Characterization of extracellular matrix proteins and resident cells in calcified pulmonary arteries in horses.
Luis G. Arroyo Associate Professor | Lic. Med. Vet., DVSc, PhD, DACVIM

Recently, we observed fibro-calcification of the tunica media of large elastic arteries, predominantly of the pulmonary artery in greater than 80% of young racehorses in Ontario based on a post-mortem survey study. The lesions found in equine arteries resemble a well-described vascular pathology in humans involving the tunica media of major arteries. This lesion is the most important cause of stiffening of the blood vessel in humans and it has been established beyond reasonable doubt that there are strong independent associations between increased arterial stiffness or pulse pressure and increased morbidity, and mortality from cardiovascular disease. The potential clinical consequences of arterial calcification in horse’s unknown, but vascular degenerative lesions occasionally lead to vascular failure and result in death. Although the pathogenesis of this lesion is not fully understood, human and animal model studies suggest that an imbalance between calcification inhibitors and promoters in the arterial wall ultimately tip this balance to cause calcification. Ectopic calcification promoters (bone morphogenetic proteins and elastin degradation products) and inhibitors (matrix Gla protein (MGP), osteopontin and osteoprotegerin) have been found at the arterial calcification site. In addition, cells associated with the artery calcification site have osteo / chondrocytic morphological features. These histopathological similarities among different species suggested to us that there is a potential parallel of the pathogenesis process among species, although the triggering or risk factor (s) may be different and/or unrelated. The objectives of this study are to, first, investigate changes to the extracellular matrix proteins and whether proteins (i.e. collagen II, alkaline phosphatase) normally involved in bone development are present in calcified arteries of racehorses. Second, to characterize resident cells (osteo/chondrocyte-like cells) in tunica media of unaffected.

Characterization of the equine hindgut microflora and metabolites using an in-vitro model.
Luis G. Arroyo Associate Professor | Lic. Med. Vet., DVSc, PhD, DACVIM

The equine gastrointestinal tract is a large tubular structure that houses complex microbial ecosystems within multiple compartments along its length. The hindgut compartment (cecum and colon) comprises the larger portion of the tract and it is here where complex sugars are fermented by the resident microbes, providing between 60-70% of the daily energy requirements of the horse. Microbial communities (microbiota) disturbances may affect the health of the host leading to potentially life threatening disorders in horses, such as colitis and laminitis. The composition and function of the equine hindgut microflora is currently a topic undergoing intense research and much knowledge had been generated. However, a better understanding of the microbial ecosystem, in particular how disturbances in microbiota homeostasis change its function and the consequences of these changes to host health, is needed. In order to understand the effects of gut microbiota disturbances, it is imperative to know the status of the microbial ecosystem under healthy conditions. Culture of whole gut microbial communities in vitro, under physiologically relevant conditions, can be achieved by using continuous culture (chemostat) systems. The aim of this proposal is to develop an in vitro system that mimics the nutritional and environmental conditions of the hindgut of the horse, which (when seeded with GI samples obtained from horses) will allow analysis of the microbial population composition, abundance and distribution and their metabolic products.

Seroprevalence investigation and isolation of endemic strain (s) of Neorickettsia risticii (PHF) in the province of Ontario.
Luis G. Arroyo Associate Professor | Lic. Med. Vet., DVSc, PhD, DACVIM

Potomac Horse Fever (PHF) is a seasonal infectious disease caused by the bacterium Neorickettsia risticii. PHF occurs in Ontario mainly during the summer months (July to September). The clinical disease is usually sporadic with most of the cases reported from eastern and south-western Ontario. There severity of the clinical signs varies between horses, but in gereneral characteristic clinical signs include depression, decreased appetite, fever, diarrhea and colic. A high percentage of cases (up to 40%) of horses suffering from PHF may develop laminitis, which is the most serious and life threatening complication. Recent studies conducted on the life cycle of N. risticii have shown that fresh water snails and aquatic insects, such as caddisflies, mayflies, and dragonflies, are common carriers of the bacterium. Ingestion of contaminated hay, grain, pasture or drinking water with dead insects may result in clinical disease. Currently, there is an increasing concern among horse owners in Ontario because of the lack of information about the epidemiology and prevalence of the disease, natural reservoir of the bacterium, and biosecurity measures to prevent and control the disease. Preliminary data at the Ontario Veterinary College support clinical, serological and molecular evidence of the presence of the disease in different parts of the province. However, up to date there are only 2 published cases of PHF from horses in Ontario. The objectives of this preliminary study are to a)investigate the seroprevalence of PHF in horses from Ontario, and b) to isolate the endemic strain (s) of N. risticii from horses with clinical disease in Ontario for molecular analysis.

