Electrocardiographic ventricular repolarisation processes in Andalusian horses before and after physical training.

Equine ventricular repolarisation wave can be influenced by many physiological and pathological factors. T wave abnormalities have been related to a shorter time for ventricular filling, reduced stroke volume, cardiac output and exercise performance in racehorses. The present research performed electrocardiographic recordings in 14 four-year-old male Andalusian horses, when they were untrained and after three months of an aerobic training programme. Leads I, aVF, V10, V1R, V3R, V1L and V3L were used. It was aimed to assess the incidence of abnormal T waves in this breed, according to the criteria of abnormalities established for other equine breeds, to evaluate if the changes induced by training in the T wave are the same that those reported in the athletic heart syndrome in canine and human athletes and, to analyse if the abnormal T waves could have been related to changes in the plasma concentrations of Na, K, and Cl, since these electrolytes are involved in the cardiac electrical processes. It was found out that the incidence of abnormal T waves was quite high in the Andalusian breed, especially in the precordial leads. Moreover, the incidence of abnormal T waves increased in precordial leads and decreased in lead I and aVF after training. The abnormal T waves after training were shorter and had the same voltage, changes which were different to those presented for the athlete¿ heart syndrome. The horses with abnormal T waves had higher plasma K concentrations, both before and after training. Plasma Na and Cl concentration at rest decreased after training. Plasma Na concentrations were positively related to T wave duration and negatively to T wave voltage. Key words: electrocardiography, heart, horses, performance, trainin

Electrocardiographic exam in Arabian horses submitted to exercise on high-speed treadmill and vitamin E supplementation

Avaliou-se o efeito do exercício físico, treinamento e suplementação com vitamina E sobre os parâmetros eletrocardiográficos, utilizando oito éguas da raça Puro Sangue Árabe, sem treinamento prévio, distribuídos em dois grupos: controle (n=4) e suplementado com vitamina E (n=4) na dose diária de 1.000 UI. Os animais foram submetidos a uma prova de exercício progressivo (P1) em esteira de alta velocidade inclinada a +7%, em seguida a um período de treinamento de 20 dias e posteriormente uma nova prova de exercício progressivo (P2). Realizou-se a análise e interpretação dos traçados eletrocardiográficos quanto ao ritmo, frequência cardíaca, duração e amplitude das ondas e intervalos, antes, imediatamente após e 30 min após o exercício progressivo. Observou-se uma diminuição da frequência cardíaca de repouso após o treinamento. Não houve efeito da suplementação com vitamina E sobre os parâmetros eletrocardiográficos. Os resultados indicaram que o eletrocardiograma foi eficiente na detecção das alterações cardíacas promovidas pelo exercício físico, mas necessita-se de mais estudos para elucidar o seu significado clínico.The effect of physical exercise, training and vitamin E supplementation on electrocardiographic parameters was evaluated in eight untrained Arabian mares, divided into two groups: control (n=4) and supplemented with vitamin E (n=4) at the daily dose of 1.000 UI. Animals were submitted to an incremental exercise test (P1) on high-speed treadmill inclined +7%, after that to a training period of 20 days and later to a new incremental exercise test (P2). Analysis and interpretation of electrocardiographic tracings were performed regarding the rhythm, heart rate, duration and amplitude of waves and intervals, before, immediately after and 30 min after the incremental exercise. A reduction of the rest heart rate was observed after training. There was no effect of vitamin E supplementation on electrocardiographic parameters. Results indicated that the electrocardiogram was efficient in detention of cardiac alterations promoted by the physical exercise, but more studies are needed to elucidate its clinical meaning

Bibliographic survey of the parameters used to perform evaluation and physiology of exercise in horses / Levantamento bibliográfico dos parâmetros utilizados para avaliação de desempenho e fisiologia do exercício em equinos

