Professor Chris Proudman

Gut bacterial communities have a profound influence on the health of humans and animals. Early-life gut microbial community structure influences the development of immunological competence and susceptibility to disease. For the Thoroughbred racehorse, the significance of early-life microbial colonisation events on subsequent health and athletic performance is unknown. Here we present data from a three-year cohort study of horses bred for racing designed to explore interactions between early-life gut bacterial community structure, health events in later life and athletic performance on the racetrack. Our data show that gut bacterial community structure in the first months of life predicts the risk of specific diseases and athletic performance up to three years old. Foals with lower faecal bacterial diversity at one month old had a significantly increased risk of respiratory disease in later life which was also associated with higher relative abundance of faecal Pseudomonadaceae. Surprisingly, athletic performance up to three years old, measured by three different metrics, was positively associated with higher faecal bacterial diversity at one month old and with the relative abundance of specific bacterial families. We also present data on the impact of antibiotic exposure of foals during the first month of life. This resulted in significantly lower faecal bacterial diversity at 28 days old, a significantly increased risk of respiratory disease in later life and a significant reduction in average prize money earnings, a proxy for athletic performance. Our study reveals associations between early-life bacterial community profiles and health events in later life and it provides evidence of the detrimental impact of antimicrobial treatment in the first month of life on health and performance outcomes in later life. For the first time, this study demonstrates a relationship between early-life gut bacterial communities and subsequent athletic performance that has implications for athletes of all species including humans.

The records of 200 colic episodes, collected prospectively, over a two year period, from first opinion cases, were analysed and compared with a control sample selected at random from the same population. Analysis by colic type revealed 72% spasmodic/undiagnosed; 7% surgical; 5.5% flatulent; 5% pelvic flexure impactions; 9.5% other implications and 1% colitis. A possible predisposing cause was identified in 43% of the spasmodic/undiagnosed cases. The total incidence of colic in each age group showed no statistically significant differences from the control population. Stallions were significantly under represented in the colic population. When compared with the control sample the number of spasmodic/undiagnosed colic cases in the 5-10-year-old group and the number of surgical colics in the over 15-year-old group were significantly greater than expected. No statistically significant correlation was found when the seasonal incidence of colic was compared with monthly temperature, change in monthly temperature, monthly rainfall, and rainfall weighted for temperature.

Several imaging techniques were used to diagnose hypoplasia of the soft palate in a horse. The absence of the caudal soft palate, hypertrophied lymphoid tissue and the formation of a pseudouvula were observed endoscopically. Plain and contrast radiography were used to demonstrate a soft palate remnant and to identify structures rostral to the epiglottis. Retrograde endoscopy of the pharynx via a tracheotomy incision is described.

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A centrifugation/flotation technique for the identification of equine tapeworm eggs is described. It was validated by using faeces samples from 80 horses of known tapeworm status, and had a sensitivity of 61 per cent and a specificity of 98 per cent. The exclusion of false negative results in animals with less than 20 tapeworms increased the sensitivity to 92 per cent. No significant correlation was found between the number of eggs observed and the number of tapeworms present in the horses.

INTESTINAL parasites are an important cause of morbidity and mortality in horses of all types. The prevalence of many intestinal helminths is high and, within any population of horses, certain individuals can harbour large numbers of parasites, which may lead to disease. There is widespread misunderstanding and ignorance about parasite control strategies among the horse-owning public, with the consequence that many horses are unnecessarily exposed to pathogenic burdens of intestinal parasites. It is the duty of all veterinary surgeons engaged in equine preventive medicine to be able to give sound advice to horse owners on the best anthelmintic strategy for their particular situation. This article reviews the problems of the control of intestinal parasites in horses, discusses some of the pharmaceutical and managemental control measures that can be used, and offers advice on putting this information into practice.

Without an experimental model of equine grass sickness (EGS), a randomised controlled field trial (RCT) represents the only method of evaluating the efficacy of Clostridium botulinum type C vaccination in preventing naturally occurring disease. Clinical trial feasibility is an important aspect of preliminary work undertaken prior to initiating RCTs, estimating parameters that are important for study design. This cross-sectional study aimed to assess the feasibility of conducting a nationwide RCT of a candidate vaccine for EGS based on responses from a sample of British equine veterinary practices (n = 119/284). Seventy-three percent of practices had attended >= 1 EGS case within the preceding 2 years (median four cases), and 51.3% regularly attended recurrently affected premises. Veterinary surgeons had greater confidence diagnosing acute/subacute EGS based solely on history and clinical signs compared to chronic EGS. Ninety-one percent of respondents (n = 103/113) considered the proposed RCT to be important/very important to equine veterinary research. Ninety-one percent of respondents (n = 102/112) indicated preparedness to assist in owner recruitment and 92.9% (n = 104/112) indicated willingness to participate in a RCT. The most frequent reasons for practices declining to participate were low incidence of EGS (n = 4), did not believe clients would wish to participate (n = 3) and amount of paperwork/data collection involved (n = 2). There was considerable support amongst participating veterinary practices for a RCT evaluating the efficacy of Clostridium botulinum vaccination for the prevention of EGS in Britain. Substantial proportions of participating practices would be prepared to participate in the RCT and regularly attended EGS-affected premises that would meet trial inclusion criteria. (C) 2016 Elsevier Ltd. All rights reserved.

Faeces samples from 116 horses with colic and 115 non-colic cases were assayed by a centrifugation/floatation method for the presence of tapeworm eggs. Analysis of these data failed to demonstrate an association between tapeworm infection and colic of all types. The data were further analysed according to anatomical site of the lesion causing colic. The risk of ileocaecal colic was increased in the presence of tapeworms. Age, breed and sex were evaluated as possible confounding factors but had no association with the incidence of colic. This study suggests an association between tapeworms and colic of ileocaecal origin.

Whole worm extract (WWE) and excretory/secretory (E/S) antigens of Anoplocephala perfoliata were characterised by SDS-PAGE and their use in the serodiagnosis of equine cestodosis was evaluated. An enzyme-linked immunosorbent assay (ELISA) was used to compare WWE and E/S antigen as the capture layer in an antibody capture ELISA. E/S antigen gave the best differentiation between sera from tapeworm-positive and tapeworm-negative horses. The E/S-ELISA was optimised and validated against sera from horses of known tapeworm status. This assay gave a diagnostic sensitivity of 68% (n = 38) and a specificity in helminth-naive horses of 95% (n = 20). Western blot analysis of tapeworm-positive and tapeworm-negative sera revealed the immunoreactive proteins of E/S antigen to be those of 12 and 13 kDa.

There is increasing interest in the application of serological methods to macro-parasite infections to indicate infection intensity, which in turn is related to pathogenicity. Colic is the single most important cause of mortality in horses and there is evidence that a proportion of colic cases are associated with infection with the intestinal cestode Anoplocephala perfoliata. In order to develop better tools to investigate this association, the correlation between antigen-specific equine IgG and IgG(T) and infection intensity of A. perfoliata was investigated. Affinity purification of a 12/13 kDa protein doublet from crude excretory/secretory (E/S) products, and its use in enzyme linked immunosorbent assays (ELISA) is described. Its use in the immunodiagnosis of equine cestodosis and the correlation of anti-12/13 kDa IgG and IgG(T) with parasite burden is investigated using sera from 94 horses of known tapeworm infection intensity. The anti-12/13 kDa IgG and IgG(T) ELISAs gave correlation coefficients with infection intensity of 0.56 and 0.63 respectively. Linear regression analysis also indicated that anti-12/13 kDa IgG(T) was the best predictor of infection intensity. The decay of anti-12/13 kDa IgG(T) in houses following the elimination of A. perfoliata is demonstrated for four horses. Specificity of the anti-12/13 kDa IgG(T) ELISA is investigated with sera from 33 A. perfoliata negative horses with other helminth infections. Immunoblotting studies demonstrate no cross-reactivity between A. perfoliata 12/13 kDa antigen and the protein antigens of other helminths. It is concluded that assay of anti-12/13 kDa IgG(T) provides a useful tool for the assessment of A perfoliata infection intensity for clinical diagnosis and for epidemiological studies.

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The equine intestinal cestode Anoplocephala perfoliata has been the subject of recent epidemiological and immunological studies because of its suspected association with intestinal disease in the horse. We have previously shown that the IgG(T) subtype antibody response to the 12/13 kDa component of the parasite excretory/secretory (E/S) antigen is positively correlated with parasite intensity. In this study, we utilize that correlation to examine the changes in natural infection intensity with age. Infection intensity based on IgG(T) responses showed a triphasic age-dependency pattern with peak mean worm burden in the 6 months–2 years age group, falling to a lower plateau level from 3 to 15 years, and rising again in older age groups. Anti-E/S total IgG was found to have a convex age-dependency curve, with maximal response in the 6 months–2 years old age group. IgG(a) showed a triphasic response similar to the age-intensity profile of IgG(T); IgG(c) showed steadily increasing levels of antibody with age. The IgG(b) age-dependency profile was intermediate between IgG(a) and IgG(c). Age-specific correlation coefficients between anti-12/13 kDa IgG(T) (as a measure of infection intensity) and IgG(a) and IgG(b) revealed statistically significant values for many age groups. The relative importance of exposure to infection and the development of acquired immunity as determinants of the observed age-intensity pattern is considered.

One hundred and-two horses requiring to be euthanased for a variety of reasons were killed by the intravenous injection of a mixture of quinalbarbitone sodium (400 mg/ml) and cinchocaine hydrochloride (25 mg/ml). The dose rates used were 1 ml/10, 15, 20 and 30 kg bodyweight, and the time of injection was varied between 5 and 25 seconds. The average time to collapse from the start of the injection was 34 seconds and the average time to clinical death was 230 seconds. Slow injection (particularly of the low dose rates) and premedication with detomidine resulted in a longer time to collapse (median 46 seconds). Premedication with xylazine and low dose rates of the mixture resulted in an unacceptable degree of muscular activity and agonal gasping and death was delayed. Premedication with romifidine and butorphanol resulted in an apparent (but insignificant) reduction in the time to collapse and death but was also accompanied by significant agonal gasping. Without premedication quinalbarbitone and cinchocaine resulted in a smooth and quiet collapse with the cessation of cardiac and respiratory functions within three minutes in all cases, but the palpebral reflex of the horses was prolonged significantly beyond the time when all other reflex activity was lost. Occasional gasping and muscular tremors, particularly of the upper forelimb, occurred particularly when lower dose rates and either very slow or very fast rates of injection were used. One horse which was premedicated with xylazine and received a very low dose at a slow rate showed unacceptably violent muscular activity. At no other time was the procedure regarded as violent or unacceptable.

REASONS FOR PERFORMING STUDY: The frequency of recurrent colic in the UK equine general practice population is previously unreported. Elucidating risk factors for recurrent colic could provide a basis for clinical decision making and interventions. OBJECTIVES: To determine the incidence rate of and risk factors for recurrent colic. HYPOTHESES: Horse management, prophylactic health care and innate behaviour contribute to the risk of recurrent colic. METHODS: A cohort of 127 horses was enrolled at the point of a veterinary-diagnosed medical colic episode. Participating owners completed a baseline and 3 follow-up telephone questionnaires over one year. Clinical details of each colic episode were collected with data on management, behaviour and preventive healthcare. Incidence was calculated using time at risk data; non-time varying covariates were assessed for association with recurrent colic using multivariable logistic regression. RESULTS: The recurrence rate was 50 colic events/100 horse years at risk (HYAR). Including only veterinary attended recurrent colic episodes the incidence was 35 colic events per 100 HYAR. A multivariable logistic regression model was built to explore non-time varying risk factors for recurrence collected from baseline data. The model showed that horses that have a known dental problem (OR 5.5, 95% CI 1.3, 23.1) or crib-bite/windsuck (OR 12.1, 95% CI 1.4, 108.1) were at increased risk of recurrence during the year following a colic event. CONCLUSIONS AND POTENTIAL RELEVANCE: The incidence of recurrence in horses following a medical episode of colic is high in this population and represents a welfare concern. The incidence rate can be used to compare intervention efficacy in similar populations. Identified risk factors could provide the basis for management interventions or highlight at risk individuals.

As in other competitive sports, the famous Grand National steeplechase, which is held at Aintree in the United Kingdom and is watched by 600 million people worldwide, sometimes results in injury. By analysing data from the past 15 Grand National races (consisting of 560 starts by horses), we are able to identify several factors that are significantly associated with failure to complete the race: no previous experience of the course and its unique obstacles, unfavourable ground conditions (too soft or too hard), a large number of runners, and the length of the odds ('starting price'). We also find that there is an increased risk of falling at the first fence and at the jump known as Becher's Brook, which has a ditch on the landing side. Our findings indicate ways in which the Grand National could be made safer for horses and illustrate how epidemiological analysis might contribute to preventing injury in competitive sport.

The pattern of long-term survival and specific factors associated with long-term survival have not previously been evaluated in horses with a strangulating large colon volvulus (LCV).

The association between the equine intestinal tapeworm Anoplocephala perfoliata and specific types of intestinal disease was investigated by matched case-control study using coprological and serological diagnosis. We have previously shown that the host IgG(T) response to 12/13 kDa antigens of A. perfoliata correlates well with infection intensity, therefore this antibody response was used to investigate the risk of colic at different levels of parasite infection intensity. One hundred and three spasmodic colic cases with an equal number of controls matched for age, breed and gender, and 20 ileal impaction cases each with 2 similarly matched controls were obtained. Cases of spasmodic colic were much more likely (odds ratio = 8.0) to be associated with A. perfoliata infection detected coprologically than controls. Serological diagnosis revealed an increasing risk of spasmodic colic with increasing infection intensity. Calculation of an aetiological fraction suggests that 22% of spasmodic colic cases in this study were tapeworm associated. No significant association was found between colic and strongyle egg count. Conditional logistic regression analysis demonstrated that the relationship between colic and A. perfoliata infection intensity was not confounded by strongyle egg count and there was a linear relationship between infection intensity and the log-odds of spasmodic colic. For cases of ileal impaction, a strong association was found between colic and A. perfoliata as diagnosed by coprological means (odds ratio of 34.0). Serological diagnosis also revealed a strong association that increased with higher levels of infection intensity (odds ratio = 26.0). The aetiological fraction for the ileal impaction data suggests that 81% of the ileal impaction cases in this study were tapeworm associated. This study concludes that A. perfoliata is a significant risk factor for spasmodic colic and ileal impaction colic in the horse; and that the risk of spasmodic colic increases with infection intensity.