Plasma transfusions in horses: does it improve the clinical outcome?
Luis G. Arroyo Associate Professor | Lic. Med. Vet., DVSc, PhD, DACVIM

Equine frozen plasma is commonly prescribed for a diverse range of medical and surgical conditions. Veterinary textbooks dictate to clinicians that horses with hypoproteinemia require plasma transfusions. Plasma is generally administered to restore colloid oncotic pressure (COP), relieve edema, as a source of protein for nutrition support; to counteract endotoxemia and supplementation of immunoregulatory factors. Common clinical conditions in horses for which plasma is prescribed include chronic and/or acute hypoproteinemias as a result of protein losing enteropathies, intra/post-operative colic surgery and endotoxemias due to enterocolitis. The rationale for transfusions interventions in horses is poorly defined and there are very little clinical studies measuring the clinical impact (benefit) and outcomes (i.e. mortality/survival). This well accepted clinical practice has been subjected to minimal or null clinical research scrutiny, which is required to demonstrate effectiveness in equine medicine. In human medicine the only major clinical indication for plasma transfusions is to provide coagulation factors for the prevention or treatment of prolonged bleeding disorders. Plasma products are used however in a variety of horses despite the lack of evidence-based medicine to indicate any clinical benefit. In times of increasing cost consciousness and financial containment in the equine industry, since plasma transfusions are expensive and its clinical usefulness unknown, if not questionable, this clinical practice must be critically evaluated. This study aims to retrospectively evaluate the clinical uses, added treatment cost, adverse reactions associated and outcomes of plasma transfused cases at OVC. Prospectively, this project aims to evaluate the effects of plasma transfusions on COP of horses with hypoproteinemia and determine associations with clinical outcomes.

Effects of exercise and Lasix on the pulmonary artery hemodynamics of EIPH affected horses.
Luis G. Arroyo Associate Professor | Lic. Med. Vet., DVSc, PhD, DACVIM

Effects of exercise and Lasix on the pulmonary artery hemodynamics of arterial wall remodeling (calcification and fibrosis) occurs in racehorses and is the most important cause of vascular stiffness in humans. There is a growing body of evidence to support the hypothesis that excessive pressure pulsatility resulting from arterial stiffness promotes abnormalities in micro-vascular structure and function, contributing to end-organ damage and dysfunction in humans (Mitchell GF 2008; Tan W et al., 2013). We hypothesize therefore that stiffening of the pulmonary arteries of racehorse’s leads to hemodynamic changes (excessive pressure pulsatility) promoting micro-vascular structural and functional abnormalities in the lungs, and that this contributes to the development of EIPH. Our research group is a multidisciplinary team of equine and human clinicians, pathologists and bioengineers that for many years has been conducting equine pulmonary mechanics research (material properties, wall stress, pulse wave quantification, in vitro functional testing). We have access to specialized research equipment and clinical facilities necessary for this type of research project and we have already developed both innovative lab and clinical testing methods and analysis strategies. We aim to investigate the compliance of the pulmonary arterial wall of horses with arterial remodeling (i.e. fibro-calcification), the hemodynamics of the pulmonary artery blood flow, and the potential role of these alterations in the pathogenesis of EIPH in horses.