This literary review aims to gather technical information on the parameters analyzed as a result of equine sports medicine and exercise physiology. These parameters are divided into physical, hemodynamic, and biochemical. Within these three, heart rate (HR), respiratory rate (RF), electrocardiography, erythrogram, leukogram, creatine kinase (CK), lactate, aspartate aminotransferase (AST), lactated dehydrogenase (LDH), glucose and electrolytic salts (sodium, potassium, phosphorus, and calcium). Today, we seek to understand the dynamics exercised by horses during the year, which makes the growth of research in this area increasingly greater. However, crucial details are still lacking for certain reference standards to be accepted, and the idea of the different actions of the body from equine athletes to non-athlete horses is raised

Comparative Medicine: investigations into the fields of infectious and zoonotic disease research, and population-level epidemiology

Project 1: Human African Trypanosomiasis causes cardiac rhythm disturbances through hitherto unknown mechanisms. It is known that the presence of such protozoan parasites or simply their supernatant has excitatory effects on isolated cardiomyocytes, seen as increased Ca2+ wave occurrence. We investigated the theory that the protozoa are able to release a cathepsin L-like product into solution which is the initiator of such waves. Adult rat left ventricular cardiomyocytes were isolated in solution before [Ca2+] was raised to physiological extracellular concentration. The cells were then exposed to pH controlled samples of supernatant, with or without a known cathepsin L-like protease inhibitor (K11777), and all were subjected to population studies of wave incidence over 60 second timeframes. Baseline percentage of spontaneously waving cells in mock extracellular solution was found to be significantly lower than with the presence of Trypanosome supernatant. Addition of K11777 led to a significant reduction in spontaneity toward baseline. These results suggest that T. b. brucei produces a cathepsin L-like cysteine protease which causes increased incidence of Ca2+ waves. Project 2: Two survey-based studies of equine disease in the UK gathered thirty years apart were compared on the basis of disease category prevalence. The first study, published in 1965, was conducted by the British Equine Veterinary Association and covered the whole of mainland UK and Ireland. The second dataset was obtained from The Home of Rest for Horses Study, which surveyed Scotland and northern England in the mid-1990s. Musculoskeletal and alimentary diseases were the most important categories of disease in both studies, but there were significant differences seen in both the composition of the horse population in terms of breed type, and the profile of disease between the studies. The predominant horse type in the BEVA study was the pony, whereas the HRH study reported a more even distribution of ponies, hunter-type horses and racehorses within the population. Disease categories which were more commonly diagnosed in the HRH versus the BEVA study included musculoskeletal and systemic disease. Those categories which had were less commonly diagnosed comprised alimentary and respiratory disease. Project 3: Diarrhoea is the second most common cause of death of children under 5 years of age in the developing world, and accounts for more child deaths than measles, malaria and HIV combined. An important cause of such enteric disturbances is infection with pathogenic organisms, many of which may be acquired through zoonotic transmission from domestic animal species. Detailed epidemiological information regarding the causative agents implicated in diarrhoeic disease in western Kenya is lacking. The Kenyan Medical Research Institute (KEMRI) in collaboration with the Centres for Disease Control and Prevention (CDC) began conducting a case-control study of diarrhoeic children in western Kenya in 2008. Ninety seven and a half percent of compounds housed chickens, 75.9% housed dogs and 74.7% kept cattle, other species were less common. Two point four percent of pooled animal faecal samples were classified as diarrhoeic faeces, 66.7% of which were from young animals. The following pathogens were found in animal faecal samples at the stated prevalences (all animal species sample results combined); E. coli 99.3%, Salmonella enterica 6.6%, Campylobacter jejuni 1.8%, Campylobacter coli 0.7%, Salmonella typhi 0.2%, Giardia spp. 10.2% and Cryptosporidium spp. 2.5%. In stool samples from children, the following pathogens were found to be marginally more prevalent in cases versus than controls; C. jejuni, C. coli, S. enterica, and Cryptosporidium. Giardia spp. were more prevalent among control children than cases. No statistically significant associations were found between the presence of any of the identified pathogens in the animal population of a compound with its presence in the child samples of the same compound