REASONS FOR PERFORMING STUDY: Idiopathic focal eosinophilic enteritis (IFEE) and diffuse eosinophilic enteritis (DEE) are primary eosinophilic intestinal conditions without a known cause that are associated with an increasing number of surgical colic cases. Histology may be helpful in defining disease aetiology and pathogenesis. OBJECTIVES: To characterise further the inflammatory infiltrate in equine IFEE and to compare the condition with DEE. METHODS: Twenty-three IFEE cases and 5 DEE cases were examined by light microscopy including immunohistology to identify infiltrating leucocytes. Inflammatory infiltrates in mucosa and submucosa were characterised in IFEE lesions (Group 1), the intestine distant from the lesions in IFEE (Group 2) and DEE (Group 3). RESULTS AND CONCLUSIONS: IFEE lesions represented an accumulation of leucocytes in submucosa and muscularis, with dominance of eosinophils and macrophages and smaller numbers of lymphocytes, plasma cells and neutrophils. T cells represented the dominant lymphocytes. The mucosa overlying the lesion and both mucosa and submucosa in IFEE nonlesion sites and in DEE exhibited a similar composition, with different prevalence of various cell types. Macrophages were significantly more prevalent in the mucosal and submucosal infiltrates in IFEE nonlesion sites than in DEE, and lymphocytes significantly more prevalent in the mucosa in DEE than in IFEE nonlesion sites. The findings confirm IFEE as a primary eosinophilic intestinal disorder and indicate that IFEE represents a focally exacerbated inflammatory reaction in horses with DEE, possibly due to functional changes in the macrophage and T cell components, with subsequent excessive recruitment of both eosinophils and macrophages.

REASONS FOR PERFORMING STUDY: Horses requiring different methods of intestinal anastomosis during equine colic surgery may have differences in mortality and morbidity. HYPOTHESES: Horses requiring side-to-side jejunocaecal anastomosis have a higher mortality and morbidity rate than those requiring end-to-end jejunojejunal anastomosis. Morbidity and mortality of handsewn vs. stapled side-to-side jejunocaecal anastomoses are not significantly different. METHODS: A prospective, nonrandomised, observational study was conducted. Clinical and surgical details were recorded during hospitalisation and survival data acquired by periodic telephone questionnaire. Differences in mortality and morbidity rate were evaluated by survival analysis. RESULTS: A total of 184 horses underwent end-to-end jejunojejunal anastomosis and 178 underwent side-to-side jejunocaecal anastomosis. Horses with a jejunocaecal anastomosis had a significantly higher mortality rate. The incidence of post operative colic in horses requiring side-to-side jejunocaecal anastomoses was greater than those requiring end-to-end jejunojejunal anastomoses. Within the group undergoing side-to-side jejunocaecal anastomosis there was no evidence of differential survival between horses with handsewn vs. stapled anastomoses. CONCLUSIONS: Mortality rate is higher in horses that have required side-to-side jejunocaecal anastomosis than in those that needed end-to-end jejunojejunal anastomosis; and post operative colic is more common after side-to-side jejunocaecal anastomosis. No difference in mortality was found between horses with handsewn and stapled side-to-side jejunocaecal anastomoses. POTENTIAL RELEVANCE: Surgeons should be aware of the increased mortality and morbidity in horses requiring side-to-side jejunocaecal anastomosis. Our finding of no difference in mortality between handsewn and stapled side-to-side jejunocaecal anastomoses justifies surgeons exercising personal preference in their selection of anastomosis method.

REASONS FOR PERFORMING STUDY: The reasons why equine grass sickness (EGS) recurs on premises are unknown and, consequently, practical methods for reducing the risk of recurrence are not available. OBJECTIVES: To identify risk factors associated with recurrence of EGS on premises and to gain possible insights into the pathogenesis of the disease. METHODS: Data on disease history and risk factors were collected by postal questionnaire from premises with EGS cases between 1st January 1997 and 31st December 2001. Data on variation in rates of recurrence of EGS for different risk factors were analysed using Poisson regression analysis. RESULTS: Of 509 premises contacted, 305 (60%) returned useable questionnaires and 100 of these (33%) were classified as 'recurrent' premises. An overall median incidence rate for EGS of 2.1 EGS incidents/100 horses/premises/year was recorded. There was an increased rate of recurrence with higher numbers of horses, presence of younger animals, stud farms and livery/riding establishments, loam and sand soils, rearing of domestic birds and mechanical droppings removal. The rate of recurrence decreased with chalk soil, cograzing ruminants, grass cutting on pastures and removal of droppings by hand. Several statistically significant interactions were identified. CONCLUSIONS: Many of the findings are consistent with the theory that EGS is a toxico-infectious form of botulism. Several of the significant factors identified may directly or indirectly relate to soil disturbance and consequent soil contamination of grass, thereby increasing the rate of exposure of grazing horses to Clostridium botulinum, which resides in soil. POTENTIAL RELEVANCE: Identification of potentially modifiable risk factors may, ideally following validation in appropriately designed, controlled and randomised intervention studies, lead to practical measures to reduce the incidence of EGS on previously affected premises.

Cyathostomins are the most common parasitic nematodes of horses. Larval stages, which inhabit the intestinal wall, are particularly pathogenic and can cause severe colitis and colic. Despite their clinical importance, diagnostic techniques for the prepatent stages do not exist. A method that could estimate mucosal infection intensity would have a major impact on the control and diagnosis of cyathostominosis. Here, serum IgG(T) responses to two larval antigen complexes of 25 and 20 kDa were quantified in horses with experimental infections, natural infections and in horses that presented with clinical larval cyathostominosis. In experimentally-infected animals, anti-25 kDa complex IgG(T) levels correlated positively with field exposure and with early third stage larval (r(s)=0.74, P=0.015) and total mucosal parasite (r(s)=0.78, P=0.010) burdens. In naturally exposed horses whose parasite burdens were quantified upon post-mortem examination, antigen-specific IgG(T) responses were significantly higher in infected than in uninfected horses (P=0.0001 and 0.002, for anti-25 and anti-20 kDa responses, respectively). In these animals, anti-25 kDa IgG(T) levels correlated positively with mucosal and lumenal burdens (P

Equine grass sickness (EGS) is recognized as a debilitating and predominantly fatal neurodegenerative disease affecting grazing equids. The gastrointestinal tract is the most severely affected body system, resulting in the main clinical signs of colic (acute grass sickness), weight loss, or dysphagia (chronic grass sickness). EGS predominantly occurs within Great Britain, although it is also recognized in regions of mainland Europe, and mainly affects young horses with access to pasture in the springtime. There is strong evidence of an association between EGS and the type C toxins produced by the bacterium Clostridium botulinum. This article covers the clinical aspects, epidemiology, and global distribution of EGS, along with comparisons with botulism and developments in disease prevention.

The objective of this study was to identify risk factors, during racing, associated with imminent fatal distal limb fracture in Thoroughbreds. One hundred and nine cases of fatal distal limb fracture were identified from all 59 UK racecourses over a 2-year period (February 1999-January 2001). Three uninjured control horses were randomly selected from the same race as the case horse. Videos of races in which fractures occurred were viewed using a defined protocol. Fractures in flat races occurred at any time during the race, whereas 74% (45/61) of cases in national hunt type races occurred in the second half of races. More than 75% (79/103) of cases were spontaneous, i.e. there was no obvious external influence such as a fall at a fence or collision with another horse. Sixty-six percent (44/67) of horses, sustaining a forelimb fracture, fractured the forelimb they were using as lead leg at the time of fracture. When case and control horses were compared, horses that were: (a) making good progress through the race, (b) reluctant to start and (c) received encouragement in the final 10s before the time of fracture, were more likely to sustain a fracture.

Colic remains a significant problem in the horse in terms of welfare and economics; in some equine populations it is the single most common cause of death. Many causes of colic are cited in the equestrian and veterinary literature but little scientific evidence exists to substantiate these theories. Recent epidemiological investigations have confirmed that colic is complex and multi-factorial in nature. Studies have identified a number of factors that are associated with increased risk of colic including parasite burden, certain feed types, recent change in feeding practices, stabling, lack of access to pasture and water, increasing exercise and transport. These findings are reviewed together with examples of management practices that may be altered to reduce the incidence of specific types of colic. This is an opinionated, not a systematic, review focusing on those areas that are considered most relevant to the practitioner.

REASONS FOR PERFORMING STUDY: Equine grass sickness is a high mortality disease which, despite many years of investigation, is of unknown aetiology. Recent findings indicating that the disease is associated with Clostridium botulinum require support from an epidemiological study that recognises and controls for potential confounders, e.g. age, time of year and premises. HYPOTHESIS: EGS is associated with low antibody levels to C. botulinum antigens. METHODS: A matched case-control study was conducted. Data were collected from 66 histologically confirmed cases of EGS and 132 premises-matched control horses. The probability of EGS in horses was modelled using conditional logistic regression. RESULTS: EGS was significantly associated (age-adjusted P < 0.005) with low antibody levels to each of 3 clostridial antigens; C. botulinum type C and C. novyi type A surface antigens and a C. botulinum type C toxin complex toxoid. These serological risk factors for EGS remained highly significant when entered into multivariable models. This study also identified new horse-level risk factors for EGS; feeding hay or haylage was associated with a decreased risk of disease, change of feed type or quantity during the 14 days prior to disease was associated with increased risk, and the use of an ivermectin anthelmintic at both the ultimate and penultimate treatments was also associated with a significantly increased risk of EGS. CONCLUSIONS: This study provides strong support for the role of C. botulinum in the aetiology of EGS and identifies managemental risk factors for the disease. POTENTIAL RELEVANCE: Increasing anticlostridial antibody levels by vaccination and appropriate managemental interventions may decrease the risk of EGS occurring.

Intestinal parasites are an important cause of morbidity and mortality in horses of all types. The prevalence of many intestinal helminths is high and, within any population of horses, certain individuals can harbour large numbers of parasites, which may lead to disease. There is widespread misunderstanding and ignorance about parasite control strategies among the horse-owning public, with the consequence that many horses are unnecessarily exposed to pathogenic burdens of intestinal parasites. It is the duty of all veterinary surgeons engaged in equine preventive medicine to be able to give sound advice to horse owners on the best anthelmintic strategy for their particular situation. This article reviews the problems of the control of intestinal parasites in horses, discusses some of the pharmaceutical and managemental control measures that can be used, and offers advice on putting this information into practice.

REASONS FOR PERFORMING STUDY: Epiploic foramen entrapment (EFE) is a common cause of small intestinal strangulation in the horse and its epidemiology requires further investigation. OBJECTIVES: To identify horse- and management-level risk factors for EFE and to explore reasons for the apparent seasonality of this condition. HYPOTHESIS: Horses exhibiting certain behaviours and those exposed to particular management practices that vary seasonally are at increased risk of EFE. METHODS: A prospective unmatched, multicentre case-control study was conducted over 24 months in the UK. Data on 77 cases and 216 control horses were obtained from 9 collaborating clinics and logistic regression was used to identify associations between horse and management variables and the likelihood of EFE. RESULTS: In a final multivariable model crib-biting/windsucking behaviour was associated with the largest increase in likelihood of EFE. A history of colic in the previous 12 months, increased stabling in the previous 28 days and height of the horse also increased the likelihood of EFE. Horses with access to a mineral/salt lick, those easily frightened and horses not fed at the same time as others were at reduced risk of EFE. CONCLUSIONS: Horses exhibiting certain behaviours, those with a previous history of colic and horses of greater height appear to be at inherently greater risk of EFE. The increase in likelihood of EFE with increased duration of stabling may explain the apparent seasonality of this condition.

In this paper, the management and health problems of donkeys on loan to independent carers from The Donkey Sanctuary are characterised, and the demographics of the UK's donkey population are described using data from a variety of sources. All carers that fostered a donkey from The Donkey Sanctuary between September 2004 and August 2005 (1432 donkeys) were surveyed using a postal questionnaire requesting information about the donkey, its premises, daily care, health and preventive medicine. The response rate was 77.8 percent. The mean (sd) age of the donkeys was 20.8 (7.4) years. The majority (92 percent) of the donkeys were kept as pets. Approximately one-third (33.6 percent) of the donkeys were overweight. The most common medical problems were hoof, dermal and oral problems. A total of 86 percent of the donkeys had a dental examination at least every 12 months, and at least 45 percent had at least one dental problem.

Reasons for performing study: Recurrent colic occurs frequently in the general horse population but little evidence exists about what factors place horses at greater risk of recurrent colic. Objectives: To quantify time-varying and non time-varying risk factors for recurrent colic among horses attended by first-opinion veterinary surgeons in northwest England. Study design: Nested case-control study. Methods: A nested case-control study was conducted on data from a prospective longitudinal cohort study of 127 horses recruited subsequent to an episode of medical colic. Data were collected on management and recurrent colic episodes at 4-monthly intervals by telephone questionnaires. All recurrent colic episodes were selected as cases; controls were unmatched and randomly selected from all horse-time at risk. Data relating to the 30 days prior to the date of colic or control selection were used to determine exposure status. Multivariable logistic regression analyses were used to determine risk factors for recurrent colic. Results: In total, 59 cases and 177 controls were included. The final model showed that horses that displayed crib-biting/windsucking (odds ratio [OR] 10.1, 95% confidence interval [CI] 2.5-41.0) or weaving behaviour (OR 3.9, 95% CI 1.5-10.1) had an increased risk of recurrence of colic. Increasing time at pasture reduced the risk of recurrence (OR 0.99, 95% CI 0.99-1.0). A significant interaction was found suggesting that the risk associated with crib-biting/windsucking may be modified by feeding fruit/vegetables; however, further research is required before recommendations for feeding practices can be made. Conclusions: This study suggests that sufficient access to pasture may be an important means of reducing recurrent colic risk. The behavioural risk factors highlight individuals who may be at increased risk of recurrent colic and whose colic prevention strategies should be carefully managed.

Gut microbial communities are increasingly being linked to diseases in animals and humans. Obesity and its associated diseases are a concern for horse owners and veterinarians, and there is a growing interest in the link between diet, the intestinal microbiota and metabolic disease. Assess the influence of long-term hay or haylage feeding on the microbiota and metabolomes of 20 Welsh mountain ponies. Longitudinal study. Urine, faeces and blood were collected from 20 ponies on a monthly basis over a 13-month period. Urine and faeces were analysed using proton magnetic resonance ( H NMR) spectroscopy and faecal bacterial DNA underwent 16S rRNA gene sequencing. Faecal bacterial community profiles were observed to be different for the two groups, with discriminant analysis identifying 102 bacterial groups (or operational taxonomic units, OTUs) that differed in relative abundance in accordance with forage type. Urinary metabolic profiles of the hay and haylage fed ponies were significantly different during 12 of the 13 months of the study. Notably, the urinary excretion of hippurate was greater in the hay fed ponies for the duration of the study, while ethyl-glucoside excretion was higher in the haylage fed ponies. The study was undertaken over a 13-month period and both groups of ponies had access to pasture during the summer months. The data generated from this study, suggest that the choice of forage may have implications for the intestinal microbiota and metabolism of ponies and therefore, potentially their health status. Understanding the potential implication of feeding a particular type of forage will enable horse owners to make more informed choices with regard to feed, especially if their horse or pony is prone to weight gain.

The risk of fatal distal limb fractures in thoroughbreds racing in the UK was calculated and shown to vary considerably between the different types of race. Flat turf racing was associated with the lowest risk (0.4 per 1000 starts) and national hunt flat racing was associated with the highest risk (2.2 per 1000 starts). The types of fracture were classified by detailed radiographic and postmortem examinations of all the cases recorded over two years, and the distribution of the different types of fracture in the five main types of racing was examined. Overall, lateral condylar fractures of the third metacarpus were the most common, and they were also the most common in national hunt-type races (hurdle, steeplechase and national hunt flat races). In all-weather flat racing biaxial proximal sesamoid fractures were most common, and in turf flat racing fractures of the first phalanx were most common. The risk of fractures of more than one bone was greater in national hunt-type races.

