formalin fixed neuronal tissues from medulla, cervical, and lumbar spinal cord were harvested from EMND and from control horses. Sections were assigned randomly to TDP-43 treated or rabbit anti-IgG as control. Nuclear staining of TDP-43 was detected in one of the neural tissues of 75 % of EMND-positive and 0 of 0 % of control horses in the central nervous system (medulla, and/or cervical spinal cord and/or lumbar spinal cord). TDP-43 antibody was detected in the nucleus of EMND horses and no cytoplasmic staining was noted. As in ALS, there was no pattern of age clustering associated with the detection of TDP-43. This is the first report on the staining of TDP-43 in neuronal tissues of horses and suggests that TDP-43 may play a role in the pathogenesis of EMND. Further studies are needed to elucidate the etiologic role of this protein in the diseases.

Objective: To investigate the influence of oxidative stress in terms of antioxidant capacity and lipid peroxidation on the probability of motor neuron disease (MND) in horses., Animals: 88 horses with MND (cases) and 49 controls., Procedures: Blood samples were collected from all horses enrolled, and RBCs and plasma were harvested. Activity of the enzyme erythrocytic superoxide dismutase 1 (SOD1) was determined in the RBCs. Plasma concentrations of α-tocopherols and β-carotenes and activity of glutathione peroxidase were also evaluated. Degree of lipid peroxidation was measured by determining plasma concentrations of lipid hydroperoxides. Differences were evaluated between horse groups., Results: Cases had lower erythrocyte SOD1 activity than did controls, but the difference was not significant. On the other hand, plasma vitamin E concentrations differed significantly between groups, with the cases having lower concentrations. Neither plasma vitamin A concentration nor glutathione peroxidase activity differed between groups; however, cases had significantly higher concentrations of lipid hydroperoxides (18.53μM) than did controls (12.35μM)., Conclusions and Clinical Relevance: Horses with MND differed from those without MND by having a lower plasma concentration of vitamin E and higher concentrations of lipid hydroperoxides. Results parallel the findings in humans with sporadic amyotrophic sclerosis and provide evidence supporting the involvement of oxidative stress in the 2 conditions.

Reasons for Performing Study: Selenium and vitamin E deficiency have been associated with nutritional myopathy, more commonly known as white muscle disease (WMD) in horses. However, correlations between selenium concentrations and presenting clinical signs, age, breed, gender, serum vitamin E, creatine kinase (CK) and final diagnosis, have not previously been evaluated., Objectives: To determine the number of hospitalised horses in 3 age groups that were selenium tested and the proportions of horses with categorised presenting clinical signs; the association/odds risk of final diagnosis with selenium deficiency and to examine the association between selenium status, vitamin E status and serum CK in adult horses., Methods: Two hundred and seventy-one hospitalised horses with a selenium concentration evaluated between 1996 and 2011 were examined retrospectively. Records were examined in order to ascertain selenium and vitamin E concentrations, age, breed, gender, CK values, presenting clinical signs and final diagnosis. Data were analysed with proportions, Fisher's exact t test, odds ratios and multivariate linear regressions., Results: Within the < 30 day old age group, 13/20 animals had low selenium concentrations. There were 18/42 horses in the 30 days to 2 years old age group with low selenium and 77/209 horses more than 2 years of age with low selenium. There was an association between low selenium and myopathy in the < 30-day-old animals (P = 0.017), all of which were classified as having WMD. No associations were identified between nutritional myopathy and selenium status in horses between 30 days and 2 years of age or in horses more than 2 years of age., Conclusions and Potential Relevance: This study indicates that WMD occurs most commonly in foals < 30 days old and is associated with low selenium concentrations (7 out of 8 affected foals had blood Selenium levels < 1.26 microm/l). Low serum selenium concentrations are common in hospitalised adult horses while nutritional myopathy is rare in these animals.

