Horses - RIRDC Completed Projects in 2003--2004 & Research in Progress as at June 2005

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RIRDC Completed Projects in 2005-2006 & Research in Progress as at June 2006
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2.4 HORSES - COMPLETED PROJECTS

PROJECT No PROJECT TITLE RESEARCHER PHONE ORGANISATION

Information Technology for the Horse Industry

UM-66A A prospective study of pre sale radiographs of thoroughbred yearlings Prof Andrew Clarke (03) 9731 2314 University of Melbourne

US-124A Communications and information programs for the horse industry Ms Shirley Ray (02) 9352 6932 University of Sydney

To improve the management of growing and developing horses

US-129A Risk factors for injuries in thoroughbred racehorses during commercial training

Prof David Evans (02) 9351 2474 University of Sydney

To improve the overall health and welfare of the horse

OFF-1A The production of a poster and booklet on poisonous weeds for horses

Ms Mellisa Offord (02) 8901 0622

UQ-114A Penetration of pharmacological agents through equine skin

Dr Paul Mills (07) 3365 2964 University of Queensland

To more effectively quantify the environmental impact on the horse industry

UQ-119A Transfer of weeds to Qld native forests on horses hooves

Dr. Christopher Pollitt (07) 3365 2063 The University of Queensland

Respiratory Diseases-Improving the Respiratory Health of Horses

UM-67A Effects of Stud Management on Ecology of Virulent Rhodococcus equi and Further studies of the ecology of virulent R. equi on horse studs - 2002 stud season

Dr Glenn Browing (03) 8344 742 University of Melbourne

Lameness and Limb Injury-Developing More Effective Methods for Prevention, Diagnosis and Treatment

UM-58A Epidemiology and risk factor analysis of racetrack fatalities Prof. Andrew Clarke (03) 9731 2314 The University of Melbourne

UQ-100A Pasture fructan concentration as a cause of equine laminitis Dr. Christopher Pollitt (07) 3365 2063 The University of Queensland

Other

EPS-1A Prototype Environmental Management System (EMS) for the horse keeping community - implementation, auditing and monitoring Ms Jacqueline Frizenschaf (08) 8139 9919 Environment Protection Authority SA

Completed Projects – Information Technology for the Horse Industry

Project Title A prospective study of pre sale radiographs of thoroughbred yearlings

RIRDC Project No.: UM-66A

Researcher: Prof Andrew Clarke

Organisation: The University of Melbourne

Phone: (03) 9731 2314

Fax: (03) 9731 2377

Email: afclarke@unimelb.edu.au

Objectives ·1 To identify the incidence of radiographic lesions in yearling thoroughbred horses presented for sale in Australia.
·2 To provide an objective assessment of the impact these findings have on the racing careers (e.g. number of starts, earning capacity, age of first start) of the horses involved.

Background Radiograph repositories were introduced to auction sales in Australia in 2003 for pre-purchase assessment of orthopaedic lesions in Thoroughbred yearlings. There were 3,905 yearlings entered into 8 major national yearling sales in this year. Of these 2,847 had radiographs submitted to the radiograph repository. 2,412 of the 2,847 radiograph sets (81,741 individual x-rays) were made available for the present study. The radiographs were reviewed one of four veterinary specialist radiologists. In addition 167 sets were randomly selected and each radiologist read these sets twice. Sale catalogues and results were provided by the sales companies for all eight sales. The Australian Stud Book was used to verify catalogue information and ascertain if yearlings were named for racing or exported overseas. The i-RIS database was used to retrieve the race records of yearlings at two and three years of age.
Melissa Jackson is the PhD student working with this project. This project and the results generated by Melissa will be the basis of her thesis. Contact details: Melissa Jackson, Equine Centre, University of Melbourne, 250 Princes Highway, Werribee Vic 3030. Ph: 03 9731 2316, Fax: 03 9731 2246, Email: m.jackson2@pgrad.unimelb.edu.au

Research This project has five major components: 1. Comparison between radiographed and non-radiographed yearlings relating to sale and two-year-old race performance.
2. Prevalence of radiographically detectable orthopaedic lesions in radiographed yearlings.
3. Association between lesion presence and race performance at two and three years of age.
4. Race profile at two and three years of age and an economic analysis of return on money invested in the purchase of the yearling.
5. An assessment of the quality of radiographs and repeatability of assessments of radiograph quality and lesions by specialist radiologists.

Outcomes This project will be completed in January 2007. Preliminary findings to date include:
1. Overall 3.5% (2871/81741) of films were considered to be non-diagnostic. In approximately half of the radiographic sets examined the aim of a complete assessment was not achieved due to non-diagnostic or missing films. Care in positioning of FlexLM views of the fore fetlocks, LM views of the hind fetlocks and DMPaLO views of the carpus is especially important for maximising radiographic quality.
2. As yearling sale price increases, the likelihood of starting a race and earning prize money to cover the cost of training fees increases. The probability of earning more than the purchase price of the yearling decreases with increasing price category.

Implications The components of the study involving the determination of the prevalence of radiographically detected orthopaedic lesions in yearlings, and the analysis of the association of such lesions with future performance are the largest studies of their kind undertaken. This is the first study to document the prevalence of radiographically detectable orthopaedic lesions in Thoroughbred yearlings in Australia and the first Australian study to estimate the effect, if any, of these lesions on race performance.
The career profile and economic analysis component of the project is the largest and most detailed of its kind ever undertaken in Australia.
This project is providing information to the industry that will allow both owners and veterinarians to make more informed decisions relating to the significance of radiographic findings in yearlings.

Completed Projects – Information Technology for the Horse Industry

Project Title Communications and information programs for the horse industry

RIRDC Project No.: US-124A

Researcher: Ms Shirley Ray

Organisation: The University of Sydney

Phone: (02) 9463 9305

Fax: (02) 9463 9100

Email: shirleyr@camden.usyd.edu.au

Objectives To continue to ensure widespread industry support for the RIRDC Equine R&D Program and to enable the results of the R&D to be used by the industry, a strong communication program is necessary to ensure results get to end users. ·3 A database of equine organisations.
·4 A database for communication with the industry.
·5 RIRDC Equine Research News. This newsletter of 8 pages, published quarterly, will provide the industry with information on topical research from around the world as well as updates on current RIRDC funded projects, interviews with research workers, workshops and invited reviews.
·6 RIRDC Internet Web Site. An Internet web site for the RIRDC Horse Program which will include links for online purchase of RIRDC publications, download of the RIRDC Equine Research News, search facility for past issues and a subscription facility to receive the RIRDC Equine Research News via the Internet.

Research Each newsletter published had a print run of approximately 9,500 copies, all of which were distributed to the various sectors of the horse industry. The newsletter contained summaries of articles of local and international interest, interviews with researchers in particular fields, announcements of new RIRDC funded projects and updates of all currently funded projects, and items of general interest to the horse industry including RIRDC events and new publications. Articles from the newsletter have been reprinted in many local and international publications including: The Horse (USA), Sport Horse Journal, Breeding and Racing, Race Calendars and Hoofbeats.
The RIRDC Equine Research Program Web site contains articles that have been printed in the RIRDC Equine Research News; these articles are accessible through an index of subject areas. A local search engine allows the user to search the site for a keyword. The site has links to relevant pages of the RIRDC web site, including the Five Year Plan, Objectives and Strategies of the Equine Research Program and RIRDC’s e-shop. A links page includes links to local and international research institutions and other sites of interests. At least 50 websites contain a link to the RIRDC Equine Program website and these include: Cyberhorse, Australian Racing Board; Australian Horse Resources and Australian Horse Net Service. All past issues of the newsletter are available for download in PDF format which can be viewed using Adobe Acrobat. This means that anyone with Internet access can view the newsletter in its colour print format.
The mailing list for the RIRDC Equine Research Program is updated regularly and contains contact details of Australian Horse Associations, Research Institutions and Personnel, Horse Publications and other media and individuals with an interest in the horse industry. The mailing list contains over 2,000 entries

Completed Projects - To improve the management of growing and developing horses

Project Title Risk factors for injuries in thoroughbred racehorses during commercial training

RIRDC Project No.: US-129A

Researcher: Associate Professor David Evans

Organisation: The University of Sydney

Phone: (02) 9351 2474

Fax: (02) 9351 3957

Email: davide@vetsci.usyd.edu.au

Objectives ·7 The aim of this study was to investigate the epidemiology of MS injuries in Australian two- and three-year-old Thoroughbred racehorses. More specifically the aims were to describe the incidence and impact of MS and determine risk factors for MS injuries, in particular shin soreness.

Background Musculoskeletal (MS) injuries have been identified as a common health problem in Thoroughbred racehorses. The most common MS injuries in two- and three-year olds have been identified as shin soreness and problems involving the carpus and fetlock joints. These MS injuries incur both direct and indirect costs.
After reviewing the literature it is clear that there is very limited understanding of the risk factors for MS injuries. In particular, there is almost a complete absence of any investigations that have collected data on daily training in an effort to understand the training related risk factors for MS injuries. Studies that utilize daily training records are fundamental to the understanding of training related risk factors for MS injuries. An understanding of the risk factors for MS injuries would facilitate the design of strategies that may reduce the occurrence and impact of MS injuries.

Research During the study period 451 horses were enrolled during their two-year-old racing season. Training and injury information for these horses were collected directly from the trainer. The research confirmed previous research that MS injuries are a common problem in two- and three-year-old Thoroughbred racehorses. The results showed that injury prior to the first start had considerable impact on time to first start. However, preparations after the first MS injury do not appear to be adversely affected. The investigation of risk factors showed that increasing distance galloped at high speed increased the risk of MS injury.

Outcomes Since 2000 the results of this study have been presented to a number of difference associations and audiences. These were: ·1 Australian Equine Veterinary Association
·2 Society of Veterinary Epidemiology and Preventive Medicine
·3 International Society Veterinary Epidemiology and Economics
·4 Institute of Veterinary and Biomedical Science, Massey University
·5 Australian and New Zealand racetrack managers
·6 Sydney metropolitan and provincial Thoroughbred trainers
·7 Australian Veterinary Association
·8 British Equine Veterinary Association
·9 International Horse Industry Symposium

Implications The results of this study show that exposure to high-speed exercise is associated with an increased risk of injury. Unfortunately, racehorses must be exposed to high-speed exercise in order to prepare them for competition. Therefore, future research should focus on developing ways in which the risk associated with high-speed exercise can be reduced.

Publications Cogger N., Perkins H., Hodgson D.R., Reid S.W.J. and Evans, D.L. (in press). "Risk factors for musculoskeletal injuries in two-year-old Thoroughbred racehorses." Preventive Veterinary Medicine.

Completed Projects - To improve the overall health and welfare of the horse

Project Title The production of a poster and booklet on poisonous weeds for horses

RIRDC Project No.: OFF-1A

Researcher: Ms Mellisa Offord

Organisation: Mellisa Offord

Phone: (02) 8901 0622

Fax: (02) 9451 3286

Email: mofford@optusnet.com.au

Objectives ·8 To produce a poster and booklet showing plants and trees that are poisonous to horses.

Outcomes A book and poster have been produced and these were launched at the Equine Science Symposium held on the Gold Coast in June 2006.
The book is titled Plants Poisonous to Horses – An Australian Field Guide. It is 127 pages and it is a guide for horse owners to help them prevent plant poisoning in their horses. It clearly outlines the factors that influence the risk of plant poisoning and the ways to reduce this risk. The types of toxins that can be found in plants are discussed, along with the signs of poisoning that may be seen in horses that have ingested toxic plant material. A description of each plant is provided, including structure, growth habit and identifying features. The book contains over 200 colour photos of poisonous plants. Management notes suggest ways of dealing with poisonous plants on properties. The book includes a comprehensive list of ornamental plants that are known to be toxic to horses. The reference list is a useful resource for further reading and includes the journal articles, industry papers, books, CD ROMs and Internet resources that were used to create this publication.
The poster highlights 11 plants in Australia that are a risk to horses.