Idiopathic focal eosinophilic enteritis (IFEE) is a rare disease in the horse and few cases have been reported in the literature. The objective of this paper was to describe the clinical, surgical, histological features and post-operative progress of 12 horses with IFEE, a disease that had not been identified in the authors' hospital population prior to 2000. Diagnosis of IFEE was made at laparotomy and confirmed by histological examination of resected abnormal small intestine. In all 12 horses colic was associated with jejunal obstruction involving visibly striking and palpably thickened serosal plaques or circumferential constrictions. Surgical resection of affected intestine was performed in 10 horses, of which seven completely recovered. In one horse, on which surgical resection was not performed, ingesta re-obstructed post-operatively at one of the lesions. Histological examination of resected intestines revealed, in all cases, severe transmural enteritis in which eosinophilic leucocytes were the predominant inflammatory cells. No helminths were identified and the cause of this lesion is not known. IFEE is an uncommon but significant cause of small intestinal obstruction for which surgical resection can be curative. The gross lesions may be under-recognised and histology is essential for diagnosis.

REASONS FOR PERFORMING STUDY: Epiploic foramen entrapment (EFE) has been associated with reduced post operative survival compared to other types of colic but specific factors associated with reduced long-term survival of these cases have not been evaluated in a large number of horses using survival analysis. OBJECTIVE: To describe post operative survival of EFE cases and to identify factors associated with long-term survival. METHODS: A prospective, multicentre, international study was conducted using clinical data and long-term follow-up information for 126 horses diagnosed with EFE during exploratory laparotomy at 15 clinics in the UK, Ireland and USA. Descriptive data were generated and survival analysis performed to identify factors associated with reduced post operative survival. RESULTS: For the EFE cohort that recovered following anaesthesia, survival to hospital discharge was 78.5%. Survival to 1 and 2 years post operatively was 50.6 and 34.3%, respectively. The median survival time of EFE cases undergoing surgery was 397 days. Increased packed cell volume (PCV) and increased length of small intestine (SI) resected were significantly associated with increased likelihood of mortality when multivariable analysis of pre- and intraoperative variables were analysed. When all pre-, intra- and post operative variables were analysed separately, only horses that developed post operative ileus (POI) were shown to be at increased likelihood of mortality. CONCLUSIONS: Increased PCV, increased length of SI resected and POI are all associated with increased likelihood of mortality of EFE cases. This emphasises the importance of early diagnosis and treatment and the need for improved strategies in the management of POI in order to reduce post operative mortality in these cases. POTENTIAL RELEVANCE: The present study provides evidence-based information to clinicians and owners of horses undergoing surgery for EFE about long-term survival. These results are applicable to university and large private clinics over a wide geographical area.

A 6-year-old mare was presented for acute abdominal pain unresponsive to analgesics. Exploratory laparotomy revealed entrapment of the small colon through a 12 cm rent in the mesocolon. The incarcerated small colon was manually reduced and the rent in the mesocolon was sutured closed. The mare made excellent postoperative recovery and was discharged from the hospital 4 days later. The cause of the rent, which was chronic in appearance, is unknown.

Intestinal motility disorders are an important problem in horses and donkeys and this study was carried out in order to evaluate the enteric neurons in animals with and without intestinal disease. Surplus intestinal tissue samples were collected from 28 horses undergoing exploratory laparotomy for colic. In addition, surplus intestinal samples from 17 control horses were collected immediately following humane destruction for clinical conditions not relating to the intestinal tract. Similar samples were also collected during routine post-mortem examinations from 12 aged donkeys; six animals were humanely destroyed for conditions related to the intestinal tract, while the remaining six were humanely destroyed for other reasons including dental and orthopaedic diseases. Tissue samples were fixed in formalin and immunohistochemical labelling was performed targeting the enteric neurons using a polyclonal antibody specific for the neuronal marker PGP 9.5. The distribution and density of neuronal networks were assessed qualitatively and semiquantitatively. There was strong PGP 9.5 expression in both the horse and donkey samples and labelling was detected throughout the tissue sections. In both species, PGP 9.5-immunoreactive nerve fibres were detected in all layers of the intestinal tract, both in large and small intestinal samples. Networks of enteric neurons were present in the donkey with a similar distribution to that seen in the horse. There was no demonstrable difference in enteric neuronal density and distribution in the groups of animals with intestinal disease compared with those without, apart from two (out of 28) horses with intestinal disease that showed a marked reduction in PGP 9.5 immunoreactivity. Apart from these two animals, this total cohort analysis differs from some previously observed findings in horses with intestinal disease and may therefore reflect the different pathophysiological processes occurring in varying intestinal conditions resulting in colic both in the donkey and the horse. © 2013 Elsevier Ltd.

A 12/13 kDa antigen, tapeworm ELISA test, developed for use in horses, was used to detect parasite-specific serum antibody, IgG(T), in the serum of donkeys. In a pilot study the 12/13 kDa antigen was tested and proved to detect the antibody, IgG(T), in donkey sera. Blood samples from 797 donkeys, naturally exposed to cestode infection, from four geographical localities were collected and sera were prepared and analysed. There was substantial serological evidence that donkeys were potentially infected with A. perfoliata. A range of ELISA OD values were obtained from the serological assay. Over 26% and 7.5% of the donkeys were moderately and highly infected, respectively, showing at least a 34% sero-prevalence. The rest, 66.1%, were either with low infection intensity or negative for A. perfoliata infection. The risk of infections, both in sero-prevalence and intensity, as determined by ELISA optical density (OD), were highest in the highland areas of Ethiopia where pastures are low-lying and wet, and permanent pasture management is regularly practised. Sex, age and body condition of the donkeys had no significant effect either on prevalence of the infection or on the serum antibody level. These results indicate a risk of intestinal disorders, particularly, colic, associated with A. perfoliata infection in donkeys.

We present here the use of targeted, long-read sequencing of the myostatin (MSTN) gene as a model to detect potential gene editing events in Thoroughbred horses. MSTN is a negative regulator of muscle development, making the gene a prime candidate target for gene doping. By sequencing the complete gene in one PCR product, we can catalogue all mutations without the need to produce short-fragment libraries. A panel of reference material fragments with defined mutations was constructed and successfully sequenced by both Oxford Nanopore and Illumina-based methods, showing that gene doping editing events can be detected using this technology. To ascertain the normal variation within the population, we sequenced the MSTN gene in 119 UK Thoroughbred horses. Variants from the reference genome were assigned to haplotypes and eight distinct patterns, designated Hap1 (reference genome) to Hap8, were determined with haplotypes Hap2 and Hap3 (which includes the 'speed gene' variant) being far the most prevalent. Hap3 was most abundant in flat-racing horses, whereas Hap2 was most abundant in jump-racing. Within this data set, results for 105 racehorses from out-of-competition sampling were compared between matrices of extracted DNA and direct PCR of whole blood from lithium heparin gel tubes, and strong agreement was found between the two methods. The direct-blood PCR was achieved without compromising the sample prior to plasma separation for analytical chemistry, and could thus be used as part of a routine screening workflow for gene editing detection.

Colic surgery is a frequently performed operation with high postoperative mortality. This study was undertaken to identify variables associated with decreased postoperative survival. We used data from 321 horse years of postoperative survival time to model the probability of survival following recovery from colic surgery. Continuous variables were modelled using a 6 variable, penalised Cox regression model. This demonstrated approximately linear relationships between survival and the following variables: increase in packed cell volume (PCV), intestinal resection length, time to surgery (interval between onset of colic and surgery) and duration of surgery. No significant decrease in survival was demonstrated with increasing age of the patient or with heart rate. The only categorical variable to be significantly associated with decreased survival was epiploic foramen entrapment. The final, fixed effects Cox proportional hazards model of postoperative survival included the variables epiploic foramen entrapment, PCV, resection length and duration of surgery, each variable adjusted for the nonlinear relationship with time to surgery. Residual variation in postoperative survival attributable to professional personnel (referring veterinary surgeon, anaesthetist and surgeon) was explored by fitting each as a random effects term in the model. Little of the residual variation could be attributed to any category of personnel. Model diagnostics indicated little influence by individual outliers on model parameters and little evidence of subjects poorly predicted by the final model. The study highlights factors influencing the long-term survival of horses recovering from colic surgery and proposes a model that can be used to inform prognosis.

Due to the ease of collection, transport and storage, the use of dried blood spots (DBS) offers an attractive alternative matrix for detection of the abuse of gene therapy, otherwise known as gene doping. This study evaluated the recovery, extraction efficiency and resulting detection capability of DNA from DBS by evaluating different target types, DNA extraction kits, the number of punches and blood tube preservatives. The long-term storage stability of low-copy-number transgene targets in DBS was not assessed in this study but would be noteworthy to investigate further. DNA was quantified using two detection methods: qPCR and digital PCR (dPCR). Using six punches with the Qiagen Investigator kit gave the best overall DNA yield compared with other extraction methods. Including three punches, however, gave better DNA extraction efficiency. Reference material could be detected using qPCR and dPCR in DBS spiked with 5000 copies/mL of blood (approximately 15 copies per 3 mm of punch). The optimal DNA extraction protocol was used on DBS samples from a custom recombinant adeno-associated virus administration study and showed successful detection of vector targets in DBS samples. DNA extraction methods for dried blood spots (DBS) were evaluated to determine the suitability of the use of DBS as an alternative matrix for transgene detection towards gene doping screening. DNA was quantified using two detection methods: qPCR and dPCR. Using the Qiagen Investigator kit for DNA extraction with six DBS punches gave a better recovery of targets than three. However, three punches gave better extraction efficiency. DNA can be detected down to 5000 copies/mL of blood using qPCR and dPCR. image

The serological changes in two groups of horses known to be harbouring Anoplocephala perfoliata were studied; 12 were treated with 1.5 mg/kg praziquantel and 200 microg/kg ivermectin, and 14 were treated with 200 microg/kg ivermectin. Serological and faecal analyses were carried out on each animal at intervals for 758 days. The titres of antibodies specific for A perfoliata decreased from the day of treatment to day 28 in both groups, and continued to decrease in the group treated with praziquantel and ivermectin, with the first significant decrease from the other group at day 121. From day 151 to day 295 the first significant increase in antibody levels in the group treated with both drugs was observed; no A perfoliata eggs were detected in the faeces of these animals until day 295 when five of the 10 were positive.

The clinical features of 71 cases in 70 horses in which part of the small intestine became entrapped in the epiploic foramen are described. The horses' sex, age and breed, the month during which they were affected, and whether they exhibited stereotypic behaviour were compared with the same variables in 1279 horses which suffered other types of surgical colic during the same period. Thoroughbred and thoroughbred cross horses were over-represented among the 70 affected horses. There was no age or sex predilection. Fifty-five (77.5 per cent) of the cases occurred between October and March and 15 (21.1 per cent) occurred in January. The cases were significantly more likely to have a history of crib-biting/windsucking than the control group (odds ratio 7.9, 95 per cent confidence interval 4.1 to 15.3). The condition had recurred in two of the horses. Fifty-eight (81.7 per cent) recovered from surgery and 49 (69 per cent) survived until they were discharged from hospital. The median survival time of 31 of the affected horses discharged from the hospital was 700 days, whereas 417 horses with other types of surgical colic had a median survival time in excess of 1931 days.

REASONS FOR PERFORMING STUDY: Incisional drainage and suppuration occurs commonly following exploratory laparotomy; any technique with the potential to reduce the incidence of this complication warrants investigation. OBJECTIVES: To determine if abandoning the use of subcutaneous sutures in laparotomy wound closure is safe and whether it reduces the risk of suppuration. METHODS: A randomised controlled study was carried out at 2 referral hospitals in the UK, involving 309 horses undergoing exploratory laparotomy; 150 horses underwent '2-layer' closure without a subcutaneous suture, while 159 underwent conventional '3-layer' closure. Information regarding degree of oedema and gross types of discharge was recorded daily; suppuration was defined as discharge of pus. Telephone follow-up was carried out 30 days after hospital discharge to identify those complications occurring after that date and, thereafter, every 3 months. The influence of closure method on risk of wound suppuration was assessed by Chi-squared analysis and by logistic regression. Time to suppuration was modelled using a Cox proportional hazards model. RESULTS: No catastrophic failures of 2-layer closures were recorded. Prevalence of suppuration was not significantly different, being 18.7% and 23.9% for 2- and 3-layer closures, respectively (OR = 1.37, 0.79-2.37, P = 0.263). CONCLUSIONS AND POTENTIAL RELEVANCE: This study found no significant difference in prevalence or rate of wound suppuration in 2-layer closures compared to conventional 3-layer closure. Two-layer closure is recommended as a safe alternative means of achieving ventral midline abdominal closure in horses.

Between March 1, 2000 and August 31, 2001, a case-control study was conducted on 12 racecourses in England and Wales to identify and quantify the risk factors associated with horse falls in hurdle races. The cases and controls were defined so that variables relating to the horse, the jockey, the race and racecourse, and the jump could be considered. The cases were defined as a jumping effort at a hurdle flight that resulted in a fall, and the controls were defined as a successful jump over a hurdle at any of the 12 racecourses within 14 days before or after the case fall. Conditional logistic regression was used to examine the univariable and multivariable relationships between the predictor variables and the risk of falling. The risk of falling was significantly associated with the position of the jump in the race, and with the distance and speed of the race. A horse's previous racing experience and history were also significantly associated with the risk of falling and horses participating in their first hurdle race were at almost five times greater risk of falling than horses that had hurdled before.

REASONS FOR PERFORMING STUDY: Adipose tissue is an important source of inflammatory cytokines (adipokines) and adiposity has been identified as having a significant effect on human morbidity and mortality. Obesity is also an emerging welfare problem in the UK horse population, but the role that it plays in secondary diseases is unclear. OBJECTIVES: To examine the expression of inflammation-related adipokine genes in retroperitoneal adipose tissue of horses undergoing emergency abdominal surgery and to explore associations with adiposity and post operative survival. METHODS: Retroperitoneal adipose tissue samples were obtained from 76 horses undergoing emergency abdominal surgery. Real-time PCR was used to measure gene expression for leptin, adiponectin, tumour necrosis factor-alpha, macrophage chemoattractant protein-1, macrophage inhibitory factor, serum amyloid A, haptoglobin and interleukin-1. Multivariate patterns of adipokine expression were explored with principal component analysis (PCA), whilst univariable associations with post operative survival were tested in a Cox proportional hazards model. RESULTS: Leptin gene expression was higher in overweight and obese horses than in lean animals. Expression of mRNA encoding adiponectin mRNA in visceral adipose tissue was positively associated with increased post operative mortality (hazard ratio 1.31, 95% CI 1.05-1.65). However, PCA did not demonstrate multivariable patterns of adipokine gene expression from visceral adipose tissue associated with body mass index or with survival. CONCLUSIONS: In horses presented with acute intestinal disease, increased adiponectin gene expression from retroperitoneal adipose tissue is associated with an increased risk of mortality. Obesity assessed by BMI had no association with increased post operative mortality in horses with primary gastrointestinal disease. POTENTIAL RELEVANCE: Further study is warranted on the expression and effects of adipokines, particularly adiponectin, and correlation with postoperative outcome.