Changes in ELISA serology are frequently used to determine antibiotic treatment success for Lyme disease in horses. This concept was based upon a previous report showing a marked decline in ELISA values in experimentally infected and antibiotic-treated ponies. Changes in Lyme serology following antibiotic treatment in naturally infected horses have not been reported. The objective of this study was to compare Borrelia ELISA antibody concentrations in naturally exposed horses both before and following antibiotic treatment for Lyme disease. A retrospective study was performed comparing oxytetracycline- or doxycyclinetreated (n = 68) and untreated (n = 183) horses from a single equine practice and their change in Borrelia ELISA values over a similar time period. Antibiotictreated horses had a decline in ELISA values in comparison to control horses (P ≤ 0.05) and untreated horses were twice as likely to have their ELISA values increase (OR = 0.5; 95% C.I. = 0.3-0.9) compared to treated horses. The magnitude of the decline in ELISA units following treatments was small compared to that previously reported in experimentally infected and treated ponies. Field-exposed horses with high Borrelia burgdorferi ELISA values who are treated with either oxytetracycline or doxycycline can be expected to have only a small decline in ELISA values following treatment. Persistently high ELISA titres following appropriate treatments for Lyme disease may not, without appropriate clinical signs, be a reason for more prolonged treatment.

Borrelia burgdorferi, the etiologic agent of Lyme disease is a tick born spirochetal infection. Clinical signs of Lyme borreliosis are uncommon in horses, but when present they are often vague and nonspecific. In horses, Lyme borreliosis has been implicated in musculoskeletal, neurological, reproductive, and ocular disorders, including uveitis, but definitive diagnosis can be challenging as the causative agent is rarely isolated and serologic tests can be unreliable and do not confirm active disease. Here, we report two cases of equine uveitis associated with B. burgdorferi based on the identification of spirochetes within ocular fluids and confirmed with PCR testing. The two cases illustrate some of the challenges encountered in the recognition and diagnosis of equine Lyme borreliosis. Although only one of many possible causes of equine uveitis, Lyme disease should be considered a differential diagnosis, especially in endemic areas. Given the possibility for false negative results of serum tests during uveitis associated with B. burgdorferi and the failure of such tests to confirm active infection, a combination of cytologic assessment, antibody, and/or PCR testing of ocular fluids may be worthwhile if the clinical suspicion for Lyme uveitis is high., (© 2012 American College of Veterinary Ophthalmologists.)

Background: Research in humans has demonstrated that high serum iron (sFe) concentration can predispose to infection, and many infections subsequently result in alterations of host sFe. A decrease in sFe concentration is an early and sensitive indicator of systemic inflammation caused by tissue necrosis, bacterial infections, or endotoxemia in horses. Serum iron parameters in acute equine herpesvirus type 1 (EHV-1) infection have not been evaluated previously., Objectives: To document the sFe response to EHV-1 infection and to determine whether or not significant differences in sFe concentration exist between EHV-1 infected horses that develop neurologic disease and those that do not., Animals: A total of 14 horses experimentally infected with EHV-1., Methods: Data were collected as an ancillary data set during a blinded experimental EHV-1 infection. Horses were infected with the rAb4 strain of EHV-1. Temperature, neurologic score, packed cell volume (PCV), and sFe parameters (sFe concentration, % saturation, and total iron-binding capacity) were recorded daily for 2 weeks. Data were evaluated using Wilcoxon signed rank tests and Wilcoxon rank sum tests with Bonferroni corrections., Conclusions and Clinical Relevance: Serum iron concentration decreases significantly in a biphasic pattern after EHV-1 infection. There was no significant difference in sFe concentration in horses that developed neurologic disease and those that did not in these experimentally infected animals. Serum iron parameters may be useful in monitoring the clinical course of viral infections such as EHV-1., (Copyright © 2012 by the American College of Veterinary Internal Medicine.)