 Pathogenesis and characterization of inflammatory airway disease in Ontario Thoroughbred and Standardbred racehorses
Janet Beeler-Marfisi, Assistant Professor, Pathobiology, BA, DVM, DVSc, Dipl. ACVP

Inflammatory Airway Disease is a type of lung inflammation in horses that is similar to mild asthma in people. It is common in racehorses, especially 2 and 3 year olds that are stabled, but the asthma is not caused by an infection and the horse's temperature remains normal. The asthma causes the horse to have a persistent cough, which is made worse by exercise. We think it is partly caused by breathing in barn dust from hay, bedding and the floor (during sweeping). In part, barn dust is made up of very fine mould spores (even from good looking hay), and little pieces of dead bacteria (from manure). Both of these things cause the horse’s lungs to react and that reaction causes the asthma. Another thing we think causes horse asthma is air pollution. It is true that in Ontario air pollution causes asthma in people, but we haven’t studied whether it does the same in horses. It’s important to study this disease because it is costly to treat and means missed training and racing days, so the earnings potential of the horse is decreased. In this project, we will work with Thoroughbred and Standardbred trainers and veterinarians at Woodbine and Mohawk racetracks. When a trainer has a horse that they think is coughing bad enough to call their veterinarian, the vet will do a physical examination on the horse and will use their scope to look in the horse’s windpipe and down into the lungs. The vet will then perform a ‘lung wash’ on the horse. This is a safe procedure where they put a small amount of sterile saline into the lung and suck it right back out. It doesn’t fix the problem, but it allows us to look at the cells and mucus in the lungs and tells us what the horse has wrong with it (asthma, lung infection, or bleeding). Based on what we find we will be able to work out approximately how many racehorses have asthma, and we will see whether air pollution, the mould spores and pieces of bacteria cause asthma. If we can get scientific evidence of these things we will be able to help racehorses to be healthier and improve their earnings potential to the benefit of all involved.

Critical gene expression in the uterus and conceptus during successful and failing equine pregnancies.
Keith J. Betteridge Professor Emeritus/Emerita (University) | BVSc Bristol, MVSc Toronto, PhD Reading, FRCVS

Timely foaling of brood mares is very important to the racing industry with its fixed ‘birthdays’ and can be disrupted by early pregnancy loss. Our aim is to identify key mechanisms by which the mare and her conceptus (the embryo, its membranes and contained fluids) interact successfully in early pregnancy, and how these change when pregnancy fails. We have found pregnancies to fail spontaneously either between Days 12-16 after breeding when the conceptus should be ‘recognized’ by the mare (so that she does not return to heat), or between Days 19-25. The latter stage coincides with deterioration and loss of the embryonic capsule (a non-cellular membrane) that envelops the conceptus and, we believe, shields it from potentially hostile uterine products as pregnancy is established. Experimentally, pregnancy can be terminated by injecting the mare with a prostaglandin (PG) which lowers progesterone concentrations. This is accomplished more immediately by PG treatment at Day 18 than at earlier stages. Recently, we have compared gene activity in the endometrium (lining of the uterus) of mares undergoing spontaneous and PG induced failures with activities found during normal pregnancy. For this we use an advanced molecular technique (transcriptome-wide sequencing of RNA) at Day 20, and have similarly examined normal pregnancies at Days 14-15.

Ciliary function in horses with inflammatory lung disease
Dr. Dorothee Bienzle, Pathobiology MSc, PhD, DVSc

Cilia are small hair-like extensions on cells that line the nose, trachea and larger airways. The function of cilia is to move a layer of mucus produced by cells in the lower airways toward the mouth for swallowing, and therefore to protect the lung from inhaled particles. Thus, cells with cilia that constantly beat to move a layer of mucus “outward” line the entire upper airway. Inhaled bacteria, molds and particles “stick” to the mucus layer and are then moved by beating ciliary toward the mouth. People and animals born with cilia that are defective in the proteins necessary for ciliary movement (“primary ciliary dyskinesia”) have recurrent pneumonia throughout their life as well as abnormalities in other organs. In conditions such as asthma and heaves, recurrent inflammation of the airways results in injury to cells lining the airways, including ciliated epithelial cells. It is thought that in these conditions cilia do not regenerate properly, leading to a vicious cycle of poor clearance of particles by shortened or poorly beating cilia and recurring inflammation. Viral infections of the nose and lungs have also very recently been recognized to affect the function of cilia. We will collect epithelial cells from healthy horses’ noses with a simple short medical brush, put the cells into culture and measure how well they move particles mimicking bacteria (fluorescent beads). We will compare the results from nasal samples to tracheal samples obtained with a bronchoscope and a longer medical brush. If cilia in samples from the nose function similar to those from the trachea, routine samples for assessments can be readily acquired. Once the functional assays are established we will collect samples from young and older horses with mild and severe lung inflammation (“inflammatory airway disease”, IAD, and “heaves”), respectively, to measure how cilia function in these conditions and to assess their appearance. If abnormal cilia function is a consistent feature of respiratory diseases of horses, inhaled medication to change the composition of mucus and/or to increase the production of mucus may be helpful.