Changes in management of the surgical colic patient over the last 30 years have resulted in considerable improvement in post operative survival rates. However, post operative complications remain common and these impact negatively on horse welfare, probability of survival, return to previous use and the costs of treatment. Multiple studies have investigated risk factors for post operative complications following surgical management of colic and interventions that might be effective in reducing the likelihood of these occurring. The findings from these studies are frequently contradictory and the evidence for many interventions is lacking or inconclusive. This review discusses the current available evidence and identifies areas where further studies are necessary and factors that should be taken into consideration in study design.

Impaction colic is the single most common type of colic diagnosed in a large population of donkeys (more than 2000 animals) at The Donkey Sanctuary, UK and the fatality rate from the disease is high. Here we identify risk factors for impaction colic in this population during 2006 using an unmatched case control study. There were 71 cases of impaction colic and multivariable analysis identified a number of variables associated with the disease. Management factors that increased the risk of impaction included paper bedding, feeding of concentrates, limited access to pasture and increasing number of carers. In addition health variables that were associated with an increased risk of impaction colic were weight loss, recent vaccination and a number of dental abnormalities. This study has identified variables which may help to identify donkeys at high risk of impaction colic; those with a history of weight loss and with concurrent dental pathology. Furthermore the study has also identified a number of variables that may be targeted to reduce the incidence of impaction colic in this donkey population, such as modification of concentrate feeding practices and pasture access.

REASONS FOR PERFORMING STUDY: Fatalities resulting from horse falls occurring during the cross-country phase of eventing competitions initiated epidemiological investigation of the risk factors associated with horse falls. OBJECTIVES: To identify variables that increased or decreased the risk of a horse fall during the cross-country phase of an eventing competition. METHODS: Data were collected from randomly selected British Eventing competitions held in Great Britain during 2001 and 2002. Data were obtained for 173 cases (jumping efforts resulting in a fall of the horse-and-rider partnership) and 503 matched controls (jumping efforts not resulting in a fall). The risk of falling was modelled using conditional logistic regression. RESULTS: An increased risk of a horse fall was associated with jumping into or out of water; taking off from good-to-soft, soft or heavy ground; fences with a drop landing; nonangled fences with a spread > or =2 m; and angled fences. Other risk factors included riders who knew that they were in the lead within the competition before the cross-country phase; an inappropriate speed of approach to the fence (too fast or too slow); horse-and-rider partnerships that had not incurred refusals at earlier fences; and riders who received cross-country tuition. CONCLUSIONS: This study has identified modifiable course- and fence-level risk factors for horse falls during the cross-country phase of eventing competitions. The risk of horse and rider injury at eventing competitions should be reduced by 3 simple measures; maintaining good to firm take-off surfaces at fences, reducing the base spread of fences to

REASON FOR PERFORMING STUDY: Epiploic foramen entrapment (EFE) is one of the most common causes of small intestinal strangulation in the horse. Identification of risk factors would generate hypotheses about causation and may suggest preventive strategies. HYPOTHESIS: Horses exhibiting certain behavioural patterns and those exposed to particular management practices are at increased risk of EFE. METHODS: A matched case-control study was conducted on EFE cases admitted to hospitals in the UK, Ireland and USA. Data on 109 cases and 310 control horses were obtained by telephone questionnaire and conditional logistic regression was used to identify associations between horse- and management-level variables and the risk of EFE. RESULTS: Crib-biting/windsucking behaviour was strongly associated with increased risk of EFE (OR 67.3, 95%CI 15.3-296.5). A history of colic in the previous 12 months (OR 4.4, 95%CI 1.5-12.7) and horses of greater height (OR/cm 1.05, 95%CI 1.01-1.08) were also at increased risk. The person(s) responsible for horses' daily care (nonowner/relative/spouse OR 5.5, 95%CI 2.3-13.3) and a number of behavioural features, including response to a stimulus causing fright (easily frightened OR 0.4, 95%CI 0.1-1.0) or excitement (sweats up easily/occasionally OR 0.3, 95%CI 0.1-0.8), reaction to their surroundings (inquisitive OR 0.4, 95%CI 0.2-0.8) and feeding behaviour when stressed (goes off food in full/part OR 0.3, 95%CI 0.1-1.0) were also associated with altered risk of EFE. CONCLUSIONS: The association between horses of greater height and those with a previous history of colic and increased risk of EFE suggests that some horses may be inherently predisposed to EFE. Furthermore, a behavioural pattern has been characterised that is common to horses at increased risk of EFE. Further research is required to investigate the causal pathway linking behavioural traits with gastrointestinal dysfunction and to determine whether behavioural modification reduces the risk of EFE. POTENTIAL RELEVANCE: The findings of the present study have relevance to horses in the UK, Ireland and USA.

Previous immunohistochemical studies targeting the receptor tyrosine kinase (c-Kit) have demonstrated an apparent reduction in the number of gastrointestinal pacemaker cells--the interstitial cells of Cajal (ICC)--in horses with intestinal motility disorders. This study compared the level of transcription of the c-kit gene encoding this receptor in horses with and without such motility disorders. Transcription levels of this gene were also compared to the density of ICC immunohistochemically positive for the c-Kit antigen. Intestinal samples were collected from 18 horses with intestinal disease and from 15 control animals. Following gene extraction and identification, real-time quantitative analysis of c-kit and a control gene, ACTB (β-actin), was carried out on all samples and the density of the c-Kit-positive ICC compared. There was a significant reduction in c-Kit immunoreactivity in the ICC of horses with large intestinal obstructive disorders relative to controls but no significant difference in the transcription of the c-kit gene between normal and affected animals. Further studies will be required to elucidate the mechanisms regulating c-Kit expression and to assess the pathophysiological significance of these findings.

Reasons for performing study: The intestinal bacterial community of the horse is a key determinant of intestinal and whole body health. Understanding the bacterial community structure and function is an important foundation for studies of intestinal health and disease. Objectives: To describe the faecal bacterial community and volatile organic compounds (VOCs) of the faecal metabolome of healthy Thoroughbred racehorses and to characterise responses to dietary supplementation with amylase-rich malt extract. Study design: Intervention study. Methods: Faecal samples were collected noninvasively before and 6 weeks after supplementation in 8 privately owned Thoroughbred racehorses in active race training. Faecal metabolome was characterised using thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS), with spectral analysis performed using AMDIS and compared against the NIST database. Taxonomic description of the faecal microbiota was achieved using error-corrected 454 pyrosequencing data from 16S rRNA gene amplicons. Results: The faecal metabolome of our study population was dominated by organic acids, alcohols and ketones. We identified 81 different VOCs only 28 of which were present in >50% of samples indicating functional diversity. Faecal VOC profiles differed between first and second sampling point, some VOCs being significantly reduced post supplementation, consistent with a marked response to dietary amylase-rich malt extract. Faecal microbiota was characterised as highly diverse; samples demonstrated verifiable diversity in the range 1200-3000 operational taxonomic units (OTUs) per individual. The methods used also describe high levels of infrequent, low abundance OTUs. Faecal microbial community structure was found to be different following dietary supplementation. Differences in several low abundance bacterial taxa were detected and also some evidence of interhorse variation in response. Conclusions: The volatile faecal metabolome of Thoroughbred racehorses is dominated by organic acids, alcohols and ketones; this study demonstrates that dietary supplementation with amylase-rich malt extract may significantly alter the profile of VOCs. The faecal microbiome is highly diverse, dominated by Firmicutes and Bacteroidetes. Small but significant changes in microbial community structure were detected following dietary supplementation. This study describes the faecal metabolome and microbiome of healthy Thoroughbred racehorses against which future studies of disease and dietary intervention can be benchmarked.

REASONS FOR PERFORMING STUDY: Epiploic foramen entrapment (EFE) has been associated with a particularly poor post operative prognosis for equine colic cases, but the reasons for this are unknown. OBJECTIVES: To identify variables associated with post operative survival following surgery for small intestinal disease; develop a model describing long-term post operative survival; and identify reasons for the poor prognosis associated with EFE. METHODS: Data from 382 horses undergoing surgery were used to identify variables associated with survival. A multivariable Cox proportional hazards model for post operative survival was developed and model fit evaluated. RESULTS: The final model included the variables total plasma protein (TP) and packed cell volume (PCV) at admission, duration of surgery and the dichotomous variable relaparotomy (yes/no). Risk of death was positively associated with increasing PCV, but negatively associated with increasing TP (which decreased the probability of death). In a univariable model, EFE cases had a significantly higher death rate than other types of small intestinal disease (hazard ratio = 1.7, P = 0.035). Multivariable modelling indicated that some of the increased risk associated with EFE cases was due to lower TP values and longer duration of surgery. CONCLUSIONS: Preoperative TP is associated negatively with the risk of post operative death in horses recovering from small intestinal surgery. Other variables associated with the probability of survival are preoperative PCV, duration of surgery and relaparotomy. The increased post operative death rate of EFE cases can be explained in part by lower TP and longer surgery times of these cases. POTENTIAL RELEVANCE: Total plasma protein may be not simply a measure of hydration status in small intestinal colic cases, but an important determinant of survival. Further investigation of this relationship is warranted. Our model for post operative survival highlights the importance of preoperative TP, PCV and duration of surgery as prognostic indicators. This information should allow a more accurate post operative prognosis following small intestinal surgery.

Experimental and epidemiological evidence suggests that consumption of hydrolyzable carbohydrate, hCHO (grain), by horses is an important risk factor for colic, a common cause of equine mortality. It is unknown whether the small intestinal capacity to digest hCHO and/or to absorb monosaccharides is limiting, or even if horses can adapt to increased carbohydrate load. We investigated changes in the brush-border membrane carbohydrate digestive enzymes and glucose absorptive capacity of horse small intestine in response to increased hCHO. Expression of the Na(+)/glucose co-transporter, SGLT1, was assessed by Western blotting, immunohistochemistry, Northern blotting, QPCR, and Na(+)-dependent D-glucose transport. Glucose transport rates, SGLT1 protein, and mRNA expression were all 2-fold higher in the jejunum and 3- to 5-fold higher in the ileum of horses maintained on a hCHO-enriched diet compared to pasture forage. Activity of the disaccharidases was unaltered by diet. In a well-controlled study, we determined SGLT1 expression in the duodenal and ileal biopsies of horses switched, gradually over a 2-month period, from low (

BACKGROUND: Colic is an important cause of mortality and morbidity in domesticated horses yet many questions about this condition remain to be answered. One such question is: does season have an effect on the occurrence of colic? Time-series analysis provides a rigorous statistical approach to this question but until now, to our knowledge, it has not been used in this context. Traditional time-series modelling approaches have limited applicability in the case of relatively rare diseases, such as specific types of equine colic. In this paper we present a modelling approach that respects the discrete nature of the count data and, using a regression model with a correlated latent variable and one with a linear trend, we explored the seasonality of specific types of colic occurring at a UK referral hospital between January 1995-December 2004. RESULTS: Six- and twelve-month cyclical patterns were identified for all colics, all medical colics, epiploic foramen entrapment (EFE), equine grass sickness (EGS), surgically treated and large colon displacement/torsion colic groups. A twelve-month cyclical pattern only was seen in the large colon impaction colic group. There was no evidence of any cyclical pattern in the pedunculated lipoma group. These results were consistent irrespective of whether we were using a model including latent correlation or trend. Problems were encountered in attempting to include both trend and latent serial dependence in models simultaneously; this is likely to be a consequence of a lack of power to separate these two effects in the presence of small counts, yet in reality the underlying physical effect is likely to be a combination of both. CONCLUSION: The use of a regression model with either an autocorrelated latent variable or a linear trend has allowed us to establish formally a seasonal component to certain types of colic presented to a UK referral hospital over a 10 year period. These patterns appeared to coincide with either times of managemental change or periods when horses are more likely to be intensively managed. Further studies are required to identify the determinants of the observed seasonality. Importantly, this type of regression model has applications beyond the study of equine colic and it may be useful in the investigation of seasonal patterns in other, relatively rare, conditions in all species.

Cyathostomins are important equine gastrointestinal parasites. Mass emergence of mucosal stage larvae causes a potentially fatal colitis. Mucosal stages are undetectable non-invasively. An assay that would estimate mucosal larval stage infection would greatly assist in treatment, control and prognosis. Previously, we identified two putative diagnostic antigens (20 and 25 kDa) in somatic larval preparations. Here, we describe their purification and antigen-specific IgG(T) responses to them. Western blots confirmed the purity of the antigens and showed that epitopes in the 20 kDa complex were specific to larval cyathostomins. No cross-reactive antigens appeared to be present in Parascaris equorum or Strongyloides westeri species. Low levels of cross-reactivity were observed in Strongylus edentatus and Strongylus vulgaris species. Use of purified antigens greatly reduced background binding in equine sera. These results indicate that both antigen complexes may be of use in a diagnostic assay.

The characteristics of, and pre-existing pathology associated with 75 cases of fatal lateral condylar fracture sustained by Thoroughbreds while racing in the UK were described. Cases were identified from 220 cases of fatal distal limb fracture submitted as part of studies designed to identify risk factors for all fatal distal limb fractures. Fractures were most common in hurdle races and affected the right forelimb twice as often as the left forelimb. Fracture dimensions were similar to previous reports, however there was a much greater prevalence of articular and diaphyseal comminution and of concurrent fractures in the current report. Pre-existing pathology was particularly common in the medial and lateral parasagittal grooves of the distal articular surfaces of the third metacarpus/metatarsus. The degree of this pathology was not associated with horse age, length of career or number of career starts.

REASONS FOR PERFORMING STUDY: Considerable variation in the rates of equine fatality at different racecourses draws attention to probable risk factors at the level of course or race that might be partly responsible. Distal limb fractures are the most common cause of equine fatality on UK racecourses and identification of risk factors for such injuries and subsequent implementation of intervention strategies could significantly reduce the total number of racecourse fatalities. OBJECTIVES: To identify race- and course-level risk factors for fatal distal limb fracture in Thoroughbreds on UK racecourses. METHODS: A case-control study design was used. Case races were defined as those in which one or more horses sustained fatal fracture of the distal limb. Controls were selected in 2 different ways. Firstly, 3 races in which no fracture occurred were selected from all races of the same type held within 5 days of the case race (Analysis 1). Secondly, 3 control races were selected for each case race from all races of the same type held in the same year (Analysis 2). One hundred and nine cases were included in the study. Information about the race and the racecourses was collected from Computer Raceform. Conditional logistic regression was used to identify the relationship between a number of independent variables and the likelihood of fracture in a race. RESULTS: Longer races with a larger number of runners were more likely to contain a fracture. Firmer going and fewer days since the last race on the same course were associated with an increased risk of fracture. The going at the course at the previous race meeting was also associated with the likelihood of fracture. CONCLUSIONS: Modifications to the going on the day of a race and greater emphasis on ground maintenance between race meetings may have an impact on the risk of fatal distal limb fracture during racing. POTENTIAL RELEVANCE: Modification of risk factors such as the going and number of days since the last race meeting could reduce the number of equine fatalities on UK racecourses. The condition of the racecourse may be an important risk factor and future research should focus on the identification of course maintenance techniques that produce the safest possible racing surfaces.

