Clinical Research Abstracts of the British Equine Veterinary Association Congress 2015.

REASONS FOR PERFORMING STUDY: Although fasciolosis is an important livestock disease worldwide, the public health importance of human fasciolosis has increased in recent years and it is recognised as an important re-emerging zoonotic disease, its epidemiology and pathogenicity in donkeys, and the epidemiological role they may play have not been determined. OBJECTIVES: To investigate the epidemiology and pathogenicity of fasciolosis in donkeys. STUDY DESIGN: Cross-sectional coprological and retrospective post-mortem study. METHODS: Faecal samples collected from 803 randomly selected working donkeys from the central region of Ethiopia were analysed by a sedimentation-centrifugation-flotation technique. Further data on liver-flukes and associated pathologies were obtained by routine post mortem examinations of 112 donkeys, subjected to euthanasia on welfare grounds or died. Data were analysed using a generalised linear model and multivariate binary logistic regression in R statistical package with significance level of statistical tests set at P<0.05. RESULTS: Infection prevalences of 44.4% and 41.9% were obtained in coprologically and post mortem examined donkeys, respectively, irrespective of their age. Both Fasciola hepatica and Fasciola gigantica were identified with the mean infection intensity of 30 flukes. Older donkeys (≥8 years) were found harbouring a significantly higher worm burden (P<0.0001). Gross and histopathologies of hyperplasia and thickening of the bile ducts, fibrosis of large portal areas and irregular bile duct proliferation and hypertrophy were noted. CONCLUSIONS: The high infection prevalence of fasciolosis and the associated hepatic pathologies in working donkeys shows not only the susceptibility of donkeys and the impact it has on their health, but also indicates the important role they can play in the epidemiology of both livestock and human fasciolosis. These further demonstrate the need for these animals to be considered in the overall epidemiological studies and for sound control strategies and prevention of fasciolosis. Ethical animal research: The research underwent ethical review and the use of animals was approved by the Directors of The Donkey Sanctuary. Consent of the owners was obtained to use their animals. SOURCE OF FUNDING: The Donkey Sanctuary. Competing interests: None declared.

A metapopulation model for highly pathogenic avian influenza: implications for compartmentalization as a control measure.

Although the compartmentalization of poultry industry components has substantial economic implications, and is therefore a concept with huge significance to poultry industries worldwide, the current requirements for compartment status are generic to all OIE member countries. We examined the consequences for potential outbreaks of highly pathogenic avian influenza in the British poultry industry using a metapopulation modelling framework. This framework was used to assess the effectiveness of compartmentalization relative to zoning control, utilizing empirical data to inform the structure of potential epidemiological contacts within the British poultry industry via network links and spatial proximity. Conditions were identified where, despite the efficient isolation of poultry compartments through the removal of network-mediated links, spatially mediated airborne spread enabled spillover of infection with nearby premises making compartmentalization a more 'risky' option than zoning control. However, when zoning control did not effectively inhibit long-distance network links, compartmentalization became a relatively more effective control measure than zoning. With better knowledge of likely distance ranges for airborne spread, our approach could help define an appropriate minimum inter-farm distance to provide more specific guidelines for compartmentalization in Great Britain.

Distinguishable epidemics of multidrug-resistant Salmonella Typhimurium DT104 in different hosts.

The global epidemic of multidrug-resistant Salmonella Typhimurium DT104 provides an important example, both in terms of the agent and its resistance, of a widely disseminated zoonotic pathogen. Here, with an unprecedented national collection of isolates collected contemporaneously from humans and animals and including a sample of internationally derived isolates, we have used whole-genome sequencing to dissect the phylogenetic associations of the bacterium and its antimicrobial resistance genes through the course of an epidemic. Contrary to current tenets supporting a single homogeneous epidemic, we demonstrate that the bacterium and its resistance genes were largely maintained within animal and human populations separately and that there was limited transmission, in either direction. We also show considerable variation in the resistance profiles, in contrast to the largely stable bacterial core genome, which emphasizes the critical importance of integrated genotypic data sets in understanding the ecology of bacterial zoonoses and antimicrobial resistance.

Identifying the seasonal origins of human campylobacteriosis.