Sustained release drug formulations for equine joint conditions: Formulation and forensic considerations
M.B. Hurtig, Professor, D.V.M., M.V.SC. (SASK.), DIP. A.C.V.S.

In past studies we have studied long acting drugs that were created by a drug company for the human market to control inflammation in joints for up to 90 days. Using the sheep knee as a model we showed that these drug and polymer combinations take up residency in the joint lining where they are slowly broken down to release the drug and control pain and inflammation. The duration and drug levels in joints can be changed by altering the polymer-drug binding, so release times of a few days to several months are possible. Through a collaboration with polymer chemist Dr. Elizabeth Gillies (Western University, Ontario) we will create non-steroidal anti-inflammatory drug (NSAIDs) formulations that will last from a few days to several months after injection into joints. The advantage of this delivery system is that no drug leaks from the joint into the rest of the horse's body to cause the all too common complications of colic and gut ulcers. Dr. Gillies' role will be to create the polymer carriers from safe, food grade chemicals. These are widely used in human sustained release drugs for the eye conditions, cancer therapies etc. Dr. Gillies will modify NSAID drugs so they can be trapped in the polymer carrier system. Laboratory experiments will be used to estimate the rate of drug release into joint fluid over 21 days. With this information we will conduct trials of sustained release NSAID formulations by injecting them into the fetlocks of horses at the University of Guelph Arkell Research Station. By injecting the drug and then challenging the horse joint with an inflammation-causing molecule (lipopolysaccharide) at intervals of up to 90 days, we will be able to determine the duration of the drugs' anti-inflammatory effect. Many horses have low grade inflammation in joints that is difficult to address and requires repeated, costly treatments. Such medication regimes are expensive and not without risk, particularly when steroids are used since they may promote cartilage loss over time. An additional consideration is the potential for doping with similar drug formulations. By measuring drug levels in plasma, urine and joint fluid we will be able to estimate when drugs would be detected in doping tests. We will share all our data with racing and clean sport/doping jurisdictions and provide samples of our drug formulations for their own use. This project will provide the basis for commercialization of a sustained release formulation that would improve the safety of NSAID use in the horse and potentially reduce the incidence and rate of osteoarthritis progression in horses.

Development of Osteochondral Constructs Using Equine Umbilical Cord-Derived Mesenchymal Stromal Cells for Treating Joint Cartilage Defects
Thomas G. Koch Associate Professor | DVM Copenhagen, PhD Guelph

Co-investigators’ names and Departments: Rita A. Kandel, M.D., Department of Pathology, Mount Sinai Hospital, Toronto, ON, Mark Hurtig, Clinical Studies, OVC, Judith Koenig, Clinical Studies, OVC, Ms. Sarah Lepage, PhD Candidate, Biomed Sci, OVC