Publications Offord M (2006). Plants Poisonous to Horses: An Australian Field Guide. RIRDC Publication No. 06/048

Completed Projects - To improve the overall health and welfare of the horse

Project Title Penetration of pharmacological agents through equine skin

RIRDC Project No.: UQ-114A

Researcher: Dr Paul Mills

Organisation: The University of Queensland

Phone: (07) 3365 2964

Fax: (07) 3365 1255

Email: p.mills@uq.edu.au

Objectives ·1 To develop in vitro and in vivo methodologies to measure drug movement through equine skin
·2 To measure the regional differences (e.g. neck vs back vs axilla vs lower leg) in drug penetration
·3 To measure transdermal penetration of drugs of interest to the equine industry, including non-steroidal anti-inflammatory drugs (NSAIDs), corticosteroids and analgesics (e.g. Fentanyl)
·4 To determine the effects of vehicles (e.g. DMSO) on penetration of drugs in the horse
·5 To determine the effects of skin damage (e.g. rashes) and altered microenvironment (e.g. bandages, saddles) on transdermal drug movement in the horse

Background The skin is a protective layer preserving the integrity of the internal environment. The major barrier is the outermost layer, the stratum corneum. An increasing number of pharmaceutical agents are being developed for topical use in humans and animals. Advantages of transdermal drug penetration include; (i) ease of administration; (ii) increased compliance with use; (iii) avoidance of hepatic "first-pass" effect, and (iv) reduced gastric irritation.
It is now mandatory for any topical preparation intended for human use to undergo extensive laboratory testing prior to registration. Recent adverse developments in veterinary topical medications (e.g. Bayticol in cattle and Frontline (fipronil) in dogs and cats) have shown that inadequate studies were undertaken prior to registration. The Australian Pesticides and Veterinary Medicines Authority (APVMA) have recently taken an active interest in prerequisites for registration of veterinary topical medications.
Little is known about transdermal movement of drugs through equine skin. Many of the currently available products, such as corticosteroid creams and NSAID gels, are based on results from human or laboratory animal studies, but it is widely acknowledged that extrapolation of transdermal drug movement between species is unreliable. This may lead to adverse or unexpected results when these preparations are applied to horses. Indeed, an increasing number of positive swab samples collected during equine competition have been blamed on unexpected penetration through skin.
In this proposed study, we will use scientifically credible in vitro and in vivo methods to investigate transdermal drug movement through equine skin. Drugs that may be applied topically and are of specific interest to the equine industry will be examines, including NSAIDs, corticosteroids and other analgesics (e.g. procaine and fentanyl). We will also investigate potential factors affecting transdermal drug movement, particularly region of application (e.g. neck vs groin), the effects of altered skin integrity (rashes or bandaging) and the effects of vehicles (e.g. DMSO, propylene glycol).
The results from this study will be of benefit to all stakeholders in the equine industry. Information about where best to apply specific drugs to maximise transdermal passage, or regions to avoid if systemic concentrations are undesirable would be useful to veterinarians, owners and trainers. There in no information about drug movement through damaged skin and if medications are applied under bandages, yet many drugs are applied under these circumstances. The potential of inadvertent swab sample positives needs to the addressed by the industry and this research is a pro-active step towards developing understanding and expertise. It may also become mandatory for laboratory testing of drugs intended for topical application and having expertise and infrastructure to undertake this task would be extremely useful.

Research Preliminary information will be determined using in vitro methodologies. Equine skin will be obtained from euthanased animals and placed in Franz-type diffusion cells. Pharmacological agents, in a variety of vehicles, can be applied to the stratum corneum as a donor phase and analytes passing into and through the skin can be measured. The diffusion cell is maintained at approximately 32oC, the expected temperature of the skin surface, using a water bath. The receptor phase can be varied to mimic conditions expected in the dermis and underlying tissues of the horse. Specific measurement of drug and metabolites in skin and passing through to the receptor phase with use either high performance liquid chromatography (HPLC) or scintillation counting (if radiolabelled drug is used).
Two in vivo models will be used to confirm in vitro results and account for biological differences in living skin, such as changes in blood flow. Whole body studies where drug and/or vehicle are applied to the horse and analytes measured in blood and urine. A new technique, microdialysis, will also be developed in the horse. This involves placing semi-permeable probes into the epidermis and dermis, then directly measuring drug appearance in perfusate. Once preliminary data is obtained, models of skin damage (abrasion and/or delipidation) will be developed using harvested skin. This will permit direct testing, in vitro, of the effects of skin damage on transdermal drug movement. It is also hoped to develop and apply mathematical principles and available models to predict transdermal drug movement of new drugs or to assess the effects of alternative vehicles or the effects of skin damage.

Outcomes The techniques that may be used to investigate transdermal drug delivery in the horse were verified, particularly for in vitro methods which can be used to screen drugs and specifically compare the effects of regional differences, vehicles and skin integrity.
Skin was harvested from the thorax, groin and leg (dorsal metacarpal) of horses to investigate regional variation in the penetration of a number of commonly used drugs in horses. Significantly higher (P<0.001) maximum flux (Jmax; mol/cm2/hr) was measured when hydrocortisone was applied to skin from the leg, compared to thorax and groin, although significantly less hydrocortisone (P<0.001) was retained within skin from the leg at 24 hrs. Topical application of hydrocortisone in a vehicle containing ethanol would penetrate faster through leg skin from the lower leg, compared to the thorax or groin which, depending on cutaneous blood flow, may result in higher systemic drug concentrations or greater efficiency in treating local inflamed tissue. Similarly, a commercial formulation of methylsalicylate (Dencorub®) will achieve therapeutic levels of total salicylate beneath sites of topical application, with a faster and more pronounced response achieved through the leg region, compared to upper body. However, fentanyl Transdermal Therapeutic Systems (TTS or ‘patch’) applied to the leg region may have a small but significantly lower amount of fentanyl available systemically, compared to patches applied to the thorax or groin regions, which may affect the level of analgesia subsequently achieved in the horse.
The effects of the vehicles phosphate buffered saline (PBS), ethanol (EtOH; 50% in PBS w/w) and propylene glycol (PG; 50% in PBS w/w) on in vitro transdermal penetration of testosterone was investigated in the horse. The maximum flux (Jmax) of testosterone was significantly higher for all sites when dissolved in a vehicle containing 50% ethanol or 50% propylene glycol, compared to PBS. In contrast, higher residues of testosterone were found remaining within the skin when PBS was used as a vehicle. This study shows that variability in clinical response to testosterone could be expected with formulation design.
To investigate the effects of common skin preparations on transdermal drug penetration through equine skin in vitro, skin was exposed to the following treatments: Control (no treatment); Cleaned with aqueous chlorhexidine; cleaned with alcoholic chlorhexidine; Shaved; Tape stripped using elastoplast adhesive tape. A significantly higher proportion of methylsalicylate penetrated through alcoholic chlorhexidine and Shaved (P<0.01) and aqueous chlorhexidine and Tape stripped (P<0.05) treated skin, compared to Control, while significantly more Sa was recovered in the receptor phase of Tape stripped (P<0.01), alcoholic chlorhexidine, aqueous chlorhexidine and Shaved (P<0.05) treated skin, compared to Control. A significantly higher rate of penetration and shorter lag time was also noted for methylsalicylate following each skin pre-treatment, compared to control. The results showed that routine clinical techniques used to clean or prepare skin can significantly affect the rate and extent of penetration of a topically applied drug. This may result is more active drug available systemically enhancing efficacy and, possibly, the incidence of adverse effects to the particular drug.

Implications There are no guidelines for the topical administration of drugs to horses prior to or during competition which this project has partially addressed. These guidelines and precautions for stakeholders in the horse industry will be disseminated and can be seen as a pro-active response to inadvertent positive swab samples. We now have a centre of expertise in the area of transdermal drug penetration in the horse which has already provided expert opinion to stewards in Australia and Ireland. Importantly, we now also have the expertise to investigate transdermal penetration of other drugs relevant to the equine industry.
A peer-reviewed scientific review currently in press will provide guidelines and information for veterinarians so that they can understand transdermal drug penetration and advise their clients (owners, trainers, stewards) accordingly. The information generated about the effects of skin pre-treatment will have important implications for application of any topical medications to equine skin. For example, cleaning skin, shaving skin to remove hair or applying drugs after the removal of adhesive bandages will all affect the rate and amount of drug penetration.
This research has established scientifically-credible testing methodologies which can be used to test potential new topical remedies and to advise registration bodies (especially the APVMA) about equine topical medications. This is important as, similar to human topical medications; there are increasing requirements before topical medications can be regarded as safe and efficacious.

Publications Mills P.C. and Cross S.E. (2006) The effects of skin preparation on transdermal drug penetration in vitro through equine skin. Can J Vet Res (in press).
Mills P.C. and Cross S.E. (2006) Transdermal Drug Delivery: Basic Principles for the Veterinarian. Vet J (in press- doi 10.1016/j.tvjl.2005.09.006).
Mills P.C. and Cross S.E. (2006) Regional differences in transdermal penetration of fentanyl through equine skin. Res Vet Sci. (in press).
Mills P.C. (2006) Vehicle effects on the penetration of testosterone through equine skin. Vet.Res.Comm. (in press).
Mills P.C. and Cross S.E. (2006) Regional differences in the in vitro penetration of hydrocortisone through equine skin. J Vet.Pharm.Ther. 29: 25-30.
Mills P.C. and Cross S.E. (2006) Regional differences in the in vitro penetration of methylsalicylate through equine skin. Vet.J. (in press doi 10.1016/j.tvjl.2005.08.023).
There have also been a number of lay articles published in Australia (The Veterinarian), Ireland (Racing Times) and USA (Equus).

Completed Projects – To more effectively quantify the environmental impact on the horse industry

Project Title Transfer of weeds to Qld native forests on horses hooves

RIRDC Project No.: UQ-119A

Researcher: Dr Christopher Pollitt

Organisation: The University of Queensland

Phone: (07) 3365 2063

Fax: (07) 3365 1899

Email: c.pollitt@uq.edu.au

Objectives ·6 Do horse faeces contain viable seeds which may germinate under suitable conditions or persist in surface soil.
·7 Are the plants that germinate from horse manure found beside the trails used by horse riders in forests of SE Queensland.

Background Weeds accidentally introduced since European settlement constitute a threat to the ecological sustainability of native plant communities. Weeds may be introduced into native ecosystems via vehicular, human and animal vectors and there is anecdotal evidence that horse manure contains viable weed seed. A rigorous scientific protocol is necessary to discover if seeds in horse manure are viable and pose a threat when horses traverse areas of conservation significance.

Research Horse faeces were collected from horses grazing an 80 hectare pasture without recent weed control and from race horses maintained in stables. Half of the faeces were dried and half kept wet before being mixed with a commercial, seed free, potting mix.

Outcomes Ten plant species germinated in the pasture derived faeces (compared to one from stabled horses) and there was a higher seed load in the pasture faeces. Dominating seed germination was the grass Queensland Blue Couch (Digitaria ciliaris). Germination was 5 times greater in the dried faeces suggesting that prior drying favours grass seed germination. Only one native species, emu foot (Psoralea tenax) was found. All species that germinated in manure have current weed status.

Implications The faeces of horses grazing weed infested pastures contain viable seed that, potentially, may infest native forests. However further work is required to determine if this actually occurs. To determine the role of horse faeces as a source of weeds, plants growing on 100 m transects of the sampled pasture and transects of the trails in nearby native forests will be compared. When complete these studies will supply objective data to assess the level of threat posed by horse riding in areas of native vegetation and will help formulate rational management strategies for recreational horse riding.

Publications In press

Completed Projects – Respiratory Diseases – Improving the Respiratory Health of Horses

Project Title Effects of Stud Management on Ecology of Virulent Rhodococcus equi and Further studies of the ecology of virulent R. equi on horse studs - 2002 stud season

RIRDC Project No.: UM-47A and UM-67A

Researcher: Glenn F Browning, James R Gilkerson and Gary Muscatello

Organisation: School of Veterinary Science
The University of Melbourne

Phone: 03 8344 7342

Fax: 03 8344 7374

Email: glenfb@unimelb.edu.au

Objectives ·8 To develop methods to improve understanding of the factors that make some areas on horse farms a greater risk for development of rattles in foals and apply these to identifying factors that might be addressed by changes in management of farms.

Background Although rattles is an important disease problem in young foals there are limited options for control. Current methods rely on reducing faecal contamination and dust on studs, but we have little understanding of whether these are the only factors that account for higher infection and death rates on some studs.

Research This project aimed to develop novel methods for assessing the risk associated with particular horse farm environments. These methods were based on bacteriological sampling of air and soil on studs, then identifying those bacteria capable of causing rattles. Once these methods were developed they were applied to examination of different horse farms and different places on those farms.
The method that was developed was also used to examine the breath of foals with rattles. This suggested that foals may be able to transmit virulent R. equi to each other.