Human campylobacteriosis exhibits a distinctive seasonality in temperate regions. This paper aims to identify the origins of this seasonality. Clinical isolates [typed by multi-locus sequence typing (MLST)] and epidemiological data were collected from Scotland. Young rural children were found to have an increased burden of disease in the late spring due to strains of non-chicken origin (e.g. ruminant and wild bird strains from environmental sources). In contrast the adult population had an extended summer peak associated with chicken strains. Travel abroad and UK mainland travel were associated with up to 17% and 18% of cases, respectively. International strains were associated with chicken, had a higher diversity than indigenous strains and a different spectrum of MLST types representative of these countries. Integrating empirical epidemiology and molecular subtyping can successfully elucidate the seasonal components of human campylobacteriosis. The findings will enable public health officials to focus strategies to reduce the disease burden.

Field efficacy of praziquantel oral paste against naturally acquired equine cestodes in Ethiopia.

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.

Quantifying the sources of variability in equine faecal egg counts: implications for improving the utility of the method.

The faecal egg count (FEC) is the most widely used means of quantifying the nematode burden of horses, and is frequently used in clinical practice to inform treatment and prevention. The statistical process underlying the FEC is complex, comprising a Poisson counting error process for each sample, compounded with an underlying continuous distribution of means between samples. Being able to quantify the sources of variability contributing to this distribution of means is a necessary step towards providing estimates of statistical power for future FEC and FECRT studies, and may help to improve the usefulness of the FEC technique by identifying and minimising unwanted sources of variability. Obtaining such estimates require a hierarchical statistical model coupled with repeated FEC observations from a single animal over a short period of time. Here, we use this approach to provide the first comparative estimate of multiple sources of within-horse FEC variability. The results demonstrate that a substantial proportion of the observed variation in FEC between horses occurs as a result of variation in FEC within an animal, with the major sources being aggregation of eggs within faeces and variation in egg concentration between faecal piles. The McMaster procedure itself is associated with a comparatively small coefficient of variation, and is therefore highly repeatable when a sufficiently large number of eggs are observed to reduce the error associated with the counting process. We conclude that the variation between samples taken from the same animal is substantial, but can be reduced through the use of larger homogenised faecal samples. Estimates are provided for the coefficient of variation (cv) associated with each within animal source of variability in observed FEC, allowing the usefulness of individual FEC to be quantified, and providing a basis for future FEC and FECRT studies.

Equine cestodosis: a sero-epidemiological study of Anoplocephala perfoliata infection in Ethiopia.

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.

Assessing the probability of acquisition of meticillin-resistant Staphylococcus aureus (MRSA) in a dog using a nested stochastic simulation model and logistic regression sensitivity analysis.

Meticillin-resistant Staphylococcus aureus (MRSA) is an important nosocomial and community-acquired pathogen with zoonotic potential. The relationship between MRSA in humans and companion animals is poorly understood. This study presents a quantitative exposure assessment, based on expert opinion and published data, in the form of a second order stochastic simulation model with accompanying logistic regression sensitivity analysis that aims to define the most important factors for MRSA acquisition in dogs. The simulation model was parameterised using expert opinion estimates, along with published and unpublished data. The outcome of the model was biologically plausible and found to be dominated by uncertainty over variability. The sensitivity analysis, in the form of four separate logistic regression models, found that both veterinary and non-veterinary routes of acquisition of MRSA are likely to be relevant for dogs. The effects of exposure to, and probability of, transmission of MRSA from the home environment were ranked as the most influential predictors in all sensitivity analyses, although it is unlikely that this environmental source of MRSA is independent of alternative sources of MRSA (human and/or animal). Exposure to and transmission from MRSA positive family members were also found to be influential for acquisition of MRSA in pet dogs, along with veterinary clinic attendance and, while exposure to and transmission from the veterinary clinic environment was also found to be influential, it was difficult to differentiate between the importance of independent sources of MRSA within the veterinary clinic. The implementation of logistic regression analyses directly to the input/output relationship within the simulation model presented in this paper represents the application of a variance based sensitivity analysis technique in the area of veterinary medicine and is a useful means of ranking the relative importance of input variables.