Orthopedic injuries, constituted mainly of trauma to joint cartilage, are the most common cause of lost training days or premature retirement in the equine athlete. As cartilage tissue has a low intrinsic capacity to heal, repeated injury will eventually result in post-traumatic osteoarthritis in the joint. Though current regenerative therapies for the treatment of focal cartilage defects are showing promising preliminary results, they are wrought with potential complications from secondary surgical sites to isolate patient-specific cells or tissue. Therefore, we propose to investigate the potential of a novel cell type in the generation of osteochondral-like plugs without requiring a secondary surgical site on the patient. We have successfully and reproducibly isolated mesenchymal stromal cells from umbilical cord blood (CB-MSC) and demonstrated their ability to generate cartilage in vitro. We found that by subsequently growing the engineered cartilage on a bone substitute, we could generate an osteochondral construct that could serve as an implant. In 2006, Dr. Rita Kandel’s group in Toronto generated similar constructs and implanted them into sheep with induced cartilage injuries; the implants successfully contributed to cartilage repair at 9 months post-surgery. We believe that with further optimization, we can generate an osteochondral-like implant using CB-MSC with superior properties for in vivo repair of focal cartilage defects. The biomechanical properties of cartilage generated in vitro are historically poor, making the graft susceptible to damage upon transplantation. We propose to mechanically mature the cartilage prior to implantation in order to better withstand the large loading forces within the equine joint upon transplantation.

Allogeneic Equine Umbilical Cord Blood Mesenchymal Stem Cells for Treatment of Exercise Induced Pulmonary Hemorrhage
Thomas G. Koch Associate Professor | DVM Copenhagen, PhD Guelph

Exercised induced pulmonary hemorrhage (EIPH) is a common condition in race horses and a significant equine welfare issue. The etiology of EIPH is poorly understood but pulmonary inflammation is a downstream consequence of frequent bleeding. Mesenchymal stem cells (MSC) have non-progenitor functions through secretion of various proteins. Through such secreted factors MSC have demonstrated immune modulatory properties in vitro and in vivo. In our lab we have demonstrated that equine umbilical cord blood-derived (CB) MSC are more immune-modulatory than the more commonly used MSCs derived in vitro from adipose tissue or bone marrow aspirates. We have also shown that CB-MSC suppress lymphocyte proliferation in vitro equally immediately after thawing or after one week of culture expansion post-thawing. These CB-MSC have low to no expression of MHC-I and II, suggesting they can be applied in an allogeneic manner. In vivo studies in the horse may lend support to allogeneic application of CB-MSC. Attenuation of pulmonary inflammation from bleeding following intravenous MSC injection has been shown in vivo in humans and non-equine species. The hypothesis of this project is that allogeneic equine CB-MSC will attenuate EIPH associated pulmonary inflammation. Funds are sought to perform a feasibility study to test this hypothesis. Use of allogeneic CB-MSC holds the promise of developing a novel off-the-shelf cell-based product for this important equine disorder.

Comparison of umbilical cord tissue and cord blood stem cells
Thomas G. Koch Associate Professor | DVM Copenhagen, PhD Guelph

Equine mesenchymal stromal cells are increasingly explored as novel therapeutic agents in equine medicine for a variety of conditions. The field is increasingly focused on using these cells in an allogeneic manner due to ease of application and reduced cost of cell production. Cells from neonatal tissues have higher proliferative potential than similar cells derived from adult tissues. The use of cells with high proliferative potential is essential in order to leverage the potential advantages of using allogeneic cells. However, extended culture expansion may result in cellular drift with regards to phenotype, function and cytogenetic stability (normal chromosome numbers and sizes). The Koch lab has extensively studied equine umbilical cord blood-derived mesenchymal stromal cells and found the cells to have higher proliferative potential than similar cells from bone marrow and adipose tissue. One limitation of cord blood cells is the collection of the umbilical cord blood and the relatively few progenitor cells present in the blood. Advanced foaling management is required at the farm level and trained personnel have to be present during foaling to obtain the blood. Extensive laboratory cell expansion is needed due to the few progenitor cells present, which is associated with increased laboratory selection pressure on the cells. Mesenchymal stromal cells have been isolated from the connective tissue of equine umbilical cords. This cell source is easier to obtain and a higher number of putative progenitor cells has been reported. However, it is unclear if mesenchymal stromal cells from cord tissue and cord blood are equivalent with regards to phenotype, proliferation, function and cytogenetic stability. The goal of this project is to compare donor-matched mesenchymal stromal cells from cord tissue and cord blood with regards to the above-mentioned parameters. The influence of oxygen tension on these cell parameters will also be evaluated for both cell sources.