Outcomes The most dangerous areas on studs for foals are likely to be laneways and holding pens. Control may be aided by reducing the time that foals spend in these areas. In addition areas that had sandy, dry and/or acidic soils seemed to be a greater risk. Some control may also be achieved by avoiding close contact between foals, particularly if they are known to be suffering from rattles. Sampling air from foals may be a sensitive diagnostic technique for the early detection of infected foals.

Implications Application of methods developed in this project have provided some suggestions for reducing the incidence of rattles and further use of these methods may enable the benefit obtained from different approaches to control to be assessed. Different approaches to treatment may also be able to be assessed.

Publications Muscatello, G. Rhodococcus equi: the organism and epidemiology. Australian Equine Veterinarian 22:190-197 (2003).
Muscatello, G., and Browning, G. F. - Identification and differentiation of avirulent and virulent Rhodococcus equi using selective media and colony blotting DNA hybridization to determine their concentrations in the environment. Veterinary Microbiology, 100:121-127 (2004).
Muscatello, G., Gerbaud, S., Kennedy, C., Gilkerson, J. R., Buckley, T., Klay, M., Leadon, D. P., and Browning, G. F. – Comparisons between concentrations of Rhodococcus equi and virulent R. equi in the air of stables and paddocks on horse breeding farms in a temperate (European) climate. Equine Veterinary Journal, 38:263-265 (2006).
Phumoonna, T., Muscatello, G., Chicken, C., Gilkerson, J. R., Browning, G. F., Barton, M. D., And Heuzenroeder, M. W. - Clinical evaluation of a peptide-ELISA based upon N-terminal B cell epitope of the Vap A protein for early diagnosis of Rhodococcus equi pneumonia in foals. Journal of Veterinary Medicine B, 53:126-32 (2006).
Muscatello, G., Gilkerson, J. R., and Browning, G. F. - Comparison of two selective media for the recovery, isolation, enumeration and differentiation of Rhodococcus equi. Submitted.
Muscatello, G., Anderson, G. A., Gilkerson, J. R., and Browning, G. F. - Associations between the ecology of virulent Rhodococcus equi and the epidemiology of R. equi pneumonia on Australian Thoroughbred farms, Applied and Environmental Microbiology, in press (2006).
Muscatello, G., Gilkerson, J. R., And Browning, G. F. - Detection of virulent Rhodococcus Equi in exhaled air samples from foals. Submitted.

Completed Projects – Lameness and limb injury – Developing more effective methods of prevention, diagnosis and treatment

Project Title Epidemiology and risk factor analysis of racetrack fatalities

RIRDC Project No.: UM-58A

Researcher: Prof Andrew Clarke

Organisation: The University of Melbourne

Phone: (03) 9731 2314

Fax: (03) 9731 2377

Email: afclarke@unimelb.edu.au

Objectives ·9 This project will deliver an improved understanding of the causes of and risk factors associated with racetrack fatalities. This will translate into strategies to address this complex multifactorial problem. This in turn will result in reduced wastage within the industry and have positive welfare implications.

Background This project was the first in Australia to combine a prospective post mortem case series and large-scale retrospective observational studies to investigate the causes of and risks associated with racehorse fatality in flat and jump starts. The study was encouraged by the Victorian racing industry in response to industry and community concerns regarding high wastage rates of racehorses as well as jockey injuries and fatalities in racing. The purpose of this study was to identify risk factors that could be modified to reduce the risk of fatality in Thoroughbred racing in Victoria.
Dr Lisa Boden was the PhD student working with this project. This project and the results generated by Lisa form the basis of Lisa’s thesis which has been successfully defended. Contact details: "Lisa Boden" lboden@staffmail.ed.ac.uk

Research The following studies have been achieved as a result of this project:
A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria.
B. Risk of fatality and causes of death of Thoroughbred horses associated with racing in Victoria, Australia between 1989 and 2004.
C. Risk factors for Thoroughbred racehorse fatality in flat races in Victoria between 1989 and 2004.
D. Risk factors for Thoroughbred racehorse fatality in jump races in Victoria between 1989 and 2004.
E. Estimated attributable fractions for racehorse fatalities in Victoria.

Outcomes The results of the post mortem study showed that catastrophic limb injury was the major cause of fatality and that sudden death is a more important contributor to racing fatalities than previously reported in Victoria.
The risk of fatality in flat starts was lower in Victoria (0.44 fatalities per 1000 starts) than that reported in North America (1.4-1.7 fatalities per 1000 starts and the UK (0.8-0.9 fatalities per 1000 starts) but the risk of fatality in jump starts was greater in Victoria (8.3 fatalities per 1000 starts) than that reported overseas (3.9 fatalities per 1000 starts) in North America and 5.6 fatalities per 1000 starts in the UK.
The risk of fatality in flat starts appeared to increase over the 15 year study period but it remains unclear whether this was due to a genuine increase in risk (resulting from changes in such factors as climate or racing practices over time) or to increased stringency of reporting of proportions of fatalities.
The risk of fatality in jump starts remained constant over the study period despite jump racing reviews that recommended changes to hurdle and steeple races to improve safety.
The risk of fatality on city racecourses was 2.0 times that on country racecourses.
Risk factors associated with increased odds of fatality in flat starts were horse gender (stallions and geldings compared with mares), one or more starts in the 31-60 day period prior to the case or control start, the going of the track (fast or good surfaces), racecourse location (city compared with country tracks), increasing race length and race year .
Risk factors for jump starts included the number of flat and jump starts accumulated in the racing career prior to the study start, one or more starts in the 1-14 days prior to the case or control start, race type (steeplechase compared with hurdle races), track location (city compared with country tracks) and calendar year. Of the risk factors identified as having high population attributable fraction estimates, the most readily modifiable were: track condition in flat races (fast or good tracks), race type in jump races (steeplechase events) and the number of prior flat starts in racing career for horses in jump races.
Of the risk factors identified as having high population attributable fraction estimates, the most readily modifiable were: track condition in flat races (fast or good tracks), race type in jump races (steeplechase events) and the number of prior flat starts in racing career for horses in jump races.
Jump starts by horses with shorter racing careers were more likely to result in fatality. Over the course of a racing career, horses that started in more flat starts prior to the case or control jump start were more likely to die in the next jump start than horses that had accumulated less flat racing experience. Additionally, increased numbers of jump starts prior to the study start were also associated with increased odds of fatality.
The odds of fatality in jump starts increased with each additional flat start accumulated in the racing career. The average number of career flat starts for horses starting in flat races was 17.2 whereas horses starting in jump races had already accumulated an average of 33 career flat starts. If horses were allowed to start in up to 32 flat starts (and no more) before beginning a jump racing career, the risk of fatality in jump starts may decrease by 50% (approximately eight fatalities per year).

Implications This project identified specific risk factors for fatality in flat and jump racing in Victoria. The results of this study should be utilised to direct further research into starts, horses and races at high risk so that appropriate intervention strategies can be implemented to improve the welfare of horses racing in Victoria.
Specific recommendations made to the industry as a result of this project include:
•1 Continuation and expansion of post mortem program to include fatalities from country tracks and the collection of blood from cases of sudden death by racetrack veterinarians.
•2 Formalised monitoring of injuries and fatalities
•3 Standardisation of track rating methods and review of current policy regarding racing on ‘good’ tracks
•4 Further investigation of differences between risk in hurdles and steeplechase races
•5 Restriction of number of flat starts horses can accumulate before starting a jump racing career

Publications A very successful international two day workshop on the prevention of racetrack fatalities and injuries was hosted in Melbourne in 2005. This was the first ever international conference addressing this subject. A copy of the CD of the proceedings of the workshop has been produced. The CD includes all of the PowerPoint presentations from the symposium and has proven to be popular for those unable to attend the symposium. Other international meetings are being planned as a result of this inaugural meeting.
This project was the basis of a PhD for Dr Lisa Boden. The thesis has been completed and successfully defended. The following publications have arisen from this project to date:
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey, J., Clarke, A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. In: Proceedings of the AVA Annual Conference, Canberra, May 2004.
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey, J., Clarke, A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. Australian Equine Veterinarian Vol. 23 2004, p 148. (Abstract from the Bain Fallon Annual Conference of Australian Equine Practitioners, Darwin, July 2004).
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey, J., Clarke, A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. In: Proceedings of the Annual Conference of the Australian Society of Veterinary Pathologists, Brisbane, October 2004.
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey*, J., Clarke, A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. Proceedings of the annual conference of the Australian Veterinary Association, Canberra, May 2004
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey*, J., Clarke, A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. Australian Equine Veterinarian (23), 148.
Boden, L., Charles, J., Slocombe, R., Sandy, J., Finnin, P., McCaffrey, J.,Morton J., Clarke, A.. Sudden death in racing Thoroughbreds in Victoria, Australia. Equine Veterinary Journal (2005) 37 (3) 269.
Boden,LA. J.A. Charles, K.L. Morgan, J. Morton, T.D.H. Parkin, R.F. Slocombe, J.R. Sandy, A. F. Clarke (2005) Risk factors for racehorse fatalities in flat races in Victoria, Australia. Proceedings of Annual Conference of British Equine Veterinary Association
Boden, LA, J.A. Charles, K.L. Morgan, J.M. Morton, T.D.H. Parkin, J.R. Sandy, R.F. Slocombe, A. F. Clarke (2005) Incidence of fatalities and specific causes of death of racehorses on racecourses in Victoria, Australia between 1989 and 2004 Proceedings of Annual Conference of British Equine Veterinary Association
Boden, LA, J.A. Charles, K.L. Morgan, J.M. Morton, T.D.H. Parkin, J.R. Sandy, R.F. Slocombe, A.F. Clarke (2005) Risk factors for Thoroughbred racehorse fatalities in flat races in Victoria, Australia (1989-2004). Proceedings of Australian Equine Association
Boden, LA, Anderson, GA, Charles, JA, Finnin, PJ, Morgan, KL, Morton, JM, Parkin, TDH, Sandy, JR, Slocombe, RF, Clarke, AF. (2005) Incidence of fatalities of Thoroughbred racehorses in Victoria, Australia between 1989 and 2004. Proceedings of Australian Equine Association
Boden, LA, Charles,JA, Slocombe, RF, Sandy JR, Finnin PJ, McCaffrey, JA*, Morton, JM, Clarke, AF. (2005) Post mortem study of Thoroughbred fatalities in Victoria, Australia between 2001 and 2004. Proceedings of the American Association of Equine Practitioners.

Completed Projects – Lameness and limb injury – Developing more effective methods of prevention, diagnosis and treatment

Project Title Pasture fructan concentration as a cause of equine laminitis

RIRDC Project No.: UQ-100A

Researcher: Dr Christopher Pollitt

Organisation: The University of Queensland

Phone: (07) 3365 2063

Fax: (07) 3365 1899

Email: c.pollitt@uq.edu.au

Objectives ·1 The cause of grass founder will be understood and a strategy for diagnosis, prevention and treatment will be available
·2 A strategy for pasture and grazing management will be available so that horses and ponies do not consume sufficient fructan to develop laminitis.