From phenotype to genotype: a Bayesian solution.

The study of biological systems commonly depends on inferring the state of a 'hidden' variable, such as an underlying genotype, from that of an 'observed' variable, such as an expressed phenotype. However, this cannot be achieved using traditional quantitative methods when more than one genetic mechanism exists for a single observable phenotype. Using a novel latent class Bayesian model, it is possible to infer the prevalence of different genetic elements in a population given a sample of phenotypes. As an exemplar, data comprising phenotypic resistance to six antimicrobials obtained from passive surveillance of Salmonella Typhimurium DT104 are analysed to infer the prevalence of individual resistance genes, as well as the prevalence of a genomic island known as SGI1 and its variants. Three competing models are fitted to the data and distinguished between using posterior predictive p-values to assess their ability to predict the observed number of unique phenotypes. The results suggest that several SGI1 variants circulate in a few fixed forms through the population from which our data were derived. The methods presented could be applied to other types of phenotypic data, and represent a useful and generic mechanism of inferring the genetic population structure of organisms.

Associations between the presence of virulence determinants and the epidemiology and ecology of zoonotic Escherichia coli.

The severity of human infection with pathogenic Escherichia coli depends on two major virulence determinants (eae and stx) that, respectively, produce intimin and Shiga toxin. In cattle, both may enhance colonization, but whether this increases fitness by enhancing cattle-to-cattle transmission in the field is unknown. In E. coli O157, the almost uniform presence of the virulence determinants in cattle isolates prevents comparative analysis. The availability to this study of extensive non-O157 E. coli data, with much greater diversity in carriage of virulence determinants, provides the opportunity to gain insight into their potential impact on transmission. Dynamic models were used to simulate expected prevalence distributions for serogroups O26 and O103. Transmission parameters were estimated by fitting model outputs to prevalence data from Scottish cattle using a Bayesian Markov chain Monte Carlo (MCMC) approach. Despite similar prevalence distributions for O26 and O103, their transmission dynamics were distinct. Serogroup O26 strains appear well adapted to the cattle host. The dynamics are characterized by a basic reproduction ratio (R(0)) of >1 (allowing sustained cattle-to-cattle transmission), a relatively low transmission rate from environmental reservoirs, and substantial association with eae on transmission. The presence of stx(2) was associated with reduced transmission. In contrast, serogroup O103 appears better adapted to the noncattle environment, characterized by an R(0) value of <1 for plausible test sensitivities, a significantly higher transmission rate from noncattle sources than serogroup O26, and an absence of fitness benefits associated with the carriage of eae. Thus, the association of eae with enhanced transmission depends on the E. coli serogroup. Our results suggest that the capacity of E. coli strains to derive fitness benefits from virulence determinants influences the prevalence in the cattle population and the ecology and epidemiology of the host organism.

Putative household outbreaks of campylobacteriosis typically comprise single MLST genotypes.

During a 15-month period in Scotland a small but important number of human Campylobacter cases (3·2%) arose from 91 putative household outbreaks. Of the 26 outbreaks with known strain composition, 89% were composed of the same MLST which supports the potential use of MLST in public health epidemiology. The number of cases associated with household outbreaks is much larger than general outbreaks and there is some evidence to indicate that there may be secondary transmission, although this is relatively rare.

Risk of carcase contamination with Campylobacter in sheep sent for slaughter into an abattoir in Scotland.

Campylobacter species have been identified as the major cause of acute bacterial enteritis in the UK. However, the epidemiology of campylobacteriosis remains poorly understood. It has been suggested that the role of sheep in the epidemiology of Campylobacter has been underestimated. The objective of the present study was to assess the infection risk of Campylobacter in sheep meat as a potential risk for human campylobacteriosis and to establish any possible associations between the epidemiological factors considered in the study and the presence of Campylobacter on lamb carcases. The prevalence of Campylobacter obtained from faecal samples and swabs from fleeces and carcases was presented in a previous paper. Epidemiological data was collected through questionnaires in order to identify risk factors for the presence of Campylobacter on the carcases and to make recommendations, based on the results obtained, to prevent human campylobacteriosis.

Epidemiological features of fasciolosis in working donkeys in Ethiopia.