To investigate cytokine responses in cyathostomin infection, we quantified mucosal interleukin-4 (IL-4), interleukin-10 (IL-10), tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma by reverse transcriptase-competitive polymerase chain reaction. The analysis was performed on large intestinal wall samples obtained from six anatomical sites spanning the caecum and colon of 17 naturally exposed horses. The numbers of developing larvae (DL) and early third stage larvae (EL3) were ascertained using transmural illumination and pepsin digestion techniques, respectively. Levels of each cytokine transcript were correlated with local intestinal wall burdens of Cyathostominae larvae. IL-4 and IL-10 levels showed significant correlations with EL3 and DL burdens at several sites. No significant correlations were observed with IFNgamma. A pro-inflammatory response, typified by detection of TNFalpha transcript, was observed at a few sites in some horses with inflammatory enteropathy associated with emerging or emerged larvae. However, this cytokine was measured at an insufficient number of sites to enable statistical analysis. Levels of IL-4, IL-10 and IFNgamma transcript were compared between two groups: one group consisting of horses with low to high mucosal burdens (Group A) and the other, of horses with negative/negligible mucosal burdens (Group B). Significant differences in IL-4 (P

Risk factors for large colon volvulus (LCV) in the horse have not been previously reported. Knowledge of these risk factors may allow the introduction of measures that could be taken to minimise the incidence of LCV.

A prospective cohort study was conducted on horses starting in hurdle and steeplechase races on six UK racecourses in 2000 and 2001. Trainers or carers were questioned on the horses' pre-race routine and observational data were collected in the stables and parade ring. Some practices were common to many starters, such as withholding food and water before racing whereas other practices, such as schooling frequency, were more variable. There was a total of 2879 starts and a total of 83 injuries or medical events (28.8/1000 starts). The commonest types of injury were tendon/suspensory injuries and lacerations/wounds. Multivariable logistic regression models were used to identify the relationship between predictor variables and the risk of injury. Risk of injury or medical event was associated with distance of the race and weight carried. The risk of injury, excluding medical events, was associated with the speed of the race and foot conformation.

Background. Perioperative lidocaine treatment is commonly used in horses that undergo surgical treatment of colic, to prevent or treat postoperative ileus and reduce the effects of intestinal ischaemia-reperfusion injury. However, its clinical efficacy has not been evaluated in a large population of horses undergoing small intestinal surgery. The aim of the current study was to evaluate whether systemic lidocaine administration reduced the prevalence, volume and duration of postoperative reflux and improved rates of survival following surgical treatment of small intestinal lesions. Data were collected as a part of two prospective studies investigating postoperative survival of surgical colic patients admitted to a UK equine referral hospital during the periods 2004–2006 and 2012–2014. Kaplan-Meier plots of cumulative probability of survival and the log-rank test were used to compare survival between horses that did or did not receive lidocaine. The Wilcoxon rank-sum test was used to compare the total reflux volume and duration of reflux between the groups. A multivariable Cox proportional hazards model was used to identify pre- and intraoperative risk factors for non-survival. Results: Data from 318 horses were included in the final analysis. The overall prevalence of postoperative reflux was 24.5 %. This was significantly higher (34.8 %) in horses admitted in 2012–2014 compared to the 2004–2006 cohort (16.7). Perioperative lidocaine treatment had no effect on total reflux volume, duration of reflux or rates of postoperative survival nor was it a risk factor associated with altered postoperative survival. Variables identified to be associated with increased risk of postoperative mortality included packed cell volume on admission (hazard ratio [HR] 1.03 95 %, 95 % confidence interval [CI] 1.004–1.06, p = 0.024), heart rate on admission (HR 1.014, 95 % CI 1.004–1.024, p =0.008) and duration of surgery (HR 1.007, 95 % CI 1.002–1.01, p = 0.008). Conclusions: Lidocaine therapy had no effect on the prevalence of postoperative reflux, total reflux volume and duration of reflux nor did it have any effect on postoperative survival in horses undergoing surgical management of small intestinal disease for treatment of colic. There is a need for a well-designed multicentre, prospective randomised controlled trial to fully investigate the efficacy of lidocaine across different hospital populations.

We identified risk factors associated with falling during steeplechase racing. We used retrospective data from all steeplechase runs on UK racecourses during 1999: 10,866 starts with 647 horse falls. The relationship between continuous variables and falling was assessed using generalised additive models (GAMs). Polynomial fits then were included in a multilevel, multivariable logistic-regression model. The number of runners had a linear, positive association with the risk of falling. The distance of the race had a non-linear relationship with the risk of falling; the risk steadily increased in races up to 23 furlongs (1furlong approximately equals 198 m), and then decreased in longer races. Age also had a significant, non-linear relationship with the risk of falling: a decreasing risk up to 12 years of age followed by an increasing risk in older horses. Horses that wore visors and had raced previously were associated with a decrease in the risk of falling. Intra-class correlation coefficients (ICCs) showed that although most of the variation resided at the start (level 1), a proportion of variation in the risk of falling could be attributed to horse and race. Trainer and jockey contributed very little to the variation in the risk of falling.

The efficacy of an oral formulation of praziquantel (Equitape, Horse paste, Fort Dodge) in the reduction of cestode egg counts and serum antibody level against Anoplocephala perfoliata was assessed in 44 donkeys under field conditions. The donkeys were confirmed both by faecal examination and serum antibody assessed by an enzyme-linked immunosorbent assay to have natural infection with tapeworms. The donkeys were randomly allocated into treatment (n = 22) and control (n = 22) groups. The treatment group was treated with both praziquantel and ivermectin (Ivomec, Merial) at a dose rate of 1 mg/kg and 200 μg/kg, respectively while the control group was treated only with ivermectin. Faecal samples were collected before treatment (day-0) and 2, 6, 8, 12, and 16 weeks post-treatment while blood samples were collected before treatment and 8 and 16 weeks after treatment and analysed. The results of the study demonstrated that praziquantel paste was highly effective in reducing cestode eggs in donkeys and had an efficacy of more than 99 % until week 16 (day 112). No cestode egg reappearance by 16 weeks post-treatment in any animal in the treatment group was observed while donkeys in the control group continued shedding cestode eggs. The immunological assay also showed a significant reduction in serum antibody level against A. perfoliata in treated donkeys compared to the control group (p = 0.0001). This marked decrease in serum antibody level indicates reduced risk of cestode-associated colic and other gastrointestinal disorders and clinical diseases. No adverse reactions or clinical effects were encountered in any animal within either group throughout the trial period. © 2012 Springer-Verlag.

Gene editing and subsequent cloning techniques offer great potential not only in genetic disease correction in domestic animals but also in livestock production by enhancement of desirable traits. The existence of the technology, however, leaves it open to potential misuse in performance-led sports such as horseracing and other equestrian events. Recent advances in equine gene editing, regarding the generation of gene-edited embryos using CRISPR/Cas9 technology and somatic cell nuclear transfer, have highlighted the need to develop tools to detect potential prohibited use of the technology. One possible method involves the characterisation of the mitochondrial genome (which is not routinely preserved during cloning) and comparing it with the sequence of the registered dam. We present here our approach to whole-mitochondrial sequencing using tiled long-range PCR and next-generation sequencing. To determine whether the background mutation rate in the mitochondrial genome could potentially confound results, we sequenced 10 sets of dam and foal duos. We found variation between duos but none within duos, indicating that this method is feasible for future screening systems. Analysis of WGS data from over 100 Thoroughbred horses revealed wide variation in the mitochondria sequence within the breed, further displaying the utility of this approach.

REASONS FOR PERFORMING STUDY: Falls during racing present a risk of injury to both horse and jockey and a risk of fatality to horses. OBJECTIVES: To use video recordings of races to describe the circumstances surrounding horse falls at hurdle and steeplechase fences and to identify and quantify within-race risk factors for horse falls in National Hunt racing in the UK. METHODS: A retrospective, matched, nested case-control study using video recordings of races was conducted on 6 UK racecourses. Cases and controls were matched on both race type and jump number at which the fall occurred. Conditional logistic regression analysis was used to examine the univariable and multivariable relationship between predictor variables and the risk of falling. RESULTS: The risk of falling was significantly associated with whip use and race progress. Horses which were being whipped and progressing through the race were at greater than 7 times the risk of falling compared to horses which were not being whipped and which had no change in position or lost position through the field. CONCLUSIONS: This study has identified whip use and the position of the horse with respect to others in the field as potential risk factors for horse falls. POTENTIAL RELEVANCE: If these findings are confirmed by the use of intervention trials (e.g. with whip-free or restricted whip use races), modifications could be introduced which would reduce the frequency of horse falls, leading to improved equine welfare.

The objectives of this study were (1) to determine the prevalence of pathological abdominal adhesion formation following exploratory laparotomy; (2) to establish the site of adhesion formation and its relationship to the initial lesion; (3) to ascertain whether the development of intra-abdominal adhesions decreases long-term survival and (4) to identify risk factors for adhesion formation. Of 1014 horses treated surgically for acute gastrointestinal disease, 113 (10.1%) were subjected to repeat laparotomy, with surgical records available for 99 of these cases. Pathological adhesions were the most common diagnosis at repeat laparotomy (28%), followed by complications associated with the anastomosis (16%). Adhesions were not associated with the site of the primary lesion, resection, or endotoxaemia, consistent with the hypothesis that surgical trauma is the most important stimulus in adhesion formation. Together these findings strongly support the need for pan-abdominal, rather than site-specific adhesion prevention measures in all horses undergoing exploratory laparotomy.

A concurrent case-control study of 12 UK racecourses was made between March 1, 2000, and August 31, 2001, to identify and quantify the factors associated with the risk of horses falling in steeplechase races. Cases were defined as a jumping effort at a steeplechase fence that resulted in a fall and controls were defined as a successful jumping effort over any steeplechase fence at any of the 12 racecourses within 14 days before or after the case fall. Information on the horse, the jockey and the race were collected and all the fences on all the courses were surveyed. Conditional logistic regression was used to examine the relationships between the predictor variables and the risk of falling. There was one fall per 254 jumping efforts. The risk of a horse falling decreased the more times it had raced on a particular racecourse. The number of fences, the distance from the previous fence and the nature of the previous fence also affected the risk of falling. If the previous fence was a water jump the risk of falling increased; fences that were sited on flat or slight uphill gradients (up to approximately 1 in 25) were associated with a lower risk of horses falling than downhill fences, and higher takeoff boards were associated with a higher risk of falling.

Intestinal helminths are an important cause of equine disease. Of these parasites, the Cyathostominae are the commonest group that infect horses. These nematodes consist of a complex tribe of 51 species, although individual horses tend to harbour 10 or so common species, in addition to a few rarer species. The Cyathostominae can be extremely pathogenic, and high levels of infection result in clinical symptoms ranging from chronic weight loss to colic, diarrhoea and death. As part of their life cycle, immature cyathostomins penetrate the large intestinal wall, where they can enter a state of inhibited larval development. These larvae can exist in this state for months to years, after which they subsequently re-emerge. If larvae re-emerge in large numbers (i.e. several million), severe pathological consequences ensue. The inhibited larvae are also relatively refractory to several of the currently available anthelmintics, so that horses treated previously with anthelmintics can still carry life-threatening burdens of these parasitic stages. Little is known about the cyathostomin larvae during their mucosal phase, and current research efforts are focused on investigating the biology of these stages. Much of the research described here highlights this area of research and details studies aimed at investigating the host immune responses that the mucosal larvae invoke. As part of this research effort, molecular tools have been developed to facilitate the identification of larval and egg stages of cyathostomins. These molecular tools are now proving very useful in the investigation of the relative contributions that individual, common cyathostomin species make to the pathology and epidemiology of mixed helminth infections. At the more applied level, research is also in progress to develop an immunodiagnostic test that will allow numbers of mucosal larvae to be estimated. This test utilises antigen-specific IgG(T) serum antibody responses as markers of infection. As anthelmintic resistance will be the major constraint on the future control of the Cyathostominae, researchers are now actively investigating this area and studies aimed at elucidating the molecular mechanisms of drug resistance are described. Another parasite which has assumed a clinically important role in horses is the tapeworm, Anoplocephala perfoliata. This parasite is prevalent world-wide and has been shown to be a significant cause of equine colic. Because previous methods of estimating the infection intensity of tapeworm were inaccurate, recent research has been directed at developing an immunodiagnostic ELISA for these cestodes. Specific IgG(T) responses to antigens secreted by adult tapeworms have been shown to provide a reasonable indication of infection intensity. An ELISA based on these responses is now commercially available. The steps involved in the development of this ELISA are described here. In addition to these recent advances in research, this review also outlines the principle areas for future research into these important equine parasites.

REASONS FOR PERFORMING STUDY: The gastrointestinal pacemaker cells, the interstitial cells of Cajal (ICC), have been implicated in several human gastrointestinal dysmotility syndromes. Recently, the involvement of these cells in equine gastrointestinal diseases has been investigated in cases of equine grass sickness where a significant reduction in ICC density was observed. OBJECTIVE: To investigate ICC density in equine obstructive gastrointestinal disorders using immunohistochemical labelling methods. METHODS: Intestinal samples were analysed from 44 horses undergoing exploratory surgery for colic and from 11 control animals subjected to euthanasia for conditions not related to the gastrointestinal tract. Immunohistochemical labelling of ICC was carried out using an anti-c-Kit antibody. Two independent observers assessed ICC density using a semiquantitative grading system. RESULTS: There was a significant reduction in ICC density in horses with large colon disorders compared to the controls (P

We used data from a case-control study investigating risk factors for horse falls in the cross-country phase of eventing in Great Britain (GB) to examine evidence for memory decay and information bias. Responses to two questions obtained by telephone for 173 cases and 521 controls were examined for evidence of differential reporting according to the respondent's case-control status and performance in the dressage and cross-country phases of competitions. Information bias was found in the accuracy of reporting dressage penalty scores when analysed as a function of performance level (good/poor). Poor dressage performers were less likely to report accurate dressage scores than good performers. The accuracy of reporting dressage scores decreased as the time between the event and questionnaire completion increased, with no case-control interaction. Competitors who incurred cross-country jumping penalties at the event preceding the selected event reported their cross-country scores with less accuracy when compared with competitors who incurred no penalties. No information bias was found when the reporting of dressage and cross-country scores were analysed as a function of respondent category (case/control).