Research 1. Establish that dosing horses with Raftilose causes laminitis
Horses dosed with Raftilose or oligofructose (OF) at 3 dose rates (7.5;10 & 12.5g/kg) developed clinical and histological laminitis in at least one foot. Higher doses of OF were associated with significantly more severe (grade 3) histological laminitis (p<0.05). All the horses developed profuse diarrhoea at around 18h that ceased by 36-44h. Pyrexia and elevated heart rate peaked between 16 and 20h and returned to near normal by 48h. Faecal pH began to fall within 4 hours of dosing, reaching its lowest value (3.7) at around 18h. Blood D-lactate, of bacterial origin, peaked at 24 hours and then returned to the normal zero. Since all horses developed laminitis, the results establish that a storage carbohydrate, other than starch, can induce the disease. OF is a principal non-structural carbohydrate of pasture grasses, suggesting it is grass OF that causes laminitis and not other substance(s), such as dietary amines. Compared with traditional alimentary overload with starch, OF dosing resulted in reduced morbidity and no deaths, indicating that alimentary overload with fructan is an efficient and more humane experimental model for the induction of acute laminitis.
A paper entitled "Equine laminitis: experimental induction with oligofructose by C.C. Pollitt* and A. W. van Eps was submitted to the Equine Veterinary Journal in mid 2003. The referees were not satisfied that 3 pairs of horses dosed with oligofructose (OF) at 3 dose rates could indicate significant trends (despite our professional statistician’s explanation of the methods used). Publication has been deferred until 4 more horses have been dosed with the median dose (10/kg Bwt OF).
2. Establish surgical caecal cannulation
The difficulties that were encountered with caecal cannulation in 2002 have been overcome by using an alternative 2-step method. In stage 1 an adhesion was created between the caecum and the skin after the muscle in between was surgically removed. In stage 2, 14 days later, access to the caecal lumen was established by trocarisation of the adhesion and a soft, rubber caecal cannula was introduced. One horse has been cannulated (HGF1) and laminitis was induced successfully giving us insight into events in the caecum for the first time. Two horses have had stage 1 surgery and are ready for cannula insertion (stage 2). More cannulations and laminitis inductions are planned for 2005.
3. Establish profile of normal caecal microbiota by culture
Profiles of normal caecal microflora populations were documented through the creation of novel habitat-simulating medium containing Raftilose as the sole energy substrate (incubated equine caecal-fluid agar - IECA). Using this media, raftilose utilising organism (RUO) numbers in relation to other anaerobic bacteria were established for normal caecal contents.
4. Purify LTFs by filtration
The trigger factors responsible for initiating laminitis remain elusive. It is clear however that the factors responsible for triggering the onset of laminitis occur in the equine hindgut 8-16h post fructan administration. Furthermore, the dominance of S. bovis, during this period implicates it as the aetiological agent of equine laminitis. Attempts to purify laminitis trigger factors (LTFs) from large volumes of S bovis culture media, using centrifugal filtration equipment, were not successful. We have developed a new casein containing medium and confirmed by zymography that S bovis produces a protease in the presence of casein. Analysis by zymography in gelatin containing PAGE gels showed the S bovis protease (SBP) was a gelatinase and possibly a metalloproteinase. Since the activated MMPs that cause laminitis are also gelatinases we investigated the possibility that SBP was a LTF. SBP was not inhibited by batimastat thus diminishing its likely resemblance to lamellar MMP. Interestingly caecal liquor taken when S bovis proliferation during OF induced laminitis had peaked contained very high concentrations of SBP. SBP and clarified caecal liquor from horses developing laminitis did not induce lamellar separation in vitro suggesting that LTFs are not of bacterial origin. It seems more likely that S bovis proliferation and concomitant protease production damages the mucosal epithelium of the hindgut and that LTFs are generated from the epithelium itself, during the re-epithelialisation process.
5. Design DNA probes to estimate normal caecal ecosystem Fluorescent in situ hybridisation (FisH) probes, derived by 16s rRNA gene sequencing, successfully identified Gram positive and negative organisms in caecal and faecal material from normal horses. The results both validated FisH as a useful tool for studying hindgut bacterial populations and showed potential as a means of analysiing the caecal microflora during laminitis induction.
6. Search literature for fructan data in Australian pasture
A collection of the relevant literature applying to grazing horses has been established. Contact has been made with an expert in Colorado USA (Katy Watts) who is an agronomist specialising in soluble carbohydrates as a cause of laminitis. It would be profitable if Dr Watts was employed by RIRDC to visit Australia for 8 weeks to investigate fructan in Australian grasses and produce a report suitable for Australian horse owners.
Milestone 6 can be achieved employing a specialist agronomist.
7 Extract and cut DNA from microbes associated with laminitis
The implanted caecal fistula enabled analysis of the caecal fluid during the development of laminitis. Rapid proliferation of OF-utilising organisms was observed in all samples prior to the onset of laminitis. Caecal and faecal isolates from horses developing laminitis were subjected to RFLP analysis and formed 10 operational taxonomic units (OTU). Representative isolates from each OTU were sequenced and Streptococcus bovis was demonstrated to be the predominant, culturable, RUO found in the equine hindgut prior to the onset of equine laminitis.
8. Establish profile of laminitis caecal microbiota by culture
Changes in caecal microflora populations were documented using the novel habitat-simulating medium containing Raftilose (IECA). Using this media, changes in raftilose utilising organism (RUO) numbers in relation to other anaerobic bacteria were monitored throughout the course of experimentally induced laminitis. IECA and basal medium 10 (BM10) was used to determine viable counts for both Raftilose utilising organisms (RUO) and total anaerobes every 8 hours during Raftilose induced laminitis.
One hundred and fifty-six RUO were collected throughout the course of fructan-induced laminitis from 5 horses. Rapid proliferation of Raftilose-utilising organisms was observed in all samples prior to the onset of laminitis. Post-laminitis, however, S. bovis was not as prevalent, with Escherichia coli becoming established as the predominant RUO. RUO increased by 2.66 and 5.01 log units in caecal fluid and faeces respectively, while the total anaerobe counts obtained on basal medium 10 (BM10) increased by 1.05 and 1.92 log units in the caecum and faeces respectively. The pH of caecal fluid and faeces decreased to a pH of 4.4 at 8 hours and 4.2 at 16.25 hours and returned to normal within 16 hours in both samples.
9. DNA probes to identify laminitis causing caecal ecosystem
A fluorescence in situ hybridisation (FISH) probe targeted to S. bovis (Sb127) and a few other closely related organisms was developed and applied to the faecal samples collected throughout the course of equine laminitis. Sb127-targeted organisms were demonstrated to increase significantly between 8-16h post-fructan administration in all horses. These organisms then rapidly declined and were no longer observable in faecal specimens by 32-48h. These results both validated FISH as a useful tool for studying changes in known bacterial populations throughout the course of equine laminitis and added further strength to the hypothesis that RUO proliferate rapidly between 8-16h. These studies confirm the rapid proliferation of Streptococcus spp. observed in gram stains and by viable counting.
10. PCR the RNA from microbes associated with laminitis
The PCR products from laminitis associated isolates were subjected to restriction fragment length polymorphism (RFLP) analysis and grouped into operational taxonomic units (OTUs). Isolates with identical RFLP banding patterns were grouped into the same OTU.
11. Quantify fructan fluxes in pasture on daily and annual basis
Since commencing this project data has become available in publications by authors specializing in agronomy and animal production. The recruitment of agronomist Katy Watts (as suggested above) could achieve this milestone.

Publications Pollitt, C.C., and van Eps A.W. (2004). Prolonged, continuous distal limb cryotherapy in the horse. Equine vet. J. 36: 216-220.
Kyaw-Tanner, M. and Pollitt, C. C. Equine laminitis: increased transcription of matrix metalloproteinase-2 (MMP-2) occurs during the developmental phase. Equine vet. J. 36:221-225.
French, K.R. and Pollitt, C.C. (2004). Equine laminitis: loss of hemidesmosomes in hoof secondary epidermal lamellae correlates to dose in an oligofructose induction model: an ultrastructural study. Equine vet. J. 36: 230-235.
French, K.R. and Pollitt, C.C. (2004). Equine laminitis: cleavage of L5 associated with basement membrane dysadhesion. Equine vet. J. 36: 242-247.
van Eps, A.W. and Pollitt, C.C. (2004). Equine laminitis: cryotherapy reduces the severity of the acute lesion. Equine vet. J. 36: 255-260.
French, K.R. and Pollitt, C.C. (2004). Equine laminitis: glucose deprivation and MMP activation induce dermo-epidermal separation in vitro. Equine vet. J. 36: 261-266.
French, K.R. and Pollitt, C.C. (2004) Equine laminitis: congenital, hemidesmosomal plectin deficiency in a Quarter Horse foal. Equine vet. J. 36: 299-303.
Kyaw-Tanner, M & Pollitt C. C. (2004). Equine laminitis: increased transcription of matrix metalloproteinase-2 (MMP-2) occurs during the developmental phase. Equine vet. J. (in press).
French, K.R. & Pollitt, C.C. (2004). Equine laminitis: loss of hemidesmosome ultrastructure correlates to dose in an oligofructose induction model. Equine vet. J. (in press).
French, K.R. & Pollitt, C.C. (2004). Equine laminitis: cleavage of key hemidesmosome proteins associated with basement membrane dysadhesion. Equine vet. J. (in press.)
van Eps, A.W. & Pollitt, C.C. (2004). Equine laminitis: cryotherapy prevents development of the acute lesion. Equine vet. J. (in press).

Completed Projects - Other

Project Title Prototype Environmental Management System (EMS) for the horse keeping community - implementation, auditing and monitoring

RIRDC Project No.: EPS-1A

Researcher: Ms. Jacqueline Frizenschaf

Organisation: Environment Protection Authority (SA)

Phone: (08) 8139 9919

Fax: (08) 8139 9901

Email: frizenschaf.jacqueline@saugov.sa.gov.au

Objectives ·1 Phase 1 - Produce a practical prototype EMS framework for the horse keeping community as a prerequisite and a point of reference for performance-based auditing and development approval.
·2 Phase 2 - Develop and test a self audit kit for horse keepers through close collaboration between the EPA and the horse keeping communities in different states.
·3 Phase 3 - Develop an awareness raising and EMS adoption program to reach out to the greater horse keeping community, in the Mt. Lofty Ranges watershed (incl. and interstate component).

Background The project titled ‘Prototype Environmental Management System (EMS) for the Horse Keeping Community – Implementation, Auditing and Monitoring’ was a 3-year RIRDC-funded research project (referred to as ‘Horse EMS project’ in this report). The project explored the value of an environmental partnership approach between South Australian (SA) horse industry representatives and government to achieve environmental improvements on horse properties on a voluntary basis.

Research The project methodology included a step-by-step design towards the final outcomes, especially the development of a self-audit kit to be promoted to the horse keeping community using a targeted outreach approach. The applied framework used throughout the project was modelled according to the basic elements of International Standardisation Organisation (ISO) 14001’s EMS methodology. This provide a systematic approach to capture the potential risks of horse management activities to the environment; to prioritise action to avoid or mitigate these risks; and to monitor the success of the on-property action(s) with the implied option of continually improving on-property land management.
To achieve the project objectives, the overall methodology was strongly based on a partnership and consultative approach during each project phase. The primary project partners included the EPA and Horse SA, supported by other government and non-government entities. Principles commonly known as ‘participatory development (or management)’ were used to ensure that the project products were endorsed by all parties potentially affected or involved in the future use of the products. A pilot group, comprised of a cross section of the horse industry sector in South Australia was tasked to assist in the development, testing and review of the products. To ensure endorsement from the horse property owners as well as government, environmental land management principles (to be promoted via the products) were reviewed by scientific and government experts (to ensure a sound scientific basis), and tested by the pilot group (to ensure their likely adoption).
The project was rolled out in the Mount Lofty Ranges Watershed, the primary drinking water supply area for metropolitan Adelaide. Testing such a voluntary management effort in an environmentally sensitive region provided a sense of relevance to the project outcomes.
Additional, underpinning tools used as part of the project included: the use of workshop and review sessions; field days; monitoring performance and evaluation for both the desired project outcomes and the actual environmental improvements on the ground. Qualitative and semi-quantitative evaluation tools comprised pilot participant surveys at the beginning and end of project; feedback surveys at the end of workshops and field days; participant reports /presentations on their on farm improvements; mid-term project review with project team.
Throughout the three years or research, the project made extensive use of promotional opportunities such as media releases, and presentations at national and international conferences and local events.