A cross-sectional coprological survey in the tropical regions of Ada, Akaki, Bereh and Boset, and a retrospective post-mortem investigation were conducted to study the epidemiology of fasciolosis in working donkeys in Ethiopia. Faecal samples from 803 donkeys were collected, and the number of liver flukes recovered from 112 donkeys at post-mortem between 1995 and 2004 were analysed. There was a high prevalence of fasciolosis irrespective of the age of the donkeys. The overall prevalence of the infection was 44.4% in coprologically examined donkeys, and the prevalence in the donkeys examined post-mortem was 41.9%. The infection prevalence was significantly higher in Bereh and Ada regions than in Akaki and Boset regions. Bereh with 72.6% and Boset with 21.5% showed a significantly higher and lower infection prevalence, respectively, than the rest of the regions (P<0.001). There was no significant difference between different age groups of donkeys in the infection prevalence (P>0.05) but infection intensity was significantly higher in donkeys 8 years old and above (P<0.0001). Both Fasciola hepatica and Fasciola gigantica were identified.

Comparison of three alternative methods for analysis of equine Faecal Egg Count Reduction Test data.

The Faecal Egg Count Reduction Test (FECRT) is the most widely used method of assessing the efficacy of anthelmintics, and is the only in vivo technique currently approved for use with horses. Equine Faecal Egg Count (FEC) data are frequently characterised by a low mean, high variability, small sample size and frequent zero count observations. Accurate analysis of the data therefore depends on the use of an appropriate statistical technique. Analyses of simulated FECRT data by methods based on calculation of the empirical mean and variance, non-parametric bootstrapping, and Markov chain Monte Carlo (MCMC) are compared. The MCMC method consistently outperformed the other methods, independently of the distribution from which the data were generated. Bootstrapping produced notional 95% confidence intervals containing the true parameter as little as 40% of the time with sample sizes of less than 50. Analysis of equine FECRT data yielded inconclusive results in 53 of 63 (84%) datasets, suggesting that the routine use of prior sample size calculations should be adopted to ensure sufficient data are collected. The authors conclude that computationally intensive parametric methods such as MCMC be used for analysis of FECRT data with sample sizes of less than 50, in order to avoid erroneous inference about the true efficacy of anthelmintics in the field.

Gastrointestinal parasites of working donkeys of Ethiopia.

The general prevalence and population composition of gastrointestinal and pulmonary helminths of working donkeys were studied. For the purpose 2935 working donkeys were coprologically examined for nematode and cestode, and 215 donkeys for trematode infections. Seven donkeys that died due to various health problems or were euthanased on a welfare ground were necropsied and the parasites were recovered and identified to the species level. The study was conducted during the periods 1996-1999.Coprological examination revealed 99% strongyle, 80% Fasciola, 51% Parascaris, 30% Gastrodiscus, 11% Strongyloides westeri, 8% cestodes and 2% Oxyuris equi infection prevalence. Over 55% of donkeys had more than 1000 eggs per gram of faeces (epg). Forty two different species of parasites consisting of 33 nematodes, 3 trematodes, 3 cestodes and 3 arthropod larvae were identified from postmortem examined donkeys. Among the nematodes 17 species of Cyathostominae and 7 species of Strongylinae were identified. Other parasites identified include, Habronema muscae, Draschia megastoma, Trichostrongylus axei, Strongyloides westeri, Anoplocephala perfoliata, Anoplocephala magna, Anoplocephaloides (Paranoplocephala) mamillana, Parascaris equorum, Fasciola hepatica, Fasciola gigantica, Gastrodiscus aegyptiacus, Dictyocaulus arnfieldi, Oxyuris equi, Probstmayria vivipara, Gasterophilus intestinalis, Gasterophilus nasalis, Rhinoestrus uzbekistanicus and Setaria equina. This study revealed that working donkeys in Ethiopia are infected with a range of helminths and arthropod larvae, which are representatives of the important pathogenic parasites found in equids worldwide.

Prevalence and distribution of meticillin-resistant Staphylococcus aureus within the environment and staff of a university veterinary clinic.