Stress in equids undergoing veterinary care and the development of interventions that positively influence the horses' experience

Veterinary care can be essential for maintaining the physical health of horses and yet the impact of veterinary care on a horse’s emotional state and how that influences their behavioural responses has not been investigated previously. The term horses is used to describe both horses and ponies. Adverse behavioural responses are unwanted, not only for equine welfare but because veterinarians are frequently injured by the horses they are working with, each sustaining on average around 7-8 significant injuries over the courses of their careers. This study evaluated which aspects of veterinary care horses find stressful, their common behavioural responses in this context and potential ways of improving how they perceive veterinary care. In the context of a veterinary examination, horses are limited in which behavioural responses they can express as they will be restrained to a greater or lesser degree. To determine the range of behavioural responses that may occur and their usefulness in assessing emotional state in the veterinary clinic, a two round Delphi process was undertaken. In the first-round participants from the field of Equitation Science (n=16), Equine Veterinarians (n=10) and Animal Behaviour and Welfare Scientists (n=7) evaluated 11 videos of horse’s behavioural responses to veterinary care. They assigned a score to the degree of stress they perceived each horse to be experiencing and described any behavioural indicators of stress as free text. There was poor agreement on the perceived degree of stress the horses were experiencing and different behavioural indicators were reported by different individuals. The behavioural indicators (41) identified in round one were then re-presented to the participants in a second round where 34/41 indicators were considered useful measures of the horse’s stress response in this context. A feasibility study was then undertaken to determine the best measures of arousal and valence in the context of a veterinary clinic. Horses presenting for veterinary care at an equine hospital (n=30) were filmed during various veterinary interactions (Events) with the horses and the footage was evaluated using a detailed inventory of behavioural responses. Principle component analyses were performed, results of which suggested that the most common behavioural responses to veterinary care were characterised by freeze or flight. Evaluation of the feasibility of different measures of arousal (salivary cortisol, surface eye temperature and heart rate variability) were undertaken on the same cohort of horses. Salivary cortisol concentrations highlighted that horses were in a state of elevated arousal, likely as a consequence of transportation, even before veterinary care commenced. Elevated cortisol concentrations were maintained, most likely due to exposure to veterinary Events, but the lag between an individual Event and concentration peaks / return to baseline meant this was not feasible measure in this context. Surface eye temperature provided data that varied in real time and showed variation between horses and between individual Events. The main limitation of using this measure was the practical challenges trying to capture usable, in focus, and therefore accurate, thermal images of the eye as many horses did not remain still during Events. Heart rate variability parameters (HRV) provided data that varied in real time and showed variation between horses and between individual Events. Moreover, it could be collected remotely and regardless of any behavioural response. In the feasibility study HRV data were collected using a Polar heart rate monitor. However, validation against an ECG showed the Polar to be inaccurate, even following artefact correction. Even when an ECG was used subsequently, manual R peak detection and documentation of arrhythmias was essential to ensure accurate data for HRV analyses were generated. The methodology developed in the feasibility study was used to evaluate the responses of 60 