Outcomes · Phase 1 – Prototype EMS developed as a basis for developing a self-assessment tool for horse keepers. For the first time in Australia, a comprehensive register was compiled which included the most common horse keeping activities and their potential environmental impacts. This list was a valuable basis for a focused approach in Phase 2. The development of the register resulted in a mutual understanding of the fundamental environmental issues potentially caused by horse keeping activities. A total of three workshops, two field days and 10 site visits were held.
Additional products. During Phase 1 workshops and field days a significant knowledge gap of the pilot group (and hence, the larger horse keeping community) became apparent. There was no comprehensive understanding about how the biophysical conditions of a horse property related to the potential enforced detriment of the natural conditions if not managed right. As a result, a Step-by-Step Guide for Land Capability Assessment was devised for the pilot group using in kind support by the AMLR NRMB’s Land Management Program. The document guides the user through the process of assessing the biophysical conditions of the property and provides instructions on how to determine environmentally sustainable stocking rates on the horse property, appropriate locations for stables, yards and other intensive horse keeping structures and property maintenance requirements. In addition, a fence post sign was provided to each pilot group participant.
· Phase 2 – Self-assessment tool (now called "Action Planner") developed. The Action Planner was developed based on a series of workshops and review sessions identifying a pragmatic and user-friendly organisation and structure of the document and its scientific content. It was created to enable the horse keeping community to conduct a quick (about 20-minute long) assessment of how their property management fares. The picture-free, straightforward Action Planner has been tested interstate and received enthusiastic responses. A total of one workshop, one field day and two expert review sessions were held.
Additional products. While drafting the Action Planner, it became evident that enticing horse owners to adopt sound environmental practices would be greatly enhanced if a "how to" guide was added to the package. The Management Guidelines, now accompanying the Action Planner, were developed as a result. Supplementary funding allowed the professional production of this document, which parallels the structure of the Action Planner and contains supporting colour photographs. In addition, one environmental horse poster and several (conference) Horse EMS banners were produced by the EPA and Horse SA.
· Phase 3 – Adoption and outreach strategy developed. A well thought-through adoption and outreach strategy was considered an essential element of the project. Such a strategy would allow Horse SA, the custodian of future activities on environmental management by horse keepers, to target its efforts strategically with the goal of ultimately raising the (environmental) performance of the horse keeping community. The strategy was devised based on substantial consultation with both government and the horse keeping community. It now contains a prioritised sequence of strategic actions that provide the highest potential for on-ground environmental improvement results. One workshop and two expert review sessions were held.
Additional products. The project team soon realised that the development of an adoption and outreach strategy would benefit from expert input. Additional NHT (Envirofund) funding enabled the team to engage Porter Novelli Pty Ltd in the development of the strategy. Porter Novelli used tools such as workshops and surveys to identify different target groups for an outreach program. In addition, the strategy is underpinned by the principles of community-based social marketing as promoted by Dr. Doug McKenzie-Mohr.

Implications Although all initially specified project objectives and desired products were achieved, the overall project success truly lies in the successful application of the participatory development and management approaches used throughout the project. Environmental management for horse keepers won’t wind down after this project, but will carry on into the future, with the following new initiatives established: · community-of-practice forum;
· adoption and outreach monitoring;
· periodic updating of Action Planner and Management Guidelines; and
· extension of training and research to other ‘lifestyle’ rural land managers, which often manage horses on their properties.

Publications This project enabled the publication of the following free information resources (available from the Horse SA web site starting in August 2006):
· Community-of-practice website (maintained by Horse SA), displaying:
· The Action Planner for Horse Keepers - a tool to assist horse keeper assess and improve environmental management
· Management Guidelines – to accompany the Action Planner for horse properties

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Last updated: August 2006 Copyright © RIRDC 2006
http://www.rirdc.gov.au/comp06/hor1.html

2.4 HORSES - COMPLETED PROJECTS
PROJECT No		PROJECT TITLE		RESEARCHER	PHONE	ORGANISATION
Information Technology for the Horse Industry
UM-66A		A prospective study of pre sale radiographs of thoroughbred yearlings	Prof Andrew Clarke		(03) 9731 2314	University of Melbourne
US-124A		Communications and information programs for the horse industry	Ms Shirley Ray		(02) 9352 6932	University of Sydney
To improve the management of growing and developing horses
US-129A		Risk factors for injuries in thoroughbred racehorses during commercial training	Prof David Evans		(02) 9351 2474	University of Sydney
To improve the overall health and welfare of the horse
OFF-1A		The production of a poster and booklet on poisonous weeds for horses	Ms Mellisa Offord		(02) 8901 0622
UQ-114A		Penetration of pharmacological agents through equine skin	Dr Paul Mills		(07) 3365 2964	University of Queensland
To more effectively quantify the environmental impact on the horse industry
UQ-119A	Transfer of weeds to Qld native forests on horses hooves			Dr. Christopher Pollitt	(07) 3365 2063	The University of Queensland
Respiratory Diseases-Improving the Respiratory Health of Horses
UM-67A		Effects of Stud Management on Ecology of Virulent Rhodococcus equi and Further studies of the ecology of virulent R. equi on horse studs - 2002 stud season	Dr Glenn Browing		(03) 8344 742	University of Melbourne
Lameness and Limb Injury-Developing More Effective Methods for Prevention, Diagnosis and Treatment
UM-58A		Epidemiology and risk factor analysis of racetrack fatalities	Prof. Andrew Clarke		(03) 9731 2314	The University of Melbourne
UQ-100A		Pasture fructan concentration as a cause of equine laminitis	Dr. Christopher Pollitt		(07) 3365 2063	The University of Queensland
Other
EPS-1A		Prototype Environmental Management System (EMS) for the horse keeping community - implementation, auditing and monitoring	Ms Jacqueline Frizenschaf		(08) 8139 9919	Environment Protection Authority SA

Project Title	A prospective study of pre sale radiographs of thoroughbred yearlings
RIRDC Project No.:	UM-66A
Researcher:	Prof Andrew Clarke
Organisation:	The University of Melbourne
Phone:	(03) 9731 2314
Fax:	(03) 9731 2377
Email:	afclarke@unimelb.edu.au
Objectives	·1 To identify the incidence of radiographic lesions in yearling thoroughbred horses presented for sale in Australia. ·2 To provide an objective assessment of the impact these findings have on the racing careers (e.g. number of starts, earning capacity, age of first start) of the horses involved.
Background	Radiograph repositories were introduced to auction sales in Australia in 2003 for pre-purchase assessment of orthopaedic lesions in Thoroughbred yearlings. There were 3,905 yearlings entered into 8 major national yearling sales in this year. Of these 2,847 had radiographs submitted to the radiograph repository. 2,412 of the 2,847 radiograph sets (81,741 individual x-rays) were made available for the present study. The radiographs were reviewed one of four veterinary specialist radiologists. In addition 167 sets were randomly selected and each radiologist read these sets twice. Sale catalogues and results were provided by the sales companies for all eight sales. The Australian Stud Book was used to verify catalogue information and ascertain if yearlings were named for racing or exported overseas. The i-RIS database was used to retrieve the race records of yearlings at two and three years of age. Melissa Jackson is the PhD student working with this project. This project and the results generated by Melissa will be the basis of her thesis. Contact details: Melissa Jackson, Equine Centre, University of Melbourne, 250 Princes Highway, Werribee Vic 3030. Ph: 03 9731 2316, Fax: 03 9731 2246, Email: m.jackson2@pgrad.unimelb.edu.au
Research	This project has five major components: 1. Comparison between radiographed and non-radiographed yearlings relating to sale and two-year-old race performance. 2. Prevalence of radiographically detectable orthopaedic lesions in radiographed yearlings. 3. Association between lesion presence and race performance at two and three years of age. 4. Race profile at two and three years of age and an economic analysis of return on money invested in the purchase of the yearling. 5. An assessment of the quality of radiographs and repeatability of assessments of radiograph quality and lesions by specialist radiologists.

Project Title	Communications and information programs for the horse industry
RIRDC Project No.:	US-124A
Researcher:	Ms Shirley Ray
Organisation:	The University of Sydney
Phone:	(02) 9463 9305
Fax:	(02) 9463 9100
Email:	shirleyr@camden.usyd.edu.au
Objectives	To continue to ensure widespread industry support for the RIRDC Equine R&D Program and to enable the results of the R&D to be used by the industry, a strong communication program is necessary to ensure results get to end users. ·3 A database of equine organisations. ·4 A database for communication with the industry. ·5 RIRDC Equine Research News. This newsletter of 8 pages, published quarterly, will provide the industry with information on topical research from around the world as well as updates on current RIRDC funded projects, interviews with research workers, workshops and invited reviews. ·6 RIRDC Internet Web Site. An Internet web site for the RIRDC Horse Program which will include links for online purchase of RIRDC publications, download of the RIRDC Equine Research News, search facility for past issues and a subscription facility to receive the RIRDC Equine Research News via the Internet.
Research	Each newsletter published had a print run of approximately 9,500 copies, all of which were distributed to the various sectors of the horse industry. The newsletter contained summaries of articles of local and international interest, interviews with researchers in particular fields, announcements of new RIRDC funded projects and updates of all currently funded projects, and items of general interest to the horse industry including RIRDC events and new publications. Articles from the newsletter have been reprinted in many local and international publications including: The Horse (USA), Sport Horse Journal, Breeding and Racing, Race Calendars and Hoofbeats. The RIRDC Equine Research Program Web site contains articles that have been printed in the RIRDC Equine Research News; these articles are accessible through an index of subject areas. A local search engine allows the user to search the site for a keyword. The site has links to relevant pages of the RIRDC web site, including the Five Year Plan, Objectives and Strategies of the Equine Research Program and RIRDC’s e-shop. A links page includes links to local and international research institutions and other sites of interests. At least 50 websites contain a link to the RIRDC Equine Program website and these include: Cyberhorse, Australian Racing Board; Australian Horse Resources and Australian Horse Net Service. All past issues of the newsletter are available for download in PDF format which can be viewed using Adobe Acrobat. This means that anyone with Internet access can view the newsletter in its colour print format. The mailing list for the RIRDC Equine Research Program is updated regularly and contains contact details of Australian Horse Associations, Research Institutions and Personnel, Horse Publications and other media and individuals with an interest in the horse industry. The mailing list contains over 2,000 entries

Project Title	Risk factors for injuries in thoroughbred racehorses during commercial training
RIRDC Project No.:	US-129A
Researcher:	Associate Professor David Evans
Organisation:	The University of Sydney
Phone:	(02) 9351 2474
Fax:	(02) 9351 3957
Email:	davide@vetsci.usyd.edu.au
Objectives	·7 The aim of this study was to investigate the epidemiology of MS injuries in Australian two- and three-year-old Thoroughbred racehorses. More specifically the aims were to describe the incidence and impact of MS and determine risk factors for MS injuries, in particular shin soreness.
Background	Musculoskeletal (MS) injuries have been identified as a common health problem in Thoroughbred racehorses. The most common MS injuries in two- and three-year olds have been identified as shin soreness and problems involving the carpus and fetlock joints. These MS injuries incur both direct and indirect costs. After reviewing the literature it is clear that there is very limited understanding of the risk factors for MS injuries. In particular, there is almost a complete absence of any investigations that have collected data on daily training in an effort to understand the training related risk factors for MS injuries. Studies that utilize daily training records are fundamental to the understanding of training related risk factors for MS injuries. An understanding of the risk factors for MS injuries would facilitate the design of strategies that may reduce the occurrence and impact of MS injuries.
Research	During the study period 451 horses were enrolled during their two-year-old racing season. Training and injury information for these horses were collected directly from the trainer. The research confirmed previous research that MS injuries are a common problem in two- and three-year-old Thoroughbred racehorses. The results showed that injury prior to the first start had considerable impact on time to first start. However, preparations after the first MS injury do not appear to be adversely affected. The investigation of risk factors showed that increasing distance galloped at high speed increased the risk of MS injury.
Outcomes	Since 2000 the results of this study have been presented to a number of difference associations and audiences. These were: ·1 Australian Equine Veterinary Association ·2 Society of Veterinary Epidemiology and Preventive Medicine ·3 International Society Veterinary Epidemiology and Economics ·4 Institute of Veterinary and Biomedical Science, Massey University ·5 Australian and New Zealand racetrack managers ·6 Sydney metropolitan and provincial Thoroughbred trainers ·7 Australian Veterinary Association ·8 British Equine Veterinary Association ·9 International Horse Industry Symposium

Implications	The results of this study show that exposure to high-speed exercise is associated with an increased risk of injury. Unfortunately, racehorses must be exposed to high-speed exercise in order to prepare them for competition. Therefore, future research should focus on developing ways in which the risk associated with high-speed exercise can be reduced.
Publications	Cogger N., Perkins H., Hodgson D.R., Reid S.W.J. and Evans, D.L. (in press). "Risk factors for musculoskeletal injuries in two-year-old Thoroughbred racehorses." Preventive Veterinary Medicine.