Reasons for Performing Study: Minocycline holds great potential for use in horses not only for its antimicrobial effects but also for its anti-inflammatory and neuroprotective properties. However, there are no pharmacokinetic or safety data available regarding the use of oral minocycline in horses., Objectives: To determine pharmacokinetics, safety and penetration into plasma, synovial fluid, aqueous humour (AH) and cerebral spinal fluid (CSF) of minocycline after oral administration of multiple doses in horses and to determine the minimum inhibitory concentrations (MIC) of minocycline for equine pathogenic bacteria., Methods: Six horses received minocycline (4 mg/kg bwt q. 12 h for 5 doses). Thirty-three blood and 9 synovial fluid samples were collected over 96 h. Aqueous humour and CSF samples were collected 1 h after the final dose. Minocycline concentrations were measured using high pressure liquid chromatography. The MIC values of minocycline for equine bacterial isolates were determined., Results: At steady state, the mean ± s.d. peak concentration of minocycline in the plasma was 0.67 ± 0.26 µg/ml and the mean half-life was 11.48 ± 3.23 h. The highest trough synovial fluid minocycline concentration was 0.33 ± 0.12 µg/ml. The AH concentration of minocycline was 0.09 ± 0.03 µg/ml in normal eyes and 0.11 ± 0.04 µg/ml in blood aqueous barrier-disrupted eyes. The mean CSF concentration of minocycline was 0.38 ± 0.09 µg/ml. The MIC values were determined for 301 isolates. Minocycline concentrations were above the MIC(50) and MIC(90) for many gram-positive equine pathogens., Potential Relevance: This study supports the use of orally administered minocycline at a dose of 4 mg/kg bwt every 12 h for the treatment of nonocular infections caused by susceptible (MIC ≤ 0.25 µg/ml) organisms in horses. Further studies are required to determine the dose that would be effective for the treatment of ocular infections., (© 2011 EVJ Ltd.)

Objective: To describe the clinical, endoscopic, and serologic features of an outbreak of besnoitiosis in 2 donkey operations in northeastern Pennsylvania and to report the outcome of attempted treatment of 1 naturally infected individual., Design: Observational study., Animals: 29 donkeys (Equus asinus) in northeastern Pennsylvania., Procedures: Donkeys were examined for lesions suggestive of besnoitiosis in an outbreak investigation. Information was collected regarding the history and signalment of animals on each premises. Rhinolaryngoscopy was performed to identify nasopharyngeal and laryngeal lesions. Serum samples were collected for immunofluorescent antibody testing and immunoblotting for Besnoitia spp. Skin biopsy samples were obtained from 8 animals with lesions suggestive of besnoitiosis for histologic examination. Quantitative real-time PCR assay for Besnoitia spp was performed on tissue samples from 5 animals., Results: Besnoitiosis was confirmed in 6 of the 8 suspected cases. The most common lesion site was the nares, followed by the skin and sclera. Donkeys with clinical signs of disease had higher serum antibody titers and tested positive for a greater number of immunoblot bands than did donkeys without clinical signs of disease. All animals evaluated by PCR assay tested positive. Putative risk factors for disease included age and sex. Ponazuril was not effective at treating besnoitiosis in a naturally infected donkey., Conclusions and Clinical Relevance: Knowledge of clinical and serologic features of besnoitiosis in donkeys will assist clinicians in the diagnosis and prevention of this disease in donkey populations. Besnoitiosis may be an emerging disease of donkeys in the United States.

Hepcidin has been found to be the key regulator of iron metabolism that leads to hypoferremia during inflammation. Recent work has shown that equine hepcidin is predominantly expressed in the liver of horses. In this study, hepcidin gene expression was determined in the liver and bone marrow of six healthy horses after iv infusion of Escherichia coli O55:B5 LPS. The IL-6 gene expression was also determined in liver and bone marrow samples. Clinical and laboratory evaluations were measured at multiple time points between 0 and 240 h post-LPS infusion (PI). Liver and bone marrow biopsies were taken immediately before (baseline) and at 6 and 18 h PI. In response to endotoxin infusion, all horses showed characteristic clinical signs of endotoxemia. Plasma iron concentration was decreased significantly from the pre-infusion level at 8 h PI. Hypoferremia peak was observed at 12 h and returned to normal levels at 30 h PI. Relative real-time RT-PCR analysis showed that liver hepcidin and IL-6 mRNA expression was up-regulated at 6 h PI. Bone marrow hepcidin relative expression was not influenced by LPS infusion. In another experiment, equine monocyte cultures were stimulated with LPS (1 µg/ml). Monocyte hepcidin and IL-6 gene expression was significantly induced after 2 h of LPS stimulus and returned to baseline levels thereafter. The present study describes that, in horses, LPS infusion up-regulates hepatic hepcidin mRNA expression resulting in early observed hypoferremia and suggests that hepcidin may act as an acute-phase protein in horses.