Evaluation of the significance of arthropathy of the caudal facet joints in horses.
Judith Koenig, Dr Med Vet, DVSc, Diplomate DACVS, Diplomate ECVS

Arthritis of the neck joints is commonly present in performance horses and in many cases causes debilitating clinical signs. In older horses osteoarthritis of lower neck vertebra facet joints is classified as Type-2 Wobblers syndrome. Clinical signs include neck stiffness, neck pain, reluctance to collect, and forelimb lameness without neurological deficits. In more severe cases the osteoarthritis causes compression of the spinal cord and neurological deficits can be present. There are conflicting reports in the literature as to the significance of diagnosing osteoarthritis of lower neck vertebra facet joints on radiographs, the age at which it becomes clinically significant, how often it causes spinal cord compression, and the success rate of long-term treatment. Therefore, our objectives are to quantify degenerative changes of lower neck vertebra facet joints and to obtain inter and intravertebral measurements of the neck vertebrae on radiographs of horses with clinical signs compatible with this disease and age, breed and sports matched healthy Warmblood horses. Follow up information will be gathered by telephone survey of owners on the outcome of intra-articular treatment of osteoarthritis of lower neck vertebra facet joints to evaluate the effectiveness of this treatment method.

Evaluation of the repeated use of allogeneic umbilical cord blood mesenchymal cells in clinical cases of equine superficial digital flexor tendinitis.
Judith Koenig, Dr Med Vet, DVSc, Diplomate DACVS, Diplomate ECVS

Tendon and ligament injuries are common in the equine athlete. These types of soft tissue injuries result in significant economic loss to the equine industry each year as a result of decreased performance, prolonged rehabilitation, and recurrent injury. Even though tendons heal well, the scar tissue formed in this repair is functionally deficient in comparison to normal tendon. The result is compromised performance of the horse and high re-injury rates. Improved clinical lameness outcome and a better overall prognosis for return to exercise was noted upon injection of simple bone marrow aspirate directly into damaged suspensory ligaments. This observation spurred the interest of biologics for the treatment of equine tendon injuries. Concerns related to the volume of unaltered bone marrow and its content of bone spikes and fat cells on tendon healing lead to the use of cultured stem cells derived from bone marrow (BM) aspirates. Such BM stem cells have been used commercially to treat injured tendons in more than 500 horses with naturally occurring injuries. Only 18% of these horses were re-injured once they entered full training, which appears more favorable then a 56% re-injury rate reported in a previous study where a routine rehabilitation protocol was used. This much improved reinjury rate of only 18% was only observed when the stem cells were injected in the first 6 weeks after injury. Bone marrow derived stem cells need to be culture expanded in the lab after harvest from the horse’s bone marrow, which can take 4 weeks. This requires very prompt action after injury to ensure a diagnosis is established, bone marrow is harvested from the horse and transported to the lab for expansion and back to the veterinarian to be injected within 6 weeks after injury to achieve the reported benefits. We have shown previously, that it is safe to inject allogeneic (unrelated horse) equine umbilical cord blood stem cells into horse’s joints and that equine umbilical cord blood stem cells are more immune-modulatory than stem cells from equine bone marrow, even after prolonged freezer storage. These findings support the cell banking of umbilical cord blood stem cells for later allogeneic use, which means they can be taken off the shelf and injected into horses as needed without having to wait for culture expansion.

Effect of extracorporeal shockwave therapy on umbilical cord blood mesenchymal stromal cells
Judith Koenig, Dr Med Vet, DVSc, Diplomate DACVS, Diplomate ECVS

Shockwave has been used as a treatment to decrease the healing time and improve the quality of the repair in bone and soft tissue injuries, but the exact mechanism is not completely understood. Intra-lesional (into bowed tendons) administration of stem cells in combination with shockwave has been used as an alternative treatment for orthopedic diseases like tendon and ligament lesions in horses, without scientific evidence, but anecdotally has resulted in faster healing times and return to function. There is a need to evaluate the effect that shockwave has on stem cells. One in vitro study showed that equine fat derived stem cells grow more rapidly and differentiate faster into different tissues, but nothing is known about the secretory function of these cells or the effect of shockwave on bone marrow or umbilical cord blood derived stem cells. Labelling these cells allows tracking of these cells also in vivo. Therefore, we hypothesized that shockwave will enhance the progenitor (their ability to differentiate into different tissue) and nonprogenitor (their anti-inflammatory and secretory) functions of stem cells. Our objectives are to firstly evaluate if shockwave increases umbilical cord blood derived stem cell growth rate, proliferation rate and differentiation in comparison to untreated cells in vitro. And secondly, to evaluate the effect of shockwave on topically injected allogeneic (derived from unrelated foals) umbilical cord blood derived stem cell and follow the cells in a neck wound model in vivo.