Project Title	The production of a poster and booklet on poisonous weeds for horses
RIRDC Project No.:	OFF-1A
Researcher:	Ms Mellisa Offord
Organisation:	Mellisa Offord
Phone:	(02) 8901 0622
Fax:	(02) 9451 3286
Email:	mofford@optusnet.com.au
Objectives	·8 To produce a poster and booklet showing plants and trees that are poisonous to horses.
Outcomes	A book and poster have been produced and these were launched at the Equine Science Symposium held on the Gold Coast in June 2006. The book is titled Plants Poisonous to Horses – An Australian Field Guide. It is 127 pages and it is a guide for horse owners to help them prevent plant poisoning in their horses. It clearly outlines the factors that influence the risk of plant poisoning and the ways to reduce this risk. The types of toxins that can be found in plants are discussed, along with the signs of poisoning that may be seen in horses that have ingested toxic plant material. A description of each plant is provided, including structure, growth habit and identifying features. The book contains over 200 colour photos of poisonous plants. Management notes suggest ways of dealing with poisonous plants on properties. The book includes a comprehensive list of ornamental plants that are known to be toxic to horses. The reference list is a useful resource for further reading and includes the journal articles, industry papers, books, CD ROMs and Internet resources that were used to create this publication. The poster highlights 11 plants in Australia that are a risk to horses.
Publications	Offord M (2006). Plants Poisonous to Horses: An Australian Field Guide. RIRDC Publication No. 06/048

Project Title	Penetration of pharmacological agents through equine skin
RIRDC Project No.:	UQ-114A
Researcher:	Dr Paul Mills
Organisation:	The University of Queensland
Phone:	(07) 3365 2964
Fax:	(07) 3365 1255
Email:	p.mills@uq.edu.au
Objectives	·1 To develop in vitro and in vivo methodologies to measure drug movement through equine skin ·2 To measure the regional differences (e.g. neck vs back vs axilla vs lower leg) in drug penetration ·3 To measure transdermal penetration of drugs of interest to the equine industry, including non-steroidal anti-inflammatory drugs (NSAIDs), corticosteroids and analgesics (e.g. Fentanyl) ·4 To determine the effects of vehicles (e.g. DMSO) on penetration of drugs in the horse ·5 To determine the effects of skin damage (e.g. rashes) and altered microenvironment (e.g. bandages, saddles) on transdermal drug movement in the horse
Background	The skin is a protective layer preserving the integrity of the internal environment. The major barrier is the outermost layer, the stratum corneum. An increasing number of pharmaceutical agents are being developed for topical use in humans and animals. Advantages of transdermal drug penetration include; (i) ease of administration; (ii) increased compliance with use; (iii) avoidance of hepatic "first-pass" effect, and (iv) reduced gastric irritation. It is now mandatory for any topical preparation intended for human use to undergo extensive laboratory testing prior to registration. Recent adverse developments in veterinary topical medications (e.g. Bayticol in cattle and Frontline (fipronil) in dogs and cats) have shown that inadequate studies were undertaken prior to registration. The Australian Pesticides and Veterinary Medicines Authority (APVMA) have recently taken an active interest in prerequisites for registration of veterinary topical medications. Little is known about transdermal movement of drugs through equine skin. Many of the currently available products, such as corticosteroid creams and NSAID gels, are based on results from human or laboratory animal studies, but it is widely acknowledged that extrapolation of transdermal drug movement between species is unreliable. This may lead to adverse or unexpected results when these preparations are applied to horses. Indeed, an increasing number of positive swab samples collected during equine competition have been blamed on unexpected penetration through skin. In this proposed study, we will use scientifically credible in vitro and in vivo methods to investigate transdermal drug movement through equine skin. Drugs that may be applied topically and are of specific interest to the equine industry will be examines, including NSAIDs, corticosteroids and other analgesics (e.g. procaine and fentanyl). We will also investigate potential factors affecting transdermal drug movement, particularly region of application (e.g. neck vs groin), the effects of altered skin integrity (rashes or bandaging) and the effects of vehicles (e.g. DMSO, propylene glycol). The results from this study will be of benefit to all stakeholders in the equine industry. Information about where best to apply specific drugs to maximise transdermal passage, or regions to avoid if systemic concentrations are undesirable would be useful to veterinarians, owners and trainers. There in no information about drug movement through damaged skin and if medications are applied under bandages, yet many drugs are applied under these circumstances. The potential of inadvertent swab sample positives needs to the addressed by the industry and this research is a pro-active step towards developing understanding and expertise. It may also become mandatory for laboratory testing of drugs intended for topical application and having expertise and infrastructure to undertake this task would be extremely useful.
Research	Preliminary information will be determined using in vitro methodologies. Equine skin will be obtained from euthanased animals and placed in Franz-type diffusion cells. Pharmacological agents, in a variety of vehicles, can be applied to the stratum corneum as a donor phase and analytes passing into and through the skin can be measured. The diffusion cell is maintained at approximately 32oC, the expected temperature of the skin surface, using a water bath. The receptor phase can be varied to mimic conditions expected in the dermis and underlying tissues of the horse. Specific measurement of drug and metabolites in skin and passing through to the receptor phase with use either high performance liquid chromatography (HPLC) or scintillation counting (if radiolabelled drug is used). Two in vivo models will be used to confirm in vitro results and account for biological differences in living skin, such as changes in blood flow. Whole body studies where drug and/or vehicle are applied to the horse and analytes measured in blood and urine. A new technique, microdialysis, will also be developed in the horse. This involves placing semi-permeable probes into the epidermis and dermis, then directly measuring drug appearance in perfusate. Once preliminary data is obtained, models of skin damage (abrasion and/or delipidation) will be developed using harvested skin. This will permit direct testing, in vitro, of the effects of skin damage on transdermal drug movement. It is also hoped to develop and apply mathematical principles and available models to predict transdermal drug movement of new drugs or to assess the effects of alternative vehicles or the effects of skin damage.
Outcomes	The techniques that may be used to investigate transdermal drug delivery in the horse were verified, particularly for in vitro methods which can be used to screen drugs and specifically compare the effects of regional differences, vehicles and skin integrity. Skin was harvested from the thorax, groin and leg (dorsal metacarpal) of horses to investigate regional variation in the penetration of a number of commonly used drugs in horses. Significantly higher (P<0.001) maximum flux (Jmax; mol/cm2/hr) was measured when hydrocortisone was applied to skin from the leg, compared to thorax and groin, although significantly less hydrocortisone (P<0.001) was retained within skin from the leg at 24 hrs. Topical application of hydrocortisone in a vehicle containing ethanol would penetrate faster through leg skin from the lower leg, compared to the thorax or groin which, depending on cutaneous blood flow, may result in higher systemic drug concentrations or greater efficiency in treating local inflamed tissue. Similarly, a commercial formulation of methylsalicylate (Dencorub®) will achieve therapeutic levels of total salicylate beneath sites of topical application, with a faster and more pronounced response achieved through the leg region, compared to upper body. However, fentanyl Transdermal Therapeutic Systems (TTS or ‘patch’) applied to the leg region may have a small but significantly lower amount of fentanyl available systemically, compared to patches applied to the thorax or groin regions, which may affect the level of analgesia subsequently achieved in the horse. The effects of the vehicles phosphate buffered saline (PBS), ethanol (EtOH; 50% in PBS w/w) and propylene glycol (PG; 50% in PBS w/w) on in vitro transdermal penetration of testosterone was investigated in the horse. The maximum flux (Jmax) of testosterone was significantly higher for all sites when dissolved in a vehicle containing 50% ethanol or 50% propylene glycol, compared to PBS. In contrast, higher residues of testosterone were found remaining within the skin when PBS was used as a vehicle. This study shows that variability in clinical response to testosterone could be expected with formulation design. To investigate the effects of common skin preparations on transdermal drug penetration through equine skin in vitro, skin was exposed to the following treatments: Control (no treatment); Cleaned with aqueous chlorhexidine; cleaned with alcoholic chlorhexidine; Shaved; Tape stripped using elastoplast adhesive tape. A significantly higher proportion of methylsalicylate penetrated through alcoholic chlorhexidine and Shaved (P<0.01) and aqueous chlorhexidine and Tape stripped (P<0.05) treated skin, compared to Control, while significantly more Sa was recovered in the receptor phase of Tape stripped (P<0.01), alcoholic chlorhexidine, aqueous chlorhexidine and Shaved (P<0.05) treated skin, compared to Control. A significantly higher rate of penetration and shorter lag time was also noted for methylsalicylate following each skin pre-treatment, compared to control. The results showed that routine clinical techniques used to clean or prepare skin can significantly affect the rate and extent of penetration of a topically applied drug. This may result is more active drug available systemically enhancing efficacy and, possibly, the incidence of adverse effects to the particular drug.
Implications	There are no guidelines for the topical administration of drugs to horses prior to or during competition which this project has partially addressed. These guidelines and precautions for stakeholders in the horse industry will be disseminated and can be seen as a pro-active response to inadvertent positive swab samples. We now have a centre of expertise in the area of transdermal drug penetration in the horse which has already provided expert opinion to stewards in Australia and Ireland. Importantly, we now also have the expertise to investigate transdermal penetration of other drugs relevant to the equine industry. A peer-reviewed scientific review currently in press will provide guidelines and information for veterinarians so that they can understand transdermal drug penetration and advise their clients (owners, trainers, stewards) accordingly. The information generated about the effects of skin pre-treatment will have important implications for application of any topical medications to equine skin. For example, cleaning skin, shaving skin to remove hair or applying drugs after the removal of adhesive bandages will all affect the rate and amount of drug penetration. This research has established scientifically-credible testing methodologies which can be used to test potential new topical remedies and to advise registration bodies (especially the APVMA) about equine topical medications. This is important as, similar to human topical medications; there are increasing requirements before topical medications can be regarded as safe and efficacious.
Publications	Mills P.C. and Cross S.E. (2006) The effects of skin preparation on transdermal drug penetration in vitro through equine skin. Can J Vet Res (in press). Mills P.C. and Cross S.E. (2006) Transdermal Drug Delivery: Basic Principles for the Veterinarian. Vet J (in press- doi 10.1016/j.tvjl.2005.09.006). Mills P.C. and Cross S.E. (2006) Regional differences in transdermal penetration of fentanyl through equine skin. Res Vet Sci. (in press). Mills P.C. (2006) Vehicle effects on the penetration of testosterone through equine skin. Vet.Res.Comm. (in press). Mills P.C. and Cross S.E. (2006) Regional differences in the in vitro penetration of hydrocortisone through equine skin. J Vet.Pharm.Ther. 29: 25-30. Mills P.C. and Cross S.E. (2006) Regional differences in the in vitro penetration of methylsalicylate through equine skin. Vet.J. (in press doi 10.1016/j.tvjl.2005.08.023). There have also been a number of lay articles published in Australia (The Veterinarian), Ireland (Racing Times) and USA (Equus).

Project Title	Transfer of weeds to Qld native forests on horses hooves
RIRDC Project No.:	UQ-119A
Researcher:	Dr Christopher Pollitt
Organisation:	The University of Queensland
Phone:	(07) 3365 2063
Fax:	(07) 3365 1899
Email:	c.pollitt@uq.edu.au
Objectives	·6 Do horse faeces contain viable seeds which may germinate under suitable conditions or persist in surface soil. ·7 Are the plants that germinate from horse manure found beside the trails used by horse riders in forests of SE Queensland.
Background	Weeds accidentally introduced since European settlement constitute a threat to the ecological sustainability of native plant communities. Weeds may be introduced into native ecosystems via vehicular, human and animal vectors and there is anecdotal evidence that horse manure contains viable weed seed. A rigorous scientific protocol is necessary to discover if seeds in horse manure are viable and pose a threat when horses traverse areas of conservation significance.
Research	Horse faeces were collected from horses grazing an 80 hectare pasture without recent weed control and from race horses maintained in stables. Half of the faeces were dried and half kept wet before being mixed with a commercial, seed free, potting mix.
Outcomes	Ten plant species germinated in the pasture derived faeces (compared to one from stabled horses) and there was a higher seed load in the pasture faeces. Dominating seed germination was the grass Queensland Blue Couch (Digitaria ciliaris). Germination was 5 times greater in the dried faeces suggesting that prior drying favours grass seed germination. Only one native species, emu foot (Psoralea tenax) was found. All species that germinated in manure have current weed status.
Implications	The faeces of horses grazing weed infested pastures contain viable seed that, potentially, may infest native forests. However further work is required to determine if this actually occurs. To determine the role of horse faeces as a source of weeds, plants growing on 100 m transects of the sampled pasture and transects of the trails in nearby native forests will be compared. When complete these studies will supply objective data to assess the level of threat posed by horse riding in areas of native vegetation and will help formulate rational management strategies for recreational horse riding.
Publications	In press