OBJECTIVES: To characterise the distribution of meticillin-resistant Staphylococcus aureus within the environment of a university small animal hospital and compare this with the distribution among staff. METHODS: Samples were collected from 140 environmental sites and the anterior nares of 64 staff members at the University of Glasgow Small Animal Hospital on a single day (d1). Sixty of the environmental sites were resampled 14 days later (d14). RESULTS: Meticillin-resistant S aureus was isolated from two of 140 (1.4 per cent; 95 per cent confidence interval: 1.7 to 5.1) environmental sites on d1 and one of 60 (1.7 per cent; 95 per cent confidence interval: 0.4 to 8.9) on d14. Two of the 64 staff sampled were positive for meticillin-resistant S aureus (3.1 per cent; 95 per cent confidence interval: 0.4 to 8.4). CLINICAL SIGNIFICANCE: A lower prevalence of meticillin-resistant S aureus was observed in the environment than previously reported. The location, relatedness between isolates and the presence of Panton-Valentine leucocidin indicates that the source of the environmental meticillin-resistant S aureus was most likely to have been human rather than animal in these cases. This study presents important information regarding the potential source and distribution of meticillin-resistant S aureus within veterinary hospital environments and highlights potential variability of prevalence of meticillin-resistant S aureus within and between veterinary institutions.

Exploiting strain diversity to expose transmission heterogeneities and predict the impact of targeting supershedding.

When a few individuals generate disproportionately many secondary cases, targeted interventions can theoretically lead to highly efficient control of the spread of infection. Practical exploitation of heterogeneous transmission requires the sources of variability to be quantified, yet it is unusual to have empirical data of sufficient resolution to distinguish their effects. Here, we exploit extensive data on pathogen shedding densities and the distribution of cases, collected from the same population within the same spatio-temporal window, to expose the comparative epidemiology of independent Escherichia coli O157 strains. For this zoonotic pathogen, which exhibits high-density shedding (supershedding) and heterogeneous transmission in its cattle reservoir, whether targeting supershedding could be an effective control depends critically on the proposed link between shedding density and transmissibility. We substantiate this link by showing that our supershedder strain has nearly triple the R(0) of our non-supershedder strain. We show that observed transmission heterogeneities are strongly driven by superspreading in addition to supershedding, but that for the supershedder strain, the dominant strain in our study population, there remains sufficient heterogeneity in contribution to R(0) from different shedding densities to allow exploitation for control. However, in the presence of substantial within-host variability, our results indicate that rather than seek out supershedders themselves, the most effective controls would directly target the phenomenon of pathogen supershedding with the aim of interrupting or preventing high shedding densities. In this system, multiple sources of heterogeneity have masked the role of shedding densities-our potential targets for control. This analysis demonstrates the critical importance of disentangling the effects of multiple sources of heterogeneity when designing targeted interventions.

Incidence rate of musculoskeletal injuries and determinants of time to recovery in young Australian Thoroughbred racehorses.

OBJECTIVE: To describe the incidence rate of musculoskeletal (MS) injuries and investigate the factors that influence recovery from MS injury in young Thoroughbred horses in Australia. METHODS: Training and injury data were collected at fortnightly intervals from 14 trainers in New South Wales. Incidence rates were calculated for initial injuries stratified by age-class and sex. Survival analysis methods were used to investigate the time to recovery after the first MS injury (measured as a start in a race or barrier trial). RESULTS: For the 248 racehorses enrolled, there were 428 injuries recorded; 2-year-olds were 2.99-fold more likely than 3-year-olds to sustain an MS injury. The cumulative percentage of horses that had recovered within 6 months was 55%. The time to recovery was longest in horses whose fast gallop speed prior to injury was < 890 m/min. CONCLUSION: This study supports the finding that shin soreness is the most common MS injury in young Australian Thoroughbreds. There was an increased rate of MS injury in 2-year-olds compared with 3-year-olds. However, it was impossible to determine if the increased risk in 2-year-olds was related to age, the start of training or a combination of both. The study also found that horses performing at higher exercise speeds prior to injury were more likely to recover. There was a significant effect of trainer, suggesting that other factors related to trainer may influence the time to the recovery. Additional research is required to understand the determinants of time to recovery.