A novel serological assay which measures IgG(T) specific for a 12/13 kDa protein of the equine tapeworm Anoplocephala perfoliata was used as part of a colic outbreak investigation. A training/rehabilitation yard for Thoroughbreds and Arabs was found to have an increasing incidence of colic over a 5 year period, culminating in a peak incidence of 1.15 episodes/horse year at risk. Four animals suffered from ileal impaction colic which necessitated surgical management. A case-control study design suggested a strong association between tapeworm infection and colic, with evidence of a dose-response relationship. Intervention, in the form of anticestode anthelminthics, coincided with a decrease in the incidence of colic and a fall in anti-12/13 kDa IgG(T) antibody levels of 8 horses monitored post-treatment. This study demonstrates that anthelminthic regimens, using exclusively ivermectin, may lead to an increase in tapeworm infection intensity which may in turn lead to an increased incidence of colic. Furthermore, it provides support to the hypothesis that the risk of ileal impaction colic and spasmodic colic increases with tapeworm infection intensity. The practical application of the anti-12/13 kDa IgG(T) ELISA is demonstrated by this study.

Equine grass sickness (EGS) is a largely fatal, pasture-associated dysautonomia. Although the aetiology of this disease is unknown, there is increasing evidence that Clostridium botulinum type C plays an important role in this condition. The disease is widespread in the United Kingdom, with the highest incidence believed to occur in Scotland. EGS also shows strong seasonal variation (most cases are reported between April and July). Data from histologically confirmed cases of EGS from England and Wales in 1999 and 2000 were collected from UK veterinary diagnostic centres. The data did not represent a complete census of cases, and the proportion of all cases reported to the centres would have varied in space and, independently, in time. We consider the variable reporting of this condition and the appropriateness of the space-time K-function when exploring the spatial-temporal properties of a 'thinned' point process. We conclude that such position-dependent under-reporting of EGS does not invalidate the Monte Carlo test for space-time interaction, and find strong evidence for space time clustering of EGS cases (P < 0.001). This may be attributed to contagious or other spatially and temporally localized processes such as local climate and/or pasture management practices.

Reasons for performing study: Endotoxaemia is frequently presumed on the basis of clinical signs in horses with colic. Objective: Measurements of plasma endotoxin (LPS) are rarely made in clinical cases and there is little information on the correlations between this variable, clinical variables and outcomes. Objectives: To measure LPS levels in plasma of horses presented to the Philip Leverhulme Equine Hospital on admission and daily for up to 4 days and to relate LPS levels to selected clinical parameters, such as heart rate and packed cell volume, and outcomes. Methods: Blood samples were collected and stored at -20°C prior to assay of the plasma using a validated kinetic chromogenic Limulus amoebocyte lysate (LAL) assay. Clinical parameters and outcome variables were collected from hospital records. Associations were determined by Chi-squared test and logistic regression analysis. Results: Daily blood samples were collected from 234 horses. LPS was detected in 26.5% of the study population and in 29% of those horses presented for colic. Horses providing samples with detectable LPS were more likely to die whilst in the hospital than those that did not (P = 0.045). Horses presenting with colic were more likely to have detectable LPS in their plasma than noncolic cases (P = 0.037), although LPS was detected in the plasma of 8 out of 42 noncolic horses. A horse that did not meet the study definition of clinical endotoxaemia was 10 times less likely to provide a positive LPS sample (OR 0.10, 95% CI: 0.05-0.22). Conclusions: The proportion of horses providing samples with detectable LPS was similar to other studies. Potential relevance: LPS was detected in the minority of horses presented with colic. Increased levels of LPS positively correlated with packed cell volume and with risk of mortality in colic cases. © 2011 EVJ Ltd.

The objectives of the study were to determine whether retrospective data can be used to answer questions about the efficacy of prokinetic agents when used to treat horses with post-operative ileus (POI). We describe prevalence and mortality of POI with reference to treatment with four prokinetic agents. By combining data from two Hospitals a study population of 55 horses with POI following pedunculated lipoma obstruction (PLO) was established. Univariable and multivariable associations were determined between short term survival and potential explanatory variables. With death as outcome in multivariable models, breed and hospital were significantly associated with outcome but the use of prokinetic agents was not (P=0.15). However, sample size estimates indicate the low power of this study to detect differences in outcome. It was not possible definitively to evaluate the efficacy of prokinetics as treatment for POI following PLO using retrospective data. The data were suggestive of limited efficacy of prokinetics as treatment for POI. It is postulated that the identified association between hospital and survival reflects differences in clinician decision making. The study highlights the need for further prospective studies using randomised clinical trials to evaluate accurately the efficacy of prokinetic agents. This report illustrates difficulties with performing retrospective analysis of clinical data to determine the efficacy of treatment regimes.

Postoperative complications and mortality can occur many weeks or months after colic surgery. We are interested in the long-term outcome of these cases. This study documents patterns of mortality and morbidity among 341 horses that recovered from colic surgery March 1998-August 2000. The progress of each horse was rigorously followed by periodic telephone and postal questionnaires. Event time data were recorded for each animal and a total of 321 horse years of survival, together with death from all causes, colic-related death and various postoperative complications. Postoperative survival (of all horses excluding grass sickness cases) was triphasic over the first 600 days and there was marked mortality in the first 10 days postoperatively. The probability of survival postoperatively decreased to 0.87 by 10 days, 0.82 by 100 days and declined slowly to 0.75 at 600 days. Horses suffering from epiploic foramen entrapment had a significantly reduced probability of postoperative survival (RR = 2.1, P = 0.033). The causes of death for 104 horses that died postoperatively and the prevalence of postoperative complications are recorded for the study population. Postoperative colic was the most prevalent complication with 100 horses (29%) suffering one or more episodes. However, only 16 horses (4.6%) suffered 3 or more episodes. The incidence of postoperative colic was 0.55 episodes/horse year at risk. This study provides data that will inform the prognosis for postoperative colic cases and identifies epiploic foramen entrapment as carrying a worse prognosis for survival than other strangulating lesions.

REASONS FOR PERFORMING STUDY: Equine grass sickness (EGS) occurs repeatedly on certain premises over time. Few studies have sought, or identified, the determinants of this phenomenon in order to inform advice on disease prevention strategies. HYPOTHESIS: Premises-level risk factors are important determinants of whether EGS occurs. METHODS: A matched case-control study was undertaken. Sixty premises giving rise to one or more histologically confirmed case of EGS and 120 time-matched control premises were sampled. Data were collected on pasture management, soil nutrient content, pasture nutrient content and local weather conditions for 2 weeks prior to the onset of disease. Data were analysed by conditional logistic regression. RESULTS: Multivariable modelling identified an association between EGS and increased soil nitrogen content, pasture disturbance and previous occurrence of EGS on the premises. None of the meteorological variables recorded in this study were significantly associated with EGS occurrence. No relationship between certain management practices (e.g. harrowing, fertilisation, reseeding) and the risk of EGS was detected. CONCLUSIONS AND POTENTIAL RELEVANCE: This information is useful in understanding the causal pathway of EGS and may be used in the formulation of evidence-based disease avoidance strategies.

Gene doping in horses is a threat to the fairness in sport and has serious implications for animal welfare. To investigate the effect of long-term storage on the detection of AAV in plasma and whole blood, samples from an administration study using an adeno-associated virus serotype 6 expressing green fluorescence protein (AAV6-GFP) were stored at -20 & DEG;C for 8 months before analysis. The AAV vector was detected in stored plasma samples, following the same detection profile as the fresh plasma samples. The stored blood showed lower overall DNA detection but followed the same detection profile as the plasma samples. This study provides confidence that re-analysing plasma samples and/or analysing a frozen 'B' sample with different matrix such as whole blood after prolonged storage will still result in the detection of gene doping material. Equine whole blood and plasma samples from an administration of AAV-GFP viral vector were stored at -20 & DEG;C for 8 months and analysed with qPCR and massively parallel sequencing. The vector detection in stored plasma samples followed the same detection profile as the fresh plasma samples with minimal degradation. Stored blood samples showed lower overall DNA detection but followed the same detection profile as the plasma samples. This study provides confidence in the detection of gene doping material when re-analysing samples and/or analysing 'B' samples for confirmatory analysis.image

Aquaporins (AQPs) play fundamental roles in water and osmolyte homeostasis by facilitating water and small solute movement across plasma membranes of epithelial, endothelial, and other tissues. AQP proteins are abundantly expressed in the mammalian kidney, where they have been shown to play essential roles in fluid balance and urine concentration. Thus far, the majority of studies on renal AQPs have been carried out in laboratory rodents and sheep; no data have been published on the expression of AQPs in kidneys of equines or other large mammals. The aim of this comparative study was to determine the expression and nephron segment localization of AQP1-4 in Equus caballus by immunoblotting and immunohistochemistry with custom-designed rabbit polyclonal antisera. AQP1 was found in apical and basolateral membranes of the proximal convoluted tubules and thin descending limbs of the loop of Henle. AQP2 expression was specifically detected in apical membranes of cortical, medullary, and papillary collecting ducts. AQP3 was expressed in basolateral membranes of cortical, medullary, and papillary collecting ducts. Immunohistochemistry also confirmed AQP4 expression in basolateral membranes of cells lining the distal convoluted and connecting tubules. Western blots revealed high expression of AQP1-4 in the equine kidney. These observations confirm that AQPs are expressed in the equine kidney and are found in similar nephron locations to mouse, rat, and human kidney. Equine renal AQP proteins are likely to be involved in acute and chronic regulation of body fluid composition and may be implicated in water balance disorders brought about by colic and endotoxemia.

Until recently, the equine tapeworm Anoplocephala perfoliata was difficult to diagnose and considered to be of questionable pathogenicity. Here, Chris Proudman and Sandy Trees describe recent advances in the immunodiagnosis of this parasite that have facilitated epidemiological studies. These studies suggest that A. perfoliata may be an important cause of intestinal disease in the horse and demonstrate a dose-response relationship between infection intensity and risk of disease. If tapeworm infection is a risk factor for ileocaecal colic, the identification and treatment of infected individuals would be a rational approach to disease prevention.

REASONS FOR PERFORMING STUDY: Fractures below the level of the radius or tibia (distal limb fractures) are the most common cause of equine fatality on UK racecourses; however, little is known about their epidemiology or aetiology. Identification of risk factors could enable intervention strategies to be designed to reduce the number of fatalities. OBJECTIVES: To identify horse-level risk factors for fatal distal limb fracture in Thoroughbreds on UK racecourses. METHODS: A case-control study design was used. Fractures in case horses were confirmed by post mortem examination and 3 matched uninjured controls were selected from the race in which the case horse was running. One hundred and nine cases were included and information was collected about previous racing history, horse characteristics and training schedules. Conditional logistic regression was used to identify the relationship between a number of independent variables and the likelihood of fracture. RESULTS: Horses doing no gallop work during training and those in their first year of racing were at significantly increased risk of fracture on the racecourse. Case horses were also more likely to have trained on a sand gallop, i.e. a gallop described by trainers as being primarily composed of sand. CONCLUSIONS: Modifications to training schedules, specifically within the first year of racing, may have a large impact on the risk of fatal distal limb fracture on the racecourse. Horses should do some gallop work in training and our results suggest that the minimum distance galloped should be between 805-2012 m (4-10 furlongs)/week. POTENTIAL RELEVANCE: The information from this study can be used to alter training schedules in an attempt to reduce the incidence of fatal distal limb fracture in Thoroughbred racehorses. Training should include some gallop work, and further studies, recording the exact level of work, will help to identify an optimum range of training speeds and distances which will reduce the liklihood of catastrophic fracture on the racecourse.

REASONS FOR PERFORMING STUDY: There is limited information about risk factors and survival associated with disorders of the small colon requiring surgical management. OBJECTIVES: 1) To document the types of surgical lesion in horses where pathology of the small (descending) colon was the primary cause of abdominal pain, 2) to describe the short- and long-term survival of these cases and identify factors associated with survival and 3) to identify preoperative variables associated with localisation of a lesion to the small colon. METHODS: Clinical data and long-term follow-up were obtained for horses that underwent exploratory laparotomy over a 10-year period. Descriptive data were generated for short- and long-term survival and survival analysis performed to identify factors associated with reduced survival. Univariable and multivariable relationships were explored using a Cox proportional hazards model. Preoperative factors associated with increased likelihood of a small colon lesion were explored, using controls randomly selected from horses undergoing exploratory laparotomy for treatment of colic unrelated to the small colon. RESULTS: The study population included 84 horses. Of horses with small colon lesions recovered from anaesthesia, the percentage that survived until discharge, one year and 2 years following surgery, was 91.0, 81.0 and 73.5%, respectively. Median survival time for horses in which a resection had been performed was 1029 vs. 3072 days in the nonresection group. Small colon cases were more likely to have shown a longer duration of colic signs prior to admission (P

OBJECTIVE: To determine the effects of dobutamine and phenylephrine on intra- and postoperative survival in horses undergoing emergency abdominal surgery. STUDY DESIGN: Retrospective case analysis. ANIMALS: A total of 637 client-owned horses undergoing colic surgery. METHODS: Clinical details of horses admitted for colic surgery were recorded on a computer database. Information collected included history, clinical variables observed before surgery, anaesthesia and surgical details, and postoperative survival and morbidity rates. Details of specific importance for this study were those recorded during anaesthesia, in particular the duration of dobutamine and phenylephrine administration, separately and combined, and total anaesthesia time. Two outcomes were considered: 1) intra-operative death, i.e. death between time of pre-anaesthetic medication and recovery from anaesthesia (defined as horse walking from recovery box); and 2) all deaths, i.e. death at any time after induction of anaesthesia. The definition of 'death' included euthanasia. Univariable and multivariable statistical analyses were performed to evaluate the associations between dobutamine and/or phenylephrine use and these two outcomes. RESULTS: Results from univariable analyses suggested that dobutamine administration was not significantly associated with increased intra- or postoperative mortality. Phenylephrine administration showed univariable association with intra- and postoperative death. However, in multivariable models adjusted for the effects of heart rate and packed cell volume at admission, the phenylephrine effect was not significantly associated with intra-operative, or other types of death. CONCLUSION: This study provides no evidence to suggest that dobutamine or phenylephrine administration is associated with altered survival rates during or after colic surgery. CLINICAL RELEVANCE: Our study supports previous work, suggesting that pre-existing cardiovascular status is an important prognostic determinant in equine colic cases. It provides no evidence that dobutamine or phenylephrine administration is associated with survival.

Mortality rates for horses that have undergone emergency abdominal surgery are higher than for other procedures. Here, multivariable modelling of data from 774 surgical colic cases is used to identify pre-operative and anaesthesia-related variables associated with intra- and post-operative mortality. Intra-operative mortality was significantly (P

The misuse of gene therapy by the introduction of transgenes via plasmid or viral vectors as a doping agent is an increasing concern in human and animal sports, not only in consideration to fair competition but also in potential detrimental effects to welfare. Doping events can be detected by polymerase chain reaction (PCR) amplification of a transgene-specific region of DNA. Quantitative real-time PCR (qPCR) is particularly suited to confirmatory investigations where precise limits of detection can be calculated. To fully validate a qPCR experiment, it is highly desirable to confirm the identity of the amplicon. Although post-PCR techniques such as melt curve and fragment size analysis can provide strong evidence that the amplicon is as expected, sequence identity confirmation may be beneficial as part of regulatory proceedings. We present here our investigation into two alternative processes for the direct assessment of qPCR products for five genes using next-generation sequencing: ligation of sequence-ready adapters to qPCR products and qPCR assays performed with primers tailed with Illumina flow cell binding sites. To fully test the robustness of the techniques at concentrations required for gene doping detection, we also calculated a putative limit of detection for the assays. Both ligated adapters and tailed primers were successful in producing sequence data for the qPCR products without further amplification. Ligated adapters are preferred, however, as they do not require re-optimisation of existing qPCR assays.