Project Title	*Effects of Stud Management on Ecology of Virulent Rhodococcus equi* and Further studies of the ecology of virulent R. equi on horse studs - 2002 stud season**
RIRDC Project No.:	UM-47A and UM-67A
Researcher:	Glenn F Browning, James R Gilkerson and Gary Muscatello
Organisation:	School of Veterinary Science The University of Melbourne
Phone:	03 8344 7342
Fax:	03 8344 7374
Email:	glenfb@unimelb.edu.au
Objectives	·8 To develop methods to improve understanding of the factors that make some areas on horse farms a greater risk for development of rattles in foals and apply these to identifying factors that might be addressed by changes in management of farms.
Background	Although rattles is an important disease problem in young foals there are limited options for control. Current methods rely on reducing faecal contamination and dust on studs, but we have little understanding of whether these are the only factors that account for higher infection and death rates on some studs.
Research	This project aimed to develop novel methods for assessing the risk associated with particular horse farm environments. These methods were based on bacteriological sampling of air and soil on studs, then identifying those bacteria capable of causing rattles. Once these methods were developed they were applied to examination of different horse farms and different places on those farms. The method that was developed was also used to examine the breath of foals with rattles. This suggested that foals may be able to transmit virulent R. equi to each other.
Outcomes	The most dangerous areas on studs for foals are likely to be laneways and holding pens. Control may be aided by reducing the time that foals spend in these areas. In addition areas that had sandy, dry and/or acidic soils seemed to be a greater risk. Some control may also be achieved by avoiding close contact between foals, particularly if they are known to be suffering from rattles. Sampling air from foals may be a sensitive diagnostic technique for the early detection of infected foals.
Implications	Application of methods developed in this project have provided some suggestions for reducing the incidence of rattles and further use of these methods may enable the benefit obtained from different approaches to control to be assessed. Different approaches to treatment may also be able to be assessed.
Publications	Muscatello, G. Rhodococcus equi: the organism and epidemiology. Australian Equine Veterinarian 22:190-197 (2003). Muscatello, G., and Browning, G. F. - Identification and differentiation of avirulent and virulent Rhodococcus equi using selective media and colony blotting DNA hybridization to determine their concentrations in the environment. Veterinary Microbiology, 100:121-127 (2004). Muscatello, G., Gerbaud, S., Kennedy, C., Gilkerson, J. R., Buckley, T., Klay, M., Leadon, D. P., and Browning, G. F. – Comparisons between concentrations of Rhodococcus equi and virulent R. equi in the air of stables and paddocks on horse breeding farms in a temperate (European) climate. Equine Veterinary Journal, 38:263-265 (2006). Phumoonna, T., Muscatello, G., Chicken, C., Gilkerson, J. R., Browning, G. F., Barton, M. D., And Heuzenroeder, M. W. - Clinical evaluation of a peptide-ELISA based upon N-terminal B cell epitope of the Vap A protein for early diagnosis of Rhodococcus equi pneumonia in foals. Journal of Veterinary Medicine B, 53:126-32 (2006). Muscatello, G., Gilkerson, J. R., and Browning, G. F. - Comparison of two selective media for the recovery, isolation, enumeration and differentiation of Rhodococcus equi. Submitted. Muscatello, G., Anderson, G. A., Gilkerson, J. R., and Browning, G. F. - Associations between the ecology of virulent Rhodococcus equi and the epidemiology of R. equi pneumonia on Australian Thoroughbred farms, Applied and Environmental Microbiology, in press (2006). Muscatello, G., Gilkerson, J. R., And Browning, G. F. - Detection of virulent Rhodococcus Equi in exhaled air samples from foals. Submitted.

Project Title	Epidemiology and risk factor analysis of racetrack fatalities
RIRDC Project No.:	UM-58A
Researcher:	Prof Andrew Clarke
Organisation:	The University of Melbourne
Phone:	(03) 9731 2314
Fax:	(03) 9731 2377
Email:	afclarke@unimelb.edu.au
Objectives	·9 This project will deliver an improved understanding of the causes of and risk factors associated with racetrack fatalities. This will translate into strategies to address this complex multifactorial problem. This in turn will result in reduced wastage within the industry and have positive welfare implications.
Background	This project was the first in Australia to combine a prospective post mortem case series and large-scale retrospective observational studies to investigate the causes of and risks associated with racehorse fatality in flat and jump starts. The study was encouraged by the Victorian racing industry in response to industry and community concerns regarding high wastage rates of racehorses as well as jockey injuries and fatalities in racing. The purpose of this study was to identify risk factors that could be modified to reduce the risk of fatality in Thoroughbred racing in Victoria. Dr Lisa Boden was the PhD student working with this project. This project and the results generated by Lisa form the basis of Lisa’s thesis which has been successfully defended. Contact details: "Lisa Boden" lboden@staffmail.ed.ac.uk
Research	The following studies have been achieved as a result of this project: A. Post mortem survey of Thoroughbred mortalities on racecourses in Victoria. B. Risk of fatality and causes of death of Thoroughbred horses associated with racing in Victoria, Australia between 1989 and 2004. C. Risk factors for Thoroughbred racehorse fatality in flat races in Victoria between 1989 and 2004. D. Risk factors for Thoroughbred racehorse fatality in jump races in Victoria between 1989 and 2004. E. Estimated attributable fractions for racehorse fatalities in Victoria.
Outcomes	The results of the post mortem study showed that catastrophic limb injury was the major cause of fatality and that sudden death is a more important contributor to racing fatalities than previously reported in Victoria. The risk of fatality in flat starts was lower in Victoria (0.44 fatalities per 1000 starts) than that reported in North America (1.4-1.7 fatalities per 1000 starts and the UK (0.8-0.9 fatalities per 1000 starts) but the risk of fatality in jump starts was greater in Victoria (8.3 fatalities per 1000 starts) than that reported overseas (3.9 fatalities per 1000 starts) in North America and 5.6 fatalities per 1000 starts in the UK. The risk of fatality in flat starts appeared to increase over the 15 year study period but it remains unclear whether this was due to a genuine increase in risk (resulting from changes in such factors as climate or racing practices over time) or to increased stringency of reporting of proportions of fatalities. The risk of fatality in jump starts remained constant over the study period despite jump racing reviews that recommended changes to hurdle and steeple races to improve safety. The risk of fatality on city racecourses was 2.0 times that on country racecourses. Risk factors associated with increased odds of fatality in flat starts were horse gender (stallions and geldings compared with mares), one or more starts in the 31-60 day period prior to the case or control start, the going of the track (fast or good surfaces), racecourse location (city compared with country tracks), increasing race length and race year . Risk factors for jump starts included the number of flat and jump starts accumulated in the racing career prior to the study start, one or more starts in the 1-14 days prior to the case or control start, race type (steeplechase compared with hurdle races), track location (city compared with country tracks) and calendar year. Of the risk factors identified as having high population attributable fraction estimates, the most readily modifiable were: track condition in flat races (fast or good tracks), race type in jump races (steeplechase events) and the number of prior flat starts in racing career for horses in jump races. Of the risk factors identified as having high population attributable fraction estimates, the most readily modifiable were: track condition in flat races (fast or good tracks), race type in jump races (steeplechase events) and the number of prior flat starts in racing career for horses in jump races. Jump starts by horses with shorter racing careers were more likely to result in fatality. Over the course of a racing career, horses that started in more flat starts prior to the case or control jump start were more likely to die in the next jump start than horses that had accumulated less flat racing experience. Additionally, increased numbers of jump starts prior to the study start were also associated with increased odds of fatality. The odds of fatality in jump starts increased with each additional flat start accumulated in the racing career. The average number of career flat starts for horses starting in flat races was 17.2 whereas horses starting in jump races had already accumulated an average of 33 career flat starts. If horses were allowed to start in up to 32 flat starts (and no more) before beginning a jump racing career, the risk of fatality in jump starts may decrease by 50% (approximately eight fatalities per year).
Implications	This project identified specific risk factors for fatality in flat and jump racing in Victoria. The results of this study should be utilised to direct further research into starts, horses and races at high risk so that appropriate intervention strategies can be implemented to improve the welfare of horses racing in Victoria. Specific recommendations made to the industry as a result of this project include: •1 Continuation and expansion of post mortem program to include fatalities from country tracks and the collection of blood from cases of sudden death by racetrack veterinarians. •2 Formalised monitoring of injuries and fatalities •3 Standardisation of track rating methods and review of current policy regarding racing on ‘good’ tracks •4 Further investigation of differences between risk in hurdles and steeplechase races •5 Restriction of number of flat starts horses can accumulate before starting a jump racing career

