Development and Evaluation of a Molecular Diagnostic Method for Rapid Detection of Histoplasma capsulatum var. farciminosum, the Causative Agent of Epizootic Lymphangitis, in Equine Clinical Samples.

Histoplasma capsulatum var. farciminosum, the causative agent of epizootic lymphangitis (EZL), is endemic in parts of Africa. Diagnosis based on clinical signs and microscopy lacks specificity and is a barrier to further understanding this neglected disease. Here, a nested PCR method targeting the internal transcribed spacer (ITS) region of the rRNA operon was validated for application to equine clinical samples. Twenty-nine horses with signs of EZL from different climatic regions of Ethiopia were clinically examined. Blood samples and aspirates of pus from cutaneous nodules were taken, along with blood from a further 20 horses with no cutaneous EZL lesions. Among the 29 horses with suspected cases of EZL, H. capsulatum var. farciminosum was confirmed by extraction of DNA from pus and blood samples from 25 and 17 horses, respectively. Positive PCR results were also obtained with heat-inactivated pus (24 horses) and blood (23 horses) spotted onto Whatman FTA cards. Two positive results were obtained among blood samples from 20 horses that did not exhibit clinical signs of EZL. These are the first reports of the direct detection of H. capsulatum var. farciminosum in equine blood and at high frequency among horses exhibiting cutaneous lesions. The nested PCR outperformed conventional microscopic diagnosis, as characteristic yeast cells could be observed only in 14 pus samples. The presence of H. capsulatum var. farciminosum DNA was confirmed by sequencing the cloned PCR products, and while alignment of the ITS amplicons showed very little sequence variation, there was preliminary single nucleotide polymorphism-based evidence for the existence of two subgroups of H. capsulatum var. farciminosum This molecular diagnostic method now permits investigation of the epidemiology of EZL.

Colic in a working horse population in Egypt: Prevalence and risk factors.

REASONS FOR PERFORMING STUDY: Colic is an important health problem in managed horse populations. Currently, there is limited information about colic prevalence and risk factors for colic in working horse populations. OBJECTIVES: To determine the prevalence of, and risk factors for, colic in a working horse population in Egypt and to describe management practices in this working horse population. STUDY DESIGN: Cross-sectional survey. METHODS: Owners of 350 working horses were interviewed. Data about their horses, management and colic episodes in the preceding 12 months were collected. Dental examination was conducted on 342 horses and blood samples (n = 100) were collected for immunodiagnosis of tapeworm (Anoplocephala perfoliata) infection. Multivariable logistic regression analysis was used to identify risk factors for a history of colic in the preceding 12 months. RESULTS: The 12-month prevalence of colic was 54.6%. Severe and moderate tapeworm infection intensity was identified in 3% and 26% of horses tested, respectively. Horses that had severe dental disease (odds ratio [OR] 6.8, 95% confidence interval [CI] 1.9-24.3, P<0.001), that displayed stereotypic behaviour (OR 2.0, 95% CI 1.15-3.5, P = 0.013), were fed ground corn during the 'dry season' (OR 1.65, 95% CI 1.03-2.6, P = 0.035) or that had received an anthelmintic in the previous 6 months (OR 2.1, 95% CI 1.3-3.3, P = 0.003) were more likely to have a history of colic in the preceding 12 months. Horses fed on rice bran during the 'green season' (OR 0.47, 95% CI 0.26-0.9, P = 0.015) and that displayed geophagia/coprophagia (OR 0.19, 95% CI 0.05-0.73, P = 0.001) were less likely to have a history of colic. CONCLUSIONS: Colic is common in this working horse population and this study has identified factors associated with altered likelihood of colic. The study provides important information that may be used to inform future prospective studies investigating colic in working horse populations and to assist development of preventive healthcare strategies. The Summary is available in Chinese - see Supporting information.

Management and horse-level risk factors for recurrent colic in the UK general equine practice population.

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.

Participatory appraisal of the impact of epizootic lymphangitis in Ethiopia.

Epizootic lymphangitis (EZL) is reported to have a significant impact upon livelihoods within resource-poor settings. This study used a participatory approach to explore peoples' experiences of EZL and examine the perceived impact of disease, owner knowledge and understanding of EZL, lay management of disease and, attitudes and strategies towards disease prevention. Focus-group discussions were held with 358 cart-horse owners and drivers recruited from 7 towns attended by SPANA (Society for the protection of animals abroad) mobile veterinary clinics and 2 unexposed towns where no SPANA clinics were available. Focus group discussions explored four main research questions: (1) Is EZL recognised by animal owners, and is this considered an important disease in equids? (2) What factors do animal owners associate with the development of disease? (3) What happens to an animal with clinical disease and how does this impact upon the owner/community? (4) Are measures taken to reduce disease occurrence? These key areas were explored using photographs, disease ranking, matrices and open discussion. Data were analysed using descriptive statistics and thematic analysis. The results are presented thematically and include: recognition and descriptions of EZL, treatment strategies used, disease priorities and ranking, impact of disease, disease transmission and attitudes and approaches to disease prevention. EZL was widely recognised and ranked highly as an important disease of equids. However, there was uncertainty around identifying early cases of EZL, and this could impact upon the timing of initiating treatment and separating potentially infectious animals. People had varying knowledge of effective methods for disease prevention and reported particular difficulties with isolating infected animals. The impact of EZL was multi-dimensional and encompassed effects upon the horse, the individual owner and the wider society. Working equids provide a vital utility and source of income to many people in resource-poor settings. Often, infection with EZL resulted in a reduction in working ability which had a direct impact upon the livelihoods of owners and their dependent family members. EZL also impacted upon the welfare of the horse as sick animals continued to be worked and, in advanced cases, horses were abandoned due to ineffective or unavailable treatment. This study conceptualises the importance of EZL due to the effects of the disease on the horse and its impact upon human livelihoods. Epizootic lymphangitis is a neglected disease that requires further investigation in order to develop practical and sustainable disease control strategies within endemic regions.