REASONS FOR PERFORMING STUDY: Equine fatalities during racing continue to be a major welfare concern and falls at fences are responsible for a proportion of all equine fatalities recorded on racecourses. OBJECTIVES: To identify and quantify risk factors for horse falls in National Hunt (NH) racing and to report the frequency of falling and falling-associated fatalities. METHODS: A prospective cohort study was conducted on 2879 horse starts in hurdle and steeplechase races on 6 UK racecourses. Any horse that suffered a fall at a steeplechase or hurdle fence during the race was defined as a case. Data were obtained by interview and observations in the parade ring and from commercial databases. Multivariable logistic regression models, allowing for clustering at the level of the track, were used to identify the relationship between variables and the risk of falling. RESULTS: There were 124 falling cases (32 in hurdling and 92 in steeplechasing) identified. The injury risk of fallers was 8.9% and fatality risk 6.5%. Duration of journey to the racecourse, behaviour in the parade ring and weather at the time of the race were associated with falling in both hurdle and steeplechase racing. Age, amount of rainfall and going were also associated with falling in steeplechase racing. CONCLUSIONS: Falls at fences are significant contributors to equine fatalities during NH racing. Potentially modifiable risk factors identified were the condition of track surfaces and journey time to the racecourse. POTENTIAL RELEVANCE: It is hoped that information from this study may be used in future interventions to improve horse and jockey safety in racing. The study has also identified areas requiring further research, such as equine behaviour and its effect on racing performance, and the effect of light conditions on jumping ability.

OBJECTIVE: To determine whether there was an association between a history of cribbing and epiploic foramen entrapment (EFE) of the small intestine in horses. DESIGN: Retrospective study. ANIMALS: 68 horses examined at the University of Illinois or the University of Liverpool veterinary teaching hospitals. PROCEDURE: For horses examined at the University of Illinois that underwent surgery because of strangulating small intestine lesions, information about cribbing was obtained through telephone calls with owners. For horses examined at the University of Liverpool that underwent surgery for colic for any reason, information about cribbing was obtained through a preoperative questionnaire. RESULTS: 13 of 19 (68%) horses with EFE examined at the University of Illinois had a history of cribbing, compared with only 2 of 34 (6%) horses with other strangulating small intestine lesions (odds ratio, 34.7; 95% confidence interval, 6.2 to 194.6). Similarly, 24 of 49 (49%) horses with EFE examined at the University of Liverpool had a history of cribbing, compared with 72 of 687 (10.5%) horses with colic caused by other lesions (odds ratio, 8.2; 95% confidence interval, 4.5 to 15.1). CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that there may be an association between cribbing and EFE in horses, with horses with a history of cribbing more likely to have EFE than horses without such a history.

The intestinal microbiota of the horse, an animal of huge economic and social importance worldwide, is essential to the health of the animal. Understanding the intestinal ecosystem and its dynamic interaction with diet and dietary supplements currently requires the use of experimental animals, with consequent welfare and financial constraints. Here, we describe the development and assessment, using multiple analytical platforms, of a three-vessel, continuous-flow, in vitro model of the equine hindgut. After inoculation of the model with fresh horse feces, the bacterial communities established in each vessel had a taxonomic distribution similar to that of the source animal. Short-chain fatty acid (SCFA) and branched-chain fatty acid (BCFA) production within the model at steady state was consistent with the expected bacterial function, although higher concentrations of some SCFA/BCFA relative to those in the ex vivo gut content were apparent. We demonstrate the intermodel repeatability and the ability of the model to capture some aspects of individual variation in bacterial community profiles. The findings of this proof-of-concept study, including recognition of the limitions of the model, support its future development as a tool for investigating the impact of disease, nutrition, dietary supplementation, and medication on the equine intestinal microbiota.

The heteromeric sweet taste receptor T1R2-T1R3 is expressed on the luminal membrane of certain populations of enteroendocrine cells. Sensing of sugars and other sweet compounds by this receptor activates a pathway in enteroendocrine cells, resulting in secretion of a number of gut hormones, including glucagon-like peptide 2 (GLP-2). This subsequently leads to upregulation in the expression of intestinal Na(+)/glucose cotransporter, SGLT1, and increased intestinal glucose absorption. On the basis of the current information available on the horse genome sequence, it has been proposed that the gene for T1R2 (Tas1R2) is absent in the horse. We show here, however, that horses express both the mRNA and protein for T1R2. Equine T1R2 is most closely homologous to that in the pig and the cow. T1R2 protein, along with T1R3, α-gustducin, and GLP-2 proteins are coexpressed in equine intestinal endocrine cells. Intravenous administration of GLP-2, in rats and pigs, leads to an increase in the expression of SGLT1 in absorptive enterocytes and enhancement in blood glucose concentrations. GLP-2 receptor is expressed in enteric neurons, excluding the direct effect of GLP-2 on enterocytes. However, electric stimulation of enteric neurons generates a neural response leading to SGLT1 upregulation, suggesting that sugar in the intestine activates a reflex increase in the functional expression of SGLT1. Horses possess the ability to upregulate SGLT1 expression in response to increased dietary carbohydrates, and to enhance the capacity of the gut to absorb glucose. The gut sweet receptor provides an accessible target for manipulating the equine gut to absorb glucose (and water), allowing greater energy uptake and hydration for hard-working horses.

Anoplocephala perfoliata (Cestoda, Cyclophyllidea), the commonest intestinal tapeworm of horses, can cause colic, intussusceptions, ileal impactions and intestinal perforations. Common diagnostic techniques for A. perfoliata infection, i.e. coprology and serology, show inherent limitations in terms of sensitivity and specificity and new approaches are thus required. Hence, the present study compared the reliability of coprological, serological (i.e. ELISA) and molecular (i.e. nested PCR) methods in detecting A. perfoliata infection in naturally infected horses and in horses treated with a combination of ivermectin and praziquantel. Of 42 horses subjected to coprological examination, 16 and 26 resulted negative and positive, respectively for the presence of A. perfoliata eggs at the coprological examination. The 26 coprologically positive animals were also positive by nested PCR. Fifteen out of the 16 horses coprologically negative were negative at the molecular assay, while one yielded a PCR product detectable on an agarose gel. Eighteen out of 26 positive horses were treated with a combination of ivermectin 18.7 mg/g and praziquantel 140.3mg/g and resulted subsequently negative by coprology and nested PCR performed 2 weeks after treatment. All infected and untreated animals had a high ELISA test optical density indicating high infection intensity and associated risk of colic. However, high optical density values were also obtained in four horses post-treatment and in three horses that were negative on molecular and coprological analysis. The results of the present work indicate that the nested PCR assay represents a valid method for the specific molecular detection of A. perfoliata in faecal samples collected from naturally infected horses and may have advantages over coprological and serological approaches for diagnosing A. perfoliata infection.

BACKGROUND: Colic (abdominal pain) is a clinical condition of serious concern affecting the welfare and survival of donkeys at the Donkey Sanctuary in the UK. One of the most commonly reported causes is due to impacted ingesta in the large intestine ("impaction colic"). However little is known about the incidence of, or risk factors for, this condition. Here we describe the epidemiology of colic in donkeys, specifically impaction colic. We focus on temporal aspects of the disease and we identify environmental and management related risk factors for impaction colic in UK donkeys. RESULTS: There were 807 colic episodes in the population of 4596 donkeys between January 1st 2000 and March 31st 2005. The majority (54.8%) of episodes were due to a suspected or confirmed diagnosis of impaction of the gastrointestinal tract. The mortality risk for all colics (51.1%) was higher than reported in other equids. The incidence rate of all colics (5.9 episodes per 100 donkeys per year) and of impaction colic (3.2 episodes) was similar to that in horses. A retrospective matched case-control study of all impaction colics from January 2003 (193) indicated that older donkeys, those fed extra rations and those that previously suffered colic were at increased risk of impaction. Lighter body weight, musculo-skeletal problems, farm and dental disease were also significantly associated with a diagnosis of impaction colic. CONCLUSION: To our knowledge this is the first study to estimate the incidence rate of colic in a large population of donkeys in the UK. In contrast to other equids, impaction was the most commonly reported cause of colic. We identified several risk factors for impaction colic. Increasing age, extra rations and previous colic are known risk factors for colic in other equids. Results support the hypothesis that dental disease is associated with impaction colic. Musculo-skeletal problems may be associated with colic for various reasons including change in amount of exercise or time at pasture. Other associated factors (weight and farm) are the subject of further research. Identification of risk factors for impaction colic may highlight high risk donkeys and may allow intervention strategies to be introduced to reduce the incidence of the disease.

Two hundred and thirty-three horses were screened for the presence of roundworms by faecal egg counts (FECs) and for tapeworms by an ELISA specific for antibodies to the immunodominant 12 kDa and 13 kDa tapeworms antigen. The 62 horses were found to be infected with both parasites were treated with a combination of 0.2 mg/kg ivermectin and 1.5 mg/kg praziquantel. The treatment suppressed the median FEC of the horses to zero for 10 weeks and significantly reduced their anti-12/13 kDa antibody levels. The estimated risk of tapeworm-associated colic in these horses was halved by 12 weeks after the treatment.

The cross-country phase of eventing competitions has been associated with injuries and fatalities to horses and riders. A case-control study was carried out to identify variables that were associated with increased or decreased risk of a horse fall on the cross-country phase at event competitions. After initial analysis, the dataset was split according to the categories of one-day events as compared to two- or three-day events to establish whether significant risk factors varied between the different types of eventing competitions. Data were collected for 121 cases (horse falls) at one-day events, 59 cases at two- or three-day events and for their 540 matched controls. The data were analysed using conditional logistic regression. The variables of no previous refusals on the course, fences with a landing in water and the combined variable of the angle and the spread of the fence were significantly associated with the risk of a horse fall in both datasets. Additional risk factors for one-day event falls were: fences requiring a take-off from water, a drop landing, the rider's knowledge of their position before the cross-country phase and if the rider received cross-country tuition. Three-day event risk factors in the multivariable model included: the camber of the fence and participation in non-equestrian sports by the rider. This study identified variables that were significantly associated with an increase or a decrease in the risk of a horse fall during the cross-country phase of different types of eventing competitions. Some of these variables are modifiable and the results of this study have been reported to the governing body of the sport of eventing in the UK so that possible interventions might be considered.

The geographical spread of grass sickness between 1909 and 1999, particularly in England and Wales, is described, and the experimental investigations to identify a causal agent are summarised. The epidemiological techniques used to investigate grass sickness vary from clinical observations, to more advanced methods such as case-control studies using logistic regression analyses. Several risk factors for grass sickness have been reported consistently (age, time of year and recent movement to new pasture or premises) and several others have been reported for which the findings remain inconsistent (weather, pasture type, breed, supplementary feeding and use of anthelmintics).

Twenty-two horses with colic, referred to the University of Liverpool Equine Hospital (1992-1998), were found at laparotomy to have a partial obstruction of the large colon caused by a segmental mural lesion located in the left dorsal colon. The decision to take the horses to surgery was based on rectal examination findings of secondary impaction and mural oedema of the large colon and turbid peritoneal fluid containing large numbers of neutrophils obtained by paracentesis. The affected segment of colon showed oedema and serosal changes varying from erythema to well-defined necrosis. These pathological changes were considered to be sufficiently severe in 16 horses to merit partial resection of the left colon. Histology revealed varying degrees of mucosal necrosis, marked submucosal oedema and infiltration with large numbers of eosinophilic leucocytes. No cause was identified for the lesions. Sixteen of the 18 horses discharged from the hospital were alive 3 months to 7 years after surgery and have returned to their previous type of work and level of performance. The remaining 2 horses were subjected to euthanasia because of a recurrence of colic but no autopsy was performed.

© 2013 Bentham Science Publishers.SIFT-MS is used for the first time in profiling the volatile organic profile in faecal headspace in two groups of horses admitted to an equine hospital, one group with acute intestinal disease (colic) affecting the large colon, plus a control group of similarly managed horses admitted for non-gastrointestinal/metabolic reasons (e.g. acute orthopaedic injury). Compounds in faecal headspace which show statistically significant concentration differences between the groups are acetone and methanol. In addition, some ions at various m/z values show significantly different ion counts between the groups. Further information may be gleaned by using multivariate statistics in evaluating the differences between the two horse groups. Principal components analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were evaluated for reducing the dimensionality of the SIFT-MS data, and OPLS-DA was found to be best at discriminating between the groups, particularly with SIFT-MS data acquired using the H3O+ precursor ion. Analysis of these data also show the significance of ammonia as a discriminating ion. These results show that SIFT-MS may potentially be used on the headspace of horse faecal samples for detecting altered microbial fermentation associated with acute intestinal disease of the colon.

Reasons for performing study: Fatalities resulting from horse falls occurring during the cross-country phase of eventing competitions initiated epidemiological investigation of the risk factors associated with horse falls. Objectives: To identify variables that increased or decreased the risk of a horse fall during the cross-country phase of an eventing competition. Methods: Data were collected from randomly selected British Eventing competitions held in Great Britain during 2001 and 2002. Data were obtained for 173 cases (jumping efforts resulting in a fall of the horse-and-rider partnership) and 503 matched controls (jumping efforts not resulting in a fall). The risk of falling was modelled using conditional logistic regression. Results: An increased risk of a horse fall was associated with jumping into or out of water; taking off from good-to-soft, soft or heavy ground; fences with a drop landing; nonangled fences with a spread ≥2 m; and angled fences. Other risk factors included riders who knew that they were in the lead within the competition before the cross-country phase; an inappropriate speed of approach to the fence (too fast or too slow); horse-and-rider partnerships that had not incurred refusals at earlier fences; and riders who received cross-country tuition. Conclusions: This study has identified modifiable course- and fence-level risk factors for horse falls during the cross-country phase of eventing competitions. The risk of horse and rider injury at eventing competitions should be reduced by 3 simple measures; maintaining good to firm take-off surfaces at fences, reducing the base spread of fences to

Equine clinical larval cyathostominosis is caused by simultaneous mass emergence of previously inhibited larvae from the mucosa of the colon. Clinical signs include diarrhoea, colic, weight loss and malaise, and in up to 50% of cases, the disease results in death. Cyathostominae spend a large part of their life cycle as larval stages in the intestinal mucosa. Definitive diagnosis is difficult due to the lack of diagnostic methods for pre-patent infection. In the present study, the enzyme-linked immunosorbent assay (ELISA) was used to investigate isotype responses to larval cyathostominae somatic antigen. Measurement of anti-larval IgG(T) responses appeared to have the most immunodiagnostic potential. An increase in IgG(T) response was detected to crude larval antigen by 5 weeks post-infection (PI) in individual infected ponies. Subsequently, IgG(T) responses to larval and adult somatic extracts were examined by Western blotting using sera from experimentally-infected horses and helminth-naive animals (n=6). Two antigen complexes, designated A and B, in larval somatic antigen were recognised specifically by the infected animals by 7 weeks PI. Sera taken from 23 endemically-infected animals, whose cyathostominae burdens had been enumerated, were also used to identify putative diagnostic antigens. Eighteen horses had positive mucosal worm burdens (range 723-3,595,725) and all but two of these animals had serum IgG(T) antibody specific to either complex. Moreover, IgG(T) responses specific to antigen complexes A and B were absent in all five parasite negative horses that were tested. Serum IgG(T) responses to either of the two complexes were identified in five clinical cases tested. IgG(T) responses to adult antigen somatic extracts were more heterogeneous, with no clear pattern between experimentally-infected ponies and helminth-free controls. The results indicate that increases in serum IgG(T) to mucosal larvae occur in the pre-patent period and that two antigenic complexes within somatic preparations of these stages have immunodiagnostic potential.