horses undergoing veterinary care in the same equine hospital. The proportion of time a horse was ‘stressed’ was calculated based on the sum total of time in any negative affective state over total time. This study found marked variation regarding which veterinary Events horses appeared to find stressful, both within and between individuals. For example, 13/15 veterinary Events resulted in a range of time an individual horse was scored as ‘stressed’. Negative behavioural responses were characterised by freeze or flight but also fidgeting behaviours, which are well described in other species but poorly documented in horses. Aggressive ‘fight’ responses occurred very infrequently. Further research is required to determine whether training of equine veterinarians to recognise subtle behavioural indicators, across the range of responses (freeze, flight, fidget or fight), can be used to improve equine welfare and reduce the likelihood of occupational injuries. With regard to evaluation of individual Events: • One of the most arousing stimuli, based on behavioural response and elevated heart rate, in the veterinary environment was the sound or sight of another horse walking past and so minimising this stimulus where possible is recommended. • Horses are frequently walked onto a weigh bridge upon entering the equine hospital and this commonly evoked a behavioural response characterised by freeze or flight and elevated heart rate. It is less than ideal that their first experience in a novel environment and first interaction with staff may be a negative one. Weighing a horse after other veterinary Events have been completed or development of a low stress technique would likely be of benefit. • The use of feed during diagnostic local anaesthesia appeared to minimise the proportion of time an equid was scored as ‘Stressed’ compared to when it was not used and so is recommended. • Horses were scored as more ‘Stressed’ when an I.V. injection was administered by a student or nurse in comparison to a veterinarian. Investigation of training to try to mitigate this response should be undertaken. The final study in this thesis evaluated the way in which learning theory could be applied in veterinary practice. Classical counter conditioning was used prior to diagnostic local analgesia (nerve blocks) and the impact on the horse’s stress response was investigated. Twenty-seven cases presenting for lameness evaluation were recruited and randomly assigned to a control or treatment group. In the treatment group the provision of feed (unconditioned stimulus) was paired with someone approaching (conditioned stimulus) the limb. In the control group 14 horses received 24 nerve blocks, in the treatment group 13 horses received 35 nerve blocks. Restraint during the nerve block was at the discretion of the veterinarian performing the procedure. Video footage of horses during the nerve block was evaluated using a fixed list method Qualitative Behaviour Assessment by 5 observers blinded to treatment group. They scored the treatment group as being in a more positive affective state (at ease, relaxed and confident) than the control group (nervous, tense and fearful). However, the difference between groups was only statistically significant once other significant contributing factors such as the type of block were included in the multivariable model. This highlights that many different factors are contributing to how a horse perceives veterinary Events such as nerve blocks but that classical counter conditioning can be used to help horses perceive them more positively. In summary, a horse’s perception of veterinary care will vary markedly between individual animals but also within an individual. Constant evaluation of the horse’s emotional state, based on an understanding of subtle indicators of negative affect, will allow the veterinary team to modify their approach and optimise care of the individual. In addition, it is possible to promote a positive affective state even during aversive procedures such as nerve blocks. This can be achieved in a real-life scenario using classical counter conditioning