Project Title	Pasture fructan concentration as a cause of equine laminitis
RIRDC Project No.:	UQ-100A
Researcher:	Dr Christopher Pollitt
Organisation:	The University of Queensland
Phone:	(07) 3365 2063
Fax:	(07) 3365 1899
Email:	c.pollitt@uq.edu.au
Objectives	·1 The cause of grass founder will be understood and a strategy for diagnosis, prevention and treatment will be available ·2 A strategy for pasture and grazing management will be available so that horses and ponies do not consume sufficient fructan to develop laminitis.
Research	1. Establish that dosing horses with Raftilose causes laminitis Horses dosed with Raftilose or oligofructose (OF) at 3 dose rates (7.5;10 & 12.5g/kg) developed clinical and histological laminitis in at least one foot. Higher doses of OF were associated with significantly more severe (grade 3) histological laminitis (p<0.05). All the horses developed profuse diarrhoea at around 18h that ceased by 36-44h. Pyrexia and elevated heart rate peaked between 16 and 20h and returned to near normal by 48h. Faecal pH began to fall within 4 hours of dosing, reaching its lowest value (3.7) at around 18h. Blood D-lactate, of bacterial origin, peaked at 24 hours and then returned to the normal zero. Since all horses developed laminitis, the results establish that a storage carbohydrate, other than starch, can induce the disease. OF is a principal non-structural carbohydrate of pasture grasses, suggesting it is grass OF that causes laminitis and not other substance(s), such as dietary amines. Compared with traditional alimentary overload with starch, OF dosing resulted in reduced morbidity and no deaths, indicating that alimentary overload with fructan is an efficient and more humane experimental model for the induction of acute laminitis. A paper entitled "Equine laminitis: experimental induction with oligofructose by C.C. Pollitt* and A. W. van Eps was submitted to the Equine Veterinary Journal in mid 2003. The referees were not satisfied that 3 pairs of horses dosed with oligofructose (OF) at 3 dose rates could indicate significant trends (despite our professional statistician’s explanation of the methods used). Publication has been deferred until 4 more horses have been dosed with the median dose (10/kg Bwt OF). 2. Establish surgical caecal cannulation The difficulties that were encountered with caecal cannulation in 2002 have been overcome by using an alternative 2-step method. In stage 1 an adhesion was created between the caecum and the skin after the muscle in between was surgically removed. In stage 2, 14 days later, access to the caecal lumen was established by trocarisation of the adhesion and a soft, rubber caecal cannula was introduced. One horse has been cannulated (HGF1) and laminitis was induced successfully giving us insight into events in the caecum for the first time. Two horses have had stage 1 surgery and are ready for cannula insertion (stage 2). More cannulations and laminitis inductions are planned for 2005. 3. Establish profile of normal caecal microbiota by culture Profiles of normal caecal microflora populations were documented through the creation of novel habitat-simulating medium containing Raftilose as the sole energy substrate (incubated equine caecal-fluid agar - IECA). Using this media, raftilose utilising organism (RUO) numbers in relation to other anaerobic bacteria were established for normal caecal contents. 4. Purify LTFs by filtration The trigger factors responsible for initiating laminitis remain elusive. It is clear however that the factors responsible for triggering the onset of laminitis occur in the equine hindgut 8-16h post fructan administration. Furthermore, the dominance of S. bovis, during this period implicates it as the aetiological agent of equine laminitis. Attempts to purify laminitis trigger factors (LTFs) from large volumes of S bovis culture media, using centrifugal filtration equipment, were not successful. We have developed a new casein containing medium and confirmed by zymography that S bovis produces a protease in the presence of casein. Analysis by zymography in gelatin containing PAGE gels showed the S bovis protease (SBP) was a gelatinase and possibly a metalloproteinase. Since the activated MMPs that cause laminitis are also gelatinases we investigated the possibility that SBP was a LTF. SBP was not inhibited by batimastat thus diminishing its likely resemblance to lamellar MMP. Interestingly caecal liquor taken when S bovis proliferation during OF induced laminitis had peaked contained very high concentrations of SBP. SBP and clarified caecal liquor from horses developing laminitis did not induce lamellar separation in vitro suggesting that LTFs are not of bacterial origin. It seems more likely that S bovis proliferation and concomitant protease production damages the mucosal epithelium of the hindgut and that LTFs are generated from the epithelium itself, during the re-epithelialisation process. 5. Design DNA probes to estimate normal caecal ecosystem Fluorescent in situ hybridisation (FisH) probes, derived by 16s rRNA gene sequencing, successfully identified Gram positive and negative organisms in caecal and faecal material from normal horses. The results both validated FisH as a useful tool for studying hindgut bacterial populations and showed potential as a means of analysiing the caecal microflora during laminitis induction. 6. Search literature for fructan data in Australian pasture A collection of the relevant literature applying to grazing horses has been established. Contact has been made with an expert in Colorado USA (Katy Watts) who is an agronomist specialising in soluble carbohydrates as a cause of laminitis. It would be profitable if Dr Watts was employed by RIRDC to visit Australia for 8 weeks to investigate fructan in Australian grasses and produce a report suitable for Australian horse owners. Milestone 6 can be achieved employing a specialist agronomist. 7 Extract and cut DNA from microbes associated with laminitis The implanted caecal fistula enabled analysis of the caecal fluid during the development of laminitis. Rapid proliferation of OF-utilising organisms was observed in all samples prior to the onset of laminitis. Caecal and faecal isolates from horses developing laminitis were subjected to RFLP analysis and formed 10 operational taxonomic units (OTU). Representative isolates from each OTU were sequenced and Streptococcus bovis was demonstrated to be the predominant, culturable, RUO found in the equine hindgut prior to the onset of equine laminitis. 8. Establish profile of laminitis caecal microbiota by culture Changes in caecal microflora populations were documented using the novel habitat-simulating medium containing Raftilose (IECA). Using this media, changes in raftilose utilising organism (RUO) numbers in relation to other anaerobic bacteria were monitored throughout the course of experimentally induced laminitis. IECA and basal medium 10 (BM10) was used to determine viable counts for both Raftilose utilising organisms (RUO) and total anaerobes every 8 hours during Raftilose induced laminitis. One hundred and fifty-six RUO were collected throughout the course of fructan-induced laminitis from 5 horses. Rapid proliferation of Raftilose-utilising organisms was observed in all samples prior to the onset of laminitis. Post-laminitis, however, S. bovis was not as prevalent, with Escherichia coli becoming established as the predominant RUO. RUO increased by 2.66 and 5.01 log units in caecal fluid and faeces respectively, while the total anaerobe counts obtained on basal medium 10 (BM10) increased by 1.05 and 1.92 log units in the caecum and faeces respectively. The pH of caecal fluid and faeces decreased to a pH of 4.4 at 8 hours and 4.2 at 16.25 hours and returned to normal within 16 hours in both samples. 9. DNA probes to identify laminitis causing caecal ecosystem A fluorescence in situ hybridisation (FISH) probe targeted to S. bovis (Sb127) and a few other closely related organisms was developed and applied to the faecal samples collected throughout the course of equine laminitis. Sb127-targeted organisms were demonstrated to increase significantly between 8-16h post-fructan administration in all horses. These organisms then rapidly declined and were no longer observable in faecal specimens by 32-48h. These results both validated FISH as a useful tool for studying changes in known bacterial populations throughout the course of equine laminitis and added further strength to the hypothesis that RUO proliferate rapidly between 8-16h. These studies confirm the rapid proliferation of Streptococcus spp. observed in gram stains and by viable counting. 10. PCR the RNA from microbes associated with laminitis The PCR products from laminitis associated isolates were subjected to restriction fragment length polymorphism (RFLP) analysis and grouped into operational taxonomic units (OTUs). Isolates with identical RFLP banding patterns were grouped into the same OTU. 11. Quantify fructan fluxes in pasture on daily and annual basis Since commencing this project data has become available in publications by authors specializing in agronomy and animal production. The recruitment of agronomist Katy Watts (as suggested above) could achieve this milestone.
Publications	Pollitt, C.C., and van Eps A.W. (2004). Prolonged, continuous distal limb cryotherapy in the horse. Equine vet. J. 36: 216-220. Kyaw-Tanner, M. and Pollitt, C. C. Equine laminitis: increased transcription of matrix metalloproteinase-2 (MMP-2) occurs during the developmental phase. Equine vet. J. 36:221-225. French, K.R. and Pollitt, C.C. (2004). Equine laminitis: loss of hemidesmosomes in hoof secondary epidermal lamellae correlates to dose in an oligofructose induction model: an ultrastructural study. Equine vet. J. 36: 230-235. French, K.R. and Pollitt, C.C. (2004). Equine laminitis: cleavage of L5 associated with basement membrane dysadhesion. Equine vet. J. 36: 242-247. van Eps, A.W. and Pollitt, C.C. (2004). Equine laminitis: cryotherapy reduces the severity of the acute lesion. Equine vet. J. 36: 255-260. French, K.R. and Pollitt, C.C. (2004). Equine laminitis: glucose deprivation and MMP activation induce dermo-epidermal separation in vitro. Equine vet. J. 36: 261-266. French, K.R. and Pollitt, C.C. (2004) Equine laminitis: congenital, hemidesmosomal plectin deficiency in a Quarter Horse foal. Equine vet. J. 36: 299-303. Kyaw-Tanner, M & Pollitt C. C. (2004). Equine laminitis: increased transcription of matrix metalloproteinase-2 (MMP-2) occurs during the developmental phase. Equine vet. J. (in press). French, K.R. & Pollitt, C.C. (2004). Equine laminitis: loss of hemidesmosome ultrastructure correlates to dose in an oligofructose induction model. Equine vet. J. (in press). French, K.R. & Pollitt, C.C. (2004). Equine laminitis: cleavage of key hemidesmosome proteins associated with basement membrane dysadhesion. Equine vet. J. (in press.) van Eps, A.W. & Pollitt, C.C. (2004). Equine laminitis: cryotherapy prevents development of the acute lesion. Equine vet. J. (in press).

Project Title	Prototype Environmental Management System (EMS) for the horse keeping community - implementation, auditing and monitoring
RIRDC Project No.:	EPS-1A
Researcher:	Ms. Jacqueline Frizenschaf
Organisation:	Environment Protection Authority (SA)
Phone:	(08) 8139 9919
Fax:	(08) 8139 9901
Email:	frizenschaf.jacqueline@saugov.sa.gov.au
Objectives	·1 Phase 1 - Produce a practical prototype EMS framework for the horse keeping community as a prerequisite and a point of reference for performance-based auditing and development approval. ·2 Phase 2 - Develop and test a self audit kit for horse keepers through close collaboration between the EPA and the horse keeping communities in different states. ·3 Phase 3 - Develop an awareness raising and EMS adoption program to reach out to the greater horse keeping community, in the Mt. Lofty Ranges watershed (incl. and interstate component).
Background	The project titled ‘Prototype Environmental Management System (EMS) for the Horse Keeping Community – Implementation, Auditing and Monitoring’ was a 3-year RIRDC-funded research project (referred to as ‘Horse EMS project’ in this report). The project explored the value of an environmental partnership approach between South Australian (SA) horse industry representatives and government to achieve environmental improvements on horse properties on a voluntary basis.
Research	The project methodology included a step-by-step design towards the final outcomes, especially the development of a self-audit kit to be promoted to the horse keeping community using a targeted outreach approach. The applied framework used throughout the project was modelled according to the basic elements of International Standardisation Organisation (ISO) 14001’s EMS methodology. This provide a systematic approach to capture the potential risks of horse management activities to the environment; to prioritise action to avoid or mitigate these risks; and to monitor the success of the on-property action(s) with the implied option of continually improving on-property land management. To achieve the project objectives, the overall methodology was strongly based on a partnership and consultative approach during each project phase. The primary project partners included the EPA and Horse SA, supported by other government and non-government entities. Principles commonly known as ‘participatory development (or management)’ were used to ensure that the project products were endorsed by all parties potentially affected or involved in the future use of the products. A pilot group, comprised of a cross section of the horse industry sector in South Australia was tasked to assist in the development, testing and review of the products. To ensure endorsement from the horse property owners as well as government, environmental land management principles (to be promoted via the products) were reviewed by scientific and government experts (to ensure a sound scientific basis), and tested by the pilot group (to ensure their likely adoption). The project was rolled out in the Mount Lofty Ranges Watershed, the primary drinking water supply area for metropolitan Adelaide. Testing such a voluntary management effort in an environmentally sensitive region provided a sense of relevance to the project outcomes. Additional, underpinning tools used as part of the project included: the use of workshop and review sessions; field days; monitoring performance and evaluation for both the desired project outcomes and the actual environmental improvements on the ground. Qualitative and semi-quantitative evaluation tools comprised pilot participant surveys at the beginning and end of project; feedback surveys at the end of workshops and field days; participant reports /presentations on their on farm improvements; mid-term project review with project team. Throughout the three years or research, the project made extensive use of promotional opportunities such as media releases, and presentations at national and international conferences and local events.
Outcomes	· Phase 1 – Prototype EMS developed as a basis for developing a self-assessment tool for horse keepers. For the first time in Australia, a comprehensive register was compiled which included the most common horse keeping activities and their potential environmental impacts. This list was a valuable basis for a focused approach in Phase 2. The development of the register resulted in a mutual understanding of the fundamental environmental issues potentially caused by horse keeping activities. A total of three workshops, two field days and 10 site visits were held. Additional products. During Phase 1 workshops and field days a significant knowledge gap of the pilot group (and hence, the larger horse keeping community) became apparent. There was no comprehensive understanding about how the biophysical conditions of a horse property related to the potential enforced detriment of the natural conditions if not managed right. As a result, a Step-by-Step Guide for Land Capability Assessment was devised for the pilot group using in kind support by the AMLR NRMB’s Land Management Program. The document guides the user through the process of assessing the biophysical conditions of the property and provides instructions on how to determine environmentally sustainable stocking rates on the horse property, appropriate locations for stables, yards and other intensive horse keeping structures and property maintenance requirements. In addition, a fence post sign was provided to each pilot group participant. · Phase 2 – Self-assessment tool (now called "Action Planner") developed. The Action Planner was developed based on a series of workshops and review sessions identifying a pragmatic and user-friendly organisation and structure of the document and its scientific content. It was created to enable the horse keeping community to conduct a quick (about 20-minute long) assessment of how their property management fares. The picture-free, straightforward Action Planner has been tested interstate and received enthusiastic responses. A total of one workshop, one field day and two expert review sessions were held. Additional products. While drafting the Action Planner, it became evident that enticing horse owners to adopt sound environmental practices would be greatly enhanced if a "how to" guide was added to the package. The Management Guidelines, now accompanying the Action Planner, were developed as a result. Supplementary funding allowed the professional production of this document, which parallels the structure of the Action Planner and contains supporting colour photographs. In addition, one environmental horse poster and several (conference) Horse EMS banners were produced by the EPA and Horse SA. · Phase 3 – Adoption and outreach strategy developed. A well thought-through adoption and outreach strategy was considered an essential element of the project. Such a strategy would allow Horse SA, the custodian of future activities on environmental management by horse keepers, to target its efforts strategically with the goal of ultimately raising the (environmental) performance of the horse keeping community. The strategy was devised based on substantial consultation with both government and the horse keeping community. It now contains a prioritised sequence of strategic actions that provide the highest potential for on-ground environmental improvement results. One workshop and two expert review sessions were held. Additional products. The project team soon realised that the development of an adoption and outreach strategy would benefit from expert input. Additional NHT (Envirofund) funding enabled the team to engage Porter Novelli Pty Ltd in the development of the strategy. Porter Novelli used tools such as workshops and surveys to identify different target groups for an outreach program. In addition, the strategy is underpinned by the principles of community-based social marketing as promoted by Dr. Doug McKenzie-Mohr.
Implications	Although all initially specified project objectives and desired products were achieved, the overall project success truly lies in the successful application of the participatory development and management approaches used throughout the project. Environmental management for horse keepers won’t wind down after this project, but will carry on into the future, with the following new initiatives established: · community-of-practice forum; · adoption and outreach monitoring; · periodic updating of Action Planner and Management Guidelines; and · extension of training and research to other ‘lifestyle’ rural land managers, which often manage horses on their properties.
Publications	This project enabled the publication of the following free information resources (available from the Horse SA web site starting in August 2006): · Community-of-practice website (maintained by Horse SA), displaying: · The Action Planner for Horse Keepers - a tool to assist horse keeper assess and improve environmental management · Management Guidelines – to accompany the Action Planner for horse properties