Objective: To evaluate changes in plasma cardiac troponin I (cTnI) and L-lactate (LLt) as prognostic indicators in horses undergoing emergency abdominal surgery., Design: Prospective observational study., Setting: Veterinary teaching hospital., Animals: Thirty-four horses undergoing emergency abdominal surgery., Interventions: Serial blood sampling during various times during hospitalization (hospital admission, and 12, 24, 48, and 72 h postoperatively) evaluating cTnI and LLt concentrations., Measurements and Main Results: All horses required surgery for correction of a strangulating (n = 29) or nonstrangulating obstruction (n = 5) of the small or large intestine. Twenty-seven horses survived to discharge; 7 were euthanized either during (n = 1) or after (n = 6) surgery due to disease severity or systemic complications associated with the primary gastrointestinal lesion. Preoperative cTnI concentrations were increased above the normal reference interval in 24% of horses (8/34, median = 0.01 ng/mL, range = 0-12.23 ng/mL), whereas LLt concentrations were increased above the normal reference interval in 88% of horses (30/34, median = 3.37 mmol/L, range = 0.77-13.26 mmol/L). The LLt concentration was significantly higher (P < 0.05) in nonsurviving compared with surviving horses at admission, and at 24 and 72 hours postoperatively. No significant difference in the cTnI concentration was detected between groups at admission. However, the cTnI concentration was significantly higher (P<0.05) in nonsurviving compared with surviving horses at all time points postoperatively., Conclusions: Measurement of both LLt and cTnI concentrations may provide information for prognostication in surgical colic horses. Marked increases in admission concentrations of LLt (median 7.56 mmol/L) and even moderate postoperative increases in cTnI concentration (median 0.97 ng/mL) may both indicate a poor prognosis in critically ill horses following abdominal surgery., (© Veterinary Emergency and Critical Care Society 2012.)

Objective: To compare effects of 3 methods of topically applied cold treatment (cryotherapy) on digital laminar and venous temperatures in horses., Animals: 9 healthy adult Thoroughbreds., Procedures: Thermocouples were placed in palmar digital veins and digital laminae of both forelimbs of horses. Three methods of cryotherapy were applied to the distal aspects of the limbs: wader boot (63-cm-tall vinyl boot filled with ice and water [ice slurry]), ice bag (5-L fluid bag filled with ice slurry), and a gel pack boot (boot containing frozen gel packs). Gel packs and ice slurries were replenished every hour during cryotherapy. The forelimb that received the first treatment was randomly assigned; thereafter, control and treated forelimbs were alternated for each treatment. For each treatment, temperatures were recorded every minute during 15-minute pretreatment, 2-hour treatment, and ≥ 30 minute rewarming periods. Once temperatures had returned to within 3°C below pretreatment values, the experiment was repeated in a similar manner for other cryotherapy methods., Results: Digital venous temperatures were similar to laminar temperatures during each treatment. Ice bag and wader boot treatments caused similar cooling of digits. Gel boot treatment did not cause substantial cooling of digits., Conclusions and Clinical Relevance: Ice bag treatment caused laminar and digital venous cooling equivalent to that of wader boot treatment. Cryotherapy by use of 5-L fluid bags with an ice slurry may be a readily available, practical, and efficient method for prevention of laminitis in horses. Digital laminar and venous temperatures were similar in forelimbs of horses before and during cryotherapy.