Validación del analizador hematológico Lasercyte en burros sanos

Los burros tienen un papel central en la economía de países en desarrollado, siendo utilizados como medio de transporte, animales de carga y en labores agrarias. Más recientemente, esta especie también está siendo utilizada como mascota, así como en actividades de asinoterapia o para producción de leche. Debido a la escasa investigación en medicina asnal, muchos clínicos extrapolan rangos de referencia y datos publicados para caballos en burros. Este acercamiento se ha demostrado marcadamente erróneo, pudiendo conllevar graves errores diagnósticos, tratamientos inadecuados o innecesarios, así como aumentando un gasto injustificado para el dueño. El análisis hematológico es una de las bases del examen clínico inicial en cualquier paciente veterinario, siendo esencial establecer rangos de referencia especie-específicos y adaptados a la edad, sexo, raza, localización geográfica y equipamiento utilizado para dicha evaluación hematológica. Los analizadores automáticos hematológicos con tecnología láser son uno de los últimos avances en medicina veterinaria, permitiendo una mayor rapidez y fiabilidad en los parámetros obtenidos que los equipos basados en impedancia o las técnicas de estudio hematológico manual. Los objetivos de esta tesis doctoral fueron estudiar los resultados hematológicos en una amplia población de burros de Andalucía, determinar el nivel de acuerdo entre técnicas para esta especie y validar los resultados obtenidos con el equipo LaserCyte (analizador de tipo láser) en burros. Para desarrollar este trabajo, se obtuvo sangre de una población de 84 burros residentes en varias explotaciones de Andalucía y la misma se estudió mediante técnicas manuales, con un analizador automático de impedancia (Sysmex F820) y con el analizador LaserCyte. También se evaluó el nivel de precisión, exactitud, el error total presente en los resultados, la linealidad y otros parámetros de validación y estandarización para sangre de burro en el equipo LaserCyte. De igual forma, se comparó el grado de acuerdo entre técnicas para determinar si los resultados pudieran ser extrapolables entre sí. Por último, y ante la sospecha de marcadas diferencias en el tamaño leucocitario entre asnos y caballos que pudiesen provocar errores en el algoritmo de clasificación leucocitario de LaserCyte, se realizó un estudio morfométrico del tamaño de las distintas subpoblaciones leucocitarias en asnos. Los resultados obtenidos estuvieron dentro de los rangos hematológicos previamente reportados para esta especie, independientemente de la técnica utilizada. Se observaron diferencias significativas a nivel hematológico entre burros atendiendo al sexo (mayor número de eritrocitos circulantes pero menor hemoglobina y volumen corpuscular medio en machos respecto a hembras) y la edad (tendencia a menores contajes eritrocitarios y leucocitarios, hematocrito y hemoglobina en animales de edad avanzada respecto a jóvenes), las cuales coincidieron con las previamente observadas en esta especie por varios autores. El nivel de acuerdo entre técnicas hematológicas fue, en general, pobre, apareciendo las mayores divergencias en los contajes diferenciales leucocitarios. LaserCyte mostró un rendimiento adecuado para la mayoría de parámetros, a excepción de dichos contajes diferenciales, lo cual muy probablemente estaría relacionado con la ausencia de un algoritmo especie-específico de clasificación leucocitaria en este analizador. Esta teoría también estaría respaldada por los resultados del estudio morfométrico, que mostraron marcadas diferencias en la distribución de los tamaños leucocitarios entre asnos y caballos, en especial respecto al solapamiento entre poblaciones.Donkeys have traditionally played a central role in developing countries, being used as draft animals. In developed countries, donkeys are an emerging pet animal and new uses, such as onotherapy or milk production, are rapidly growing. Since research in donkeys is scarce, many clinicians extrapolate data and reference ranges from horses. This approach could cause misdiagnoses, unnecessary or inadequate treatments and over cost to the owner. Hematology is a basic tool in veterinary internal medicine. Species-specific intervals are crucial for a proper hematological analysis. Furthermore, differences have been reported in several parameters attending to gender, age, breed and even geographical location. Laser-based hematology analyzers are in general more accurate and costeffective compared to manual techniques and impedance analyzers. This work aims to evaluate hematological data from a donkey population from Andalusia, determine the agreement between techniques in this species and validate the LaserCyte analyzer in donkeys. Samples were obtained from 84 donkeys from several farms in Andalusia and agreement between the results from LaserCyte, Sysmex F820 and manual methods was obtained. Accuracy, precision, total error, linearity and other validation indexes for LaserCyte were calculated in those samples. A morphometric study on donkeys’ leukocytes size was carried out in order to determine any misclassification of white blood cells using equine software. Our results were within reported intervals for this species according to previous studies. Significant differences were observed attending to gender (males showing higher red blood cell counts and lower hemoglobin concentration and mean corpuscular volume) and age (lower blood and white cell counts, hematocrit and hemoglobin concentration were observed in older donkeys compared to younger ones). Those differences were in accordance to previous reports in donkeys by other authors. Agreement between techniques was poor, mostly due to incongruities in the differential leukocyte count. LaserCyte showed a reasonable performance in donkey blood, except for those differentials. This finding could be explained by the use of an equine software and misclassifications of leukocytes due to differences in size or granularity between cells in these species. Indeed, donkeys’ leukocytes size showed marked overlapping and several differences with horses in the morphometric study