An evidence-based approach to the evaluation of ethnoveterinary medicines against strongyle nematodes of equids.

Cyathostomins are the most important gastrointestinal nematode infecting equids. Their effective control is currently under threat due to widespread resistance to the broad spectrum anthelmintics licenced for use in equids. In response to similar resistance issues in other helminths, there has been increasing interest in alternative control strategies, such as bioactive plant compounds derived from traditional ethnoveterinary treatments. This study used an evidence-based approach to evaluate the potential use of plant extracts from the UK and Ethiopia to treat cyathostomins. Plants were shortlisted based on findings from a literature review and additionally, in Ethiopia, the results of a participatory rural appraisal (PRA) in the Oromia region of the country. Systematic selection criteria were applied to both groups to identify five Ethiopian and four UK plants for in vitro screening. These included Acacia nilotica (L.) Delile, Cucumis prophetarum L., Rumex abyssinicus Jacq., Vernonia amygdalina Delile. and Withania somnifera (L.) Dunal from Ethiopia and Allium sativum L. (garlic), Artemisia absinthium L., Chenopodium album L. and Zingiber officinale Roscoe. (ginger) from the UK. Plant material was collected, dried and milled prior to hydro-alcoholic extraction. Crude extracts were dissolved in distilled water (dH2O) and dimethyl sulfoxide (DMSO), serially diluted and screened for anthelmintic activity in the larval migration inhibition test (LMIT) and the egg hatch test (EHT). Repeated measures ANOVA was used to identify extracts that had a significant effect on larval migration and/or egg hatch, compared to non-treated controls. The median effective concentration (EC-50) for each extract was calculated using PROBIT analysis. Of the Ethiopian extracts A. nilotica, R. abyssinicus and C. prophetarum showed significant anthelmintic activity. Their lowest EC-50 values were 0.18 (confidence interval (CI): 0.1-0.3), 1.1 (CI 0.2-2.2) and 1.1 (CI 0.9-1.4)mg/ml, respectively. All four UK extracts, A. sativum, C. album, Z. officinale and A. absinthium, showed significant anthelmintic activity. Their lowest EC-50 values were 1.1 (CI 0.9-1.3), 2.3 (CI 1.9-2.7) and 0.3 (CI 0.2-0.4)mg/ml, respectively. Extract of A. absinthium had a relatively low efficacy and the data did not accurately fit a PROBIT model for the dose response relationship, thus an EC-50 value was not calculated. Differences in efficacy for each extract were noted, dependent on the assay and solvent used, highlighting the need for a systematic approach to the evaluation of bioactive plant compounds. This study has identified bioactive plant extracts from the UK and Ethiopia which have potential as anthelmintic forages or feed supplements in equids.

Ocular disease in working horses in Ethiopia: a cross-sectional study.

Ocular disease is a frequent finding in working horses. This study aimed to estimate the prevalence and types of ocular pathology, and explore risk factors potentially associated with disease within a population of working horses in Ethiopia. In total, 1049 horses were selected from horses attending clinics run by the Society for Protection of Animals Abroad (SPANA). Each had an ophthalmic examination conducted under field conditions using a pen-torch. All owners completed a short questionnaire. The prevalence of ocular abnormalities was 23.5 per cent (95 per cent confidence interval (CI) 18.0 per cent to 30.1 per cent) and the percentage of horses with an abnormality in at least one eye was 43 per cent (95 per cent CI 28.7 per cent to 58.4 per cent), although this varied between towns. Mild eye pathology and end-stage disease with irreversible pathology were most common. There were significantly more eye abnormalities in the right eye compared with the left, and older horses were more likely to have ocular pathology. Only 55.1 per cent of owners had noticed there was an ocular abnormality present, and only 2.2 per cent had received any previous treatment for eye disease. Only 1.9 per cent presented to the clinic because of an eye problem. There appears to be either a lack of owner awareness, or a low perception of the importance of eye disease among owners.

Recurrent colic in the horse: incidence and risk factors for recurrence in the general practice population.

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.