Visualization of colonic mesenteric vasculature during transabdominal ultrasonographic examination of horses with colic can be a predictor of right dorsal displacement of the large colon or 180° large colon volvulus, or both. Medical records of 82 horses having had surgical treatment of colic and having received a transabdominal ultrasonographic examination on admission were reviewed. Colonic mesenteric vessels were sonographically identified coursing laterally on the right side of the abdomen in 24 of the 82 cases. Horses with colonic vessels identified on ultrasound were 32.5 times more likely to be diagnosed at surgery with either large colon right dorsal displacement or 180° large colon volvulus than those in which vessels were not seen (P < 0.001). Visualization of colonic mesenteric vessels on ultrasound provided a sensitivity of 67.7%, specificity of 97.9%, positive predictive value of 95.8%, and negative predictive value of 81% for large colon right dorsal displacement or 180° large colon volvulus, or both.

This case report describes a 10-year-old horse that developed multiple dermal papules over the right masseter area following removal of a tick from the same site 3 months earlier. Histological examination of a biopsy from a papule was suggestive of either a T-cell-rich B-cell lymphoma or cutaneous lymphoid hyperplasia, a form of pseudolymphoma sometimes associated with a tick bite. Positive serological testing and PCR of the biopsy sample for Borrelia in conjunction with immunohistochemical testing of the skin biopsy, the clinical history and response to treatment with doxycycline strongly supported the diagnosis of Borrelia-associated cutaneous pseudolymphoma., (© 2011 The Authors. Veterinary Dermatology. © 2011 ESVD and ACVD.)

Severe acute pancreatitis (SAP) in foals is rare and treatment has not been documented. This paper describes the clinical, haematological and ultrasonographic findings as well as attempted treatment of SAP in two 5-day-old Appaloosa fillies. Clinical signs, including colic, diarrhoea and coma, may be mistaken for sepsis or neonatal encephalopathy. Hyperlipaemic serum and peritoneal fluid, and elevated serum and peritoneal fluid amylase and lipase activities aided the diagnosis. Severe acute pancreatitis should be included as a differential in an acutely ill foal with diarrhoea, colic, cerebral cortica dysfunction and hyperlipaemia.

Background: Myocardial disease in camelids is poorly characterized. Nutritional (selenium deficiency) and toxic (ionophore toxicity) myocardial disease have been reported in camelids. Diagnosis and management of these and other myocardial diseases might be enhanced by evaluating cardiac troponin I (cTnI) concentrations. No information about cTnI reference intervals in camelids is currently available., Hypothesis/objectives: (A) To determine cTnI concentrations obtained using a point of care i-STAT(®)1 analyzer (Heska Corporation) in healthy alpacas; (B) to compare alpaca cTnI concentrations between heparinized whole blood and plasma samples and between 2 different storage conditions (4 °C for 24 h or -80 °C for 30 days); (C) to examine assay reproducibility using the i-STAT(®)1., Animals, Materials and Methods: 23 healthy alpacas were evaluated. Blood and plasma samples were analyzed by the i-STAT(®)1 within 1 h of collection. Aliquots of plasma were stored at either 4 °C for 24 h or -80 °C for 30 days, and then analyzed. Assay reproducibility was determined by comparing 2 plasma or whole blood cTnI concentrations measured on the same sample over a 10 min period., Results: Analyzer-specific plasma cTnI concentrations in clinically normal alpacas had a median of <0.02 ng/mL (range: <0.02 ng/mL to 0.07 ng/mL). Plasma and whole blood concentrations showed good agreement. Storage did not affect cTnI concentrations (p > 0.75). Plasma cTnI concentrations had coefficient of repeatability of 0.02 ng/mL., Conclusions: The i-STAT(®)1 can measure cTnI in alpacas on both plasma and whole blood and provides similar values for both samples. Storage at 4 °C for 24 h or -80 °C for 30 days does not affect estimates of plasma cTnI. Evaluation of cTnI might be of value in assessing cardiac disease in this species., (Copyright © 2011 Elsevier B.V. All rights reserved.)