The reason for undertaking this study was that postoperative complications of colic surgery lead to patient discomfort, prolonged hospitalisation and increased cost. Potential risk factors for the 6 most common postoperative complications (jugular thrombosis, ileus, re-laparotomy, wound suppuration, incisional herniation and colic) were evaluated using multivariable models. Jugular thrombosis was associated significantly with heart rate greater than 60 beats/min and with increased packed cell volume (PCV) at admission. The risk of postoperative ileus also increased with increasing PCV at admission and was higher in horses recovering from pedunculated lipoma obstruction. Incisional herniation was strongly associated with wound suppuration and with increasing heart rate at admission. The emergence of cardiovascular parameters as risk factors for several postoperative complications is consistent with the hypothesis that endotoxaemia is important in the development of these complications. Early referral of colic cases, prior to the development of severe endotoxaemic shock, may minimise the risk of some postoperative complications. Horses that have suffered epiploic foramen entrapment, are more than 4 times as likely to undergo re-laparotomy than other horses. Horses that have suffered postoperative ileus have a similarly increased risk of undergoing re-laparotomy. The risk of postoperative colic is significantly associated with horses recovering from large colon torsion (>360 degrees) and with having undergone re-laparotomy. Hazard ratios (with 95% confidence intervals) for these last two effects are 3.1 (1.7, 5.7) and 3.4 (1.9, 6.2), respectively. Knowledge of the risk factors for postoperative complications allows more accurate prognostication postoperatively and suggests ways in which the risk of postoperative complications can be minimised.

Background The horse plays crucial roles across the globe, including in horseracing, as a working and companion animal and as a food animal. The horse hindgut microbiome makes a key contribution in turning a high fibre diet into body mass and horsepower. However, despite its importance, the horse hindgut microbiome remains largely undefined. Here, we applied culture-independent shotgun metagenomics to thoroughbred equine faecal samples to deliver novel insights into this complex microbial community. Results We performed metagenomic sequencing on five equine faecal samples to construct 123 high- or medium-quality metagenome-assembled genomes from Bacteria and Archaea. In addition, we recovered nearly 200 bacteriophage genomes. We document surprising taxonomic diversity, encompassing dozens of novel or unnamed bacterial genera and species, to which we have assigned new Candidatus names. Many of these genera are conserved across a range of mammalian gut microbiomes. Conclusions Our metagenomic analyses provide new insights into the bacterial, archaeal and bacteriophage components of the horse gut microbiome. The resulting datasets provide a key resource for future high-resolution taxonomic and functional studies on the equine gut microbiome.

A case control study was performed to identify risk factors for colic caused by simple colonic obstruction and distension (SCOD) in the horse. Case horses were recruited from 2 veterinary school clinics. Control horses were population based and matched by time of year. A number of risk factors were considered in the following areas: general carer and premises information; exercise information; husbandry information (housing- and pasture-related); feeding information; breeding information; behavioural information; travel information; preventive medicine information and previous medical information. All variables with a P value of

Dietary carbohydrates, when digested and absorbed in the small intestine of the horse, provide a substantial fraction of metabolisable energy. However, if levels in diets exceed the capacity of the equine small intestine to digest and absorb them, they reach the hindgut, cause alterations in microbial populations and the metabolite products and predispose the horse to gastrointestinal diseases. We set out to determine, at the molecular level, the mechanisms, properties and the site of expression of carbohydrate digestive and absorptive functions of the equine small intestinal brush-border membrane. We have demonstrated that the disaccharidases sucrase, lactase and maltase are expressed diversely along the length of the intestine and D-glucose is transported across the equine intestinal brush-border membrane by a high affinity, low capacity, Na+/glucose cotransporter type 1 isoform (SGLT1). The highest rate of transport is in duodenum > jejunum > ileum. We have cloned and sequenced the cDNA encoding equine SGLT1 and alignment with SGLT1 of other species indicates 85-89% homology at the nucleotide and 84-87% identity at the amino acid levels. We have shown that there is a good correlation between levels of functional SGLT1 protein and SGLT1 mRNA abundance along the length of the small intestine. This indicates that the major site of glucose absorption in horses maintained on conventional grass-based diets is in the proximal intestine, and the expression of equine intestinal SGLT1 along the proximal to distal axis of the intestine is regulated at the level of mRNA abundance. The data presented in this paper are the first to provide information on the capacity of the equine intestine to digest and absorb soluble carbohydrates and has implications for a better feed management, pharmaceutical intervention and for dietary supplementation in horses following intestinal resection.

Equine grass sickness (EGS) is a frequently fatal disease of horses, responsible for the death of 1 to 2% of the U.K. horse population annually. The etiology of this disease is currently uncharacterized, although there is evidence it is associated with Clostridium botulinum neurotoxin in the gut. Prevention is currently not possible, and ileal biopsy diagnosis is invasive. The aim of this study was to characterize the fecal microbiota and biofluid metabolic profiles of EGS horses, to further understand the mechanisms underlying this disease, and to identify metabolic biomarkers to aid in diagnosis. Urine, plasma, and feces were collected from horses with EGS, matched controls, and hospital controls. Sequencing the16S rRNA gene of the fecal bacterial population of the study horses found a severe dysbiosis in EGS horses, with an increase in Bacteroidetes and a decrease in Firmicutes bacteria. Metabolic profiling by 1H nuclear magnetic resonance spectroscopy found EGS to be associated with the lower urinary excretion of hippurate and 4-cresyl sulfate and higher excretion of O-acetyl carnitine and trimethylamine-N-oxide. The predictive ability of the complete urinary metabolic signature and using the four discriminatory urinary metabolites to classify horses by disease status was assessed using a second (test) set of horses. The urinary metabolome and a combination of the four candidate biomarkers showed promise in aiding the identification of horses with EGS. Characterization of the metabolic shifts associated with EGS offers the potential of a noninvasive test to aid premortem diagnosis.

The aim of this study was to investigate cytokine expression patterns in the large intestinal mucosa of horses, particularly in diseases associated with inflammation. Many cases of equine colitis remain without a definitive diagnosis and survival rates are poor. In humans, colitis is associated with increased expression of pro-inflammatory cytokines. To examine if similar responses occur in horses, we investigated il -2, il -4, il -5, il -10, tnfalpha, ifngamma and tgfbeta messenger rna expression in large intestinal mucosa. Samples were obtained from animals with large intestinal disease (n=15) or from horses which had different levels of cyathostomin infection (n=9) and analysed by reverse transcription-polymerase chain reaction. il -2 was detected at all sites, whilst il -4 was detected at all but one site. The presence of il -10, il -5, ifngamma and tgfbeta varied with no significant differences amongst groups (P>0.4). Detection of tnfalpha was significantly different between the group of horses that had infiltrative inflammatory bowel disease and those with larval cyathostominosis (P=0.028) and those that were helminth negative (P=0.014). These results indicate a possible role for tnfalpha in the pathogenesis of equine infiltrative inflammatory bowel disease.

REASONS FOR PERFORMING STUDY: Lateral condylar fractures of the third metacarpus/metatarsus are the most common cause of equine fatality on UK racecourses. Identification of risk factors for such injuries and the subsequent implementation of intervention strategies could significantly reduce the total number of racecourse fatalities. OBJECTIVES: To identify horse-, race- and course-level risk factors for fatal lateral condylar fracture in Thoroughbreds on UK racecourses. METHODS: Case-control study designs were used. Case horses were defined as those that were subjected to euthanasia having sustained an irreparable lateral condylar fracture while racing at any of the 59 UK racecourses. Case races were defined as those in which one or more horses sustained a fatal lateral condylar fracture. Three controls for each case horse were selected at random from the race in which the case was running. Three controls for each case race were selected at random from all races of the same type held in the same year. Ninety-eight cases were included in the study. Conditional logistic regression was used to identify the relationship between a number of independent variables and the likelihood of fracture. RESULTS: Horses doing no gallop work during training and those in their first year of racing were at significantly increased risk of fracture on the racecourse. Case horses were also more likely to have started racing as 3- or 4-year-olds. Fractures were found to be more likely in longer races with a larger number of runners, races in which professional jockeys were not permitted to ride and races in which the going was described as firm or hard. CONCLUSIONS AND POTENTIAL RELEVANCE: Modifications to training schedules, specifically within the first year of racing, may have a large impact on the risk of fatal lateral condylar fracture on the racecourse. Horses should do some gallop work in training and our results suggest that the minimum distance galloped should be between 201 m (1 furlong) and 1609 m (8 furlongs) per week. The association with age at first race requires further investigation for flat and National Hunt racing separately. A reduction in the number of races taking place on very firm going could have an impact on the number of lateral condylar fractures.

REASONS FOR PERFORMING STUDY: Large intestinal diseases are an important cause of equine colic that require surgical management, but there is little published information about long-term survival of such cases. OBJECTIVES: To identify variables associated with post operative mortality and to develop a model for post operative survival following surgery for large intestinal disease. METHODS: Clinical data and long-term follow-up information from 275 horses undergoing surgery for large intestinal disease were used. A multivariable, Cox proportional hazards model for post operative death was developed and the fit of the model evaluated. RESULTS: The continuous variables age, heart rate and packed cell volume (PCV) were associated positively with the risk of post operative death, as were the categorical variables resection (odds ratio [OR] 2.4, 95% confidence interval [CI] 1.2-5.1) and relaparotomy (OR 3.5, 95% CI 1.4-8.4). CONCLUSIONS: In the population studied, the probability of post operative survival following surgery for large intestinal disease was associated with the cardiovascular parameters heart rate and PCV on admission, age of the horse and whether it underwent resection or relaparotomy. POTENTIAL RELEVANCE: The prognosis for post operative survival for horses with large intestinal disease may be improved by early surgery, prior to the development of cardiovascular compromise. Surgeons should be aware that increasing age, resection of large colon and relaparotomy are all associated with a worsening prognosis for survival.

REASONS FOR PERFORMING STUDY: Metabonomics is emerging as a powerful tool for disease screening and investigating mammalian metabolism. This study aims to create a metabolic framework by producing a preliminary reference guide for the normal equine metabolic milieu. OBJECTIVES: To metabolically profile plasma, urine and faecal water from healthy racehorses using high resolution (1) H-nuclear magnetic resonance (NMR) spectroscopy and to provide a list of dominant metabolites present in each biofluid for the benefit of future research in this area. STUDY DESIGN: This study was performed using 7 Thoroughbreds in race training at a single time point. Urine and faecal samples were collected noninvasively and plasma was obtained from samples taken for routine clinical chemistry purposes. METHODS: Biofluids were analysed using (1) H-NMR spectroscopy. Metabolite assignment was achieved via a range of one- and 2-dimensional experiments. RESULTS: A total of 102 metabolites were assigned across the 3 biological matrices. A core metabonome of 14 metabolites was ubiquitous across all biofluids. All biological matrices provided a unique window on different aspects of systematic metabolism. Urine was the most populated metabolite matrix with 65 identified metabolites, 39 of which were unique to this biological compartment. A number of these were related to gut microbial host cometabolism. Faecal samples were the most metabolically variable between animals; acetate was responsible for the majority (28%) of this variation. Short-chain fatty acids were the predominant features identified within this biofluid by (1) H-NMR spectroscopy. CONCLUSIONS: Metabonomics provides a platform for investigating complex and dynamic interactions between the host and its consortium of gut microbes and has the potential to uncover markers for health and disease in a variety of biofluids. Inherent variation in faecal extracts along with the relative abundance of microbial-mammalian metabolites in urine and invasive nature of plasma sampling, infers that urine is the most appropriate biofluid for the purposes of metabonomic analysis.

This study reports ivermectin and moxidectin egg reappearance periods (ERP) from UK horses with persistently positive faecal egg counts (FEC), defined as positive FEC within the ERP of an anthelmintic posttreatment, or with FECs that remained positive after the normal ERP post-anthelmintic treatment. A selected population of UK pleasure horses deemed at high risk of strongyle infection was studied. The earliest ERP recorded after ivermectin or moxidectin, using first positive FEC, was 5 weeks. From 16 premises where moxidectin was used, five had ERP ≥12 weeks using two further metrics. For premises where moxidectin was administered to only one animal (present or tested), and evaluated as one group (n = 61), ERP was ≥10 weeks. For premises where ivermectin was used, the ERP was ≥5 weeks. Premises with only one horse (present or tested), dosed with ivermectin (n = 31), analysed as one group, demonstrated egg reappearance ≥6 weeks. These field data suggest shortened ERPs following macrocyclic lactone treatment compared to previously published values (8–10 and >13 weeks respectively) when these drugs were first marketed

The ovicidal activity of benzimidazole (BZ) anthelmintics is unique and not seen in other drug classes. Such ovicidal efficacy is not widely reported for equine cyathostomins, nor has this activity been tested in the face of BZ resistance. Although the product label states that fenbendazole is for use against BZ-susceptible cyathostomins, susceptibility testing is rarely performed. In this field-based study, the ovicidal efficacy of fenbendazole in horses (n = 39) harbouring BZ-resistant cyathostomins was compared when dosed at 7.5 mg/kg body weight (BW) orally, as a single dose per os (n = 21) or daily for five consecutive days in feed (n = 18). Suppression of egg hatch rate was observed in the single and five- day treatment groups; a significant difference between pre- and post-treatment egg hatch rates (P < 0.05) was observed for three days after treatment with a single dose of fenbendazole (on premises with BZ-resistant cyathostomins), and for three days after treatment for five consecutive days with fenbendazole (on premises with BZ-resistant cyathostomins). Post treatment numbers of eggs and larvae remained significantly lower (P < 0.05) than pre-treatment levels to the end of the trial. We conclude that in the face of BZ-resistant cyathostomins the ovicidal effect of fenbendazole persist for three days after both a single oral dose of 7.5 mg/kg per os and after treatment orally for five consecutive daily doses at 7.5 mg/kg in feed.