Equine Herpes Virus (EHV) and EHV-associated disease in the Ontario Equine Industry - Disease prevalence and prevention
B. Lillie, Assistant Professor, D.V.M., PH.D (GUELPH), DIP. A.C.V.P.

Equine herpesvirus-1 (EHV-1) can cause a variety of diseases in horses, from simple runny nose and fevers, to abortion or even neurologic disease and death. We are hoping to better understand abortion rates in Ontario broodmares and determine what effect EHV-1 is having on the industry. One aspect of the virus that was overlooked until recently, is that many horses become infected when they are still with their dams. Once a horse is infected with the virus, it can remain in the body and can reactivate at any time. Many characteristics of this viral infection are unknown, such as how many horses in a population are infected, when or why it reactivates within the body, and why it causes one form of disease over another. EHV-1 vaccines have been thought to be the reason for a steep reduction in abortion. However, studies have found that the vaccines are not completely protective, outbreaks occur even in vaccinated herds and over-vaccinating can actually decrease the body’s protective immune response. Considering that the acceptable rate of abortion is not known, it is difficult to gauge the effect that EHV-1 has on infected mares and therefore the value of vaccination. Vaccinating them after they are infected has not been strongly studied, so the effects of the vaccine to prevent disease in already infected horses is unclear. Further, no vaccines have been labelled protective against the neurologic form. Researchers have recently found a genetic change in the virus from horses.

Nutritional and feeding behaviour effects of steaming or soaking hay for horses
Katrina Merkies, Associate Professor

Dried hay is the most common feedstuff for horses in North America. Horse owners supply hay in a variety of formats for a variety of reasons. Two such methods in which dried hay is altered before being offered to horses is soaking and, to a lesser extent, steaming. When grass hay is soaked or steamed the resulting wet or steamed hay has less carbohydrates. Soaking, but not steaming, also results in a loss of protein and micronutrients. A lower carbohydrate-content hay can help to reduce blood glucose and this is appealing for horses with laminitis, insulin resistance (Cushings), Equine Polysaccharide Myopathy, and those who are overweight. The extent of the effects of soaking and steaming on timothy/alfalfa hay, the most commonly available hay in Ontario, is unknown as the majority of research has been conducted in the United Kingdom where grass hays are more common. Therefore, under commonly-used soaking and steaming practices, this research seeks to understand how soaking and steaming affect the nutrient content of timothy/alfalfa hay and additionally whether soaking or steaming alters the preference horses have for dried, soaked or steamed hay.

Comparing loading between the forelimbs of racing Thoroughbred to investigate possible causes of laterality in catastrophic injury
Jeffrey J. Thomason, Professor | BA, MSc, PhD Toronto

The U.S. Jockey Club’s Equine Injury Database (EID) has information on more than 1.6 million racing starts by Thoroughbreds in North America. A recent survey of the approximately 3000 catastrophic injuries (CI) appearing in the database confirmed that the frequency of CI is related to the surface type, being greater on dirt, intermediate on turf and least on synthetics. In addition the study found that the left fore and right hind limbs were significantly more prone to CI on dirt tracks than the opposite limbs, but this effect was not evident on synthetic or turf tracks. The researchers speculated that this laterality of injury might be due to uneven forces acing on the left and right limbs in turns combined with relative inconsistency of support during the stance on dirt. These findings are significant for Ontario’s TB racing industry because races are on turf or synthetics while most training is on dirt. The aims of this study are to assess whether laterality of biomechanical loading exists on dirt and synthetic surfaces, and whether epidemiological data identify that as an issue in Ontario. We will tackle this project in 4 ways: (a) We’ll collaborate with the researcher currently studying the Ontario Death Registry, to look for evidence of injury laterality. The emphasis will be on the forefeet, which sustain higher rates of injury. (b) We will take another look at data we have from previous experiment on forces acting on the feet of Thoroughbreds galloping on the straight and turns of different tracks, and will look for evidence of laterality in loading at all stages of the stance for both front feet. We have data from dirt, turf and synthetic surfaces, using the same horse but on different days. (c) We will record new biomechanical data from the front feet of horses galloping on a training dirt track and a racing synthetic track within a few minutes, to find whether and how much laterality of loading exists on each surface. (d) We combine the analyses from all of the data from parts a-c to assess whether laterality is a problem in Ontario, and to categorize which parts of the complex mechanical loading acting on the horse’ hoof during galloping might be at the root of the problem. The results should identify whether turning or the surface induces the higher risk for injury, from a biomechanical perspective.

Characterization of the epidemiology and clinical impact of Clostridium difficile infection in hospitalized foals
Scott Weese, Professor | DVM, DVSc Guelph; Dipl ACVIM

Co-investigators’ names and Departments: Nathan Slovis DVM, Dipl. ACVIM and Anne Kullmann DVM, Hagyard Equine Medical Institute (HEMI), Diego Gomez-Nieto DVM DipACVIM, Dept of Pathobiology

Neonatal foal diarrhea is a common problem in foals, and Clostridium difficile is an important cause. In Central Kentucky, there has been an increased recognition of C. difficile (CDI) infection in the foals on farms and at an equine hospital, with the latter characterized initially by a subclinical infection that develops clinical disease usually after 48 hours of hospitalization. Disease that is different from classical enterocolitis is consistent with recognition in humans that CDI is not always accompanied by overt diarrhea. Preliminary data have indicated 7 different ribotypes from nine C. difficile clinically affected foals, something that is consistent with multiple community sources rather than a hospital focus. This prospective case control study will therefore evaluate the prevalence and epidemiology of Clostridium difficile colonization in hospitalized foals versus farm-matched controls, evaluate the impact of C. difficile colonization on development of disease during hospitalization, characterize C. difficile recovered from foals, evaluate diagnostic tests for CDI and evaluate the transmission of C. difficile between mares and foals. It will involve collection of fecal samples from foals presented to two equine hospitals (Kentucky, Ontario) for reasons other than diarrhea, plus their mares, age and farm-matched controls and foals with disease potentially attributable to CDI. This study will provide important information about the nature of this important pathogen, the risks associated with C. difficile shedding at hospital admission, transmission of C. difficile between mares and foals and risk factors for disease, with a goal of better understanding the disease to better diagnose, treat and prevent infections.

Development of 3D printed models of the sacroiliac, hip, and cervical articular facet joints for ultrasound guided joint injections training of equine practitioners
Alex zur Linden, Assistant Professor | DVM, DACVR

Three-dimensional models are used to teach various skills and techniques without needing cadavers or practice on live patients. Ultrasound guided injections of the neck, hip, and sacroiliac joints are challenging to perform but very useful for diagnosing and treating arthritis in sports and racehorses. Joint injections are used for diagnosing lameness by injecting local anesthetics, or treating arthritis by injecting steroids or other similar substances. These specific injections are often only performed by radiologists or those with advanced training and access to ultrasound machines. With the increased use of ultrasound by equine veterinarians for the diagnosis of tendon injuries in racehorses, this imaging modality can also be used for diagnostic and therapeutic joint injections. One goal of this project is to determine the proper 3D printed material to mimic equine bones via ultrasound examination. The next step is to scan various body parts of a cadaver (neck, sacroiliac joints, and hips) with computed tomography (CT) to create highly accurate digital versions of these joints. These body parts will then be 3D printed in high resolution and embedded in clear ballistics gel to mimic tissue, while allowing the user to directly see the needle and underlying bones. The models will then be assessed for validity by radiologists, equine surgeons, and residents at the Ontario Veterinary College. Once validated, these models will be used to teach interested equine veterinarians in Ontario how to perform these techniques via seminars.