Working for the Betterment of Horses

"Go with your strengths, and collaborate as effectively as possible." Those were the two baseline rules that Bill Moyer, DVM, head of the Department of Large Animal Medicine & Surgery at Texas A&M's College of Veterinary Medicine, and Bryan Johnson, PhD, head of the Department of Animal Science in Texas A&M's College of Agriculture and Life Sciences, agreed upon at the inception of the Link Equine Research Endowment. Texan Patsy Link left $8.3 million to the two programs in the mid-1990s, dictating in her will that the monies be used for equine research to improve the health and welfare of horses.

The two men decided that instead of the more traditional mode of funding individual projects, they would support programs on a recurring basis, according to their previous and assumed success and production. "We went with our strengths, and made attempts to collaborate as much as possible to increase the strengths of all involved," said Moyer. "To date, this has included other departments, colleges, and universities around the world. This approach, in our opinion, has been very successful."

"We also knew we could get quite a bit more bang for our buck because we have facilities in place to take these new resources (Link funding) and create a resource base of money and expertise," said Johnson. He further explained that the Link monies could also be used to bring in matching donations from several other research funding agencies.

The Link funds give the university a recurring source of monies, since the endowment invests the balance of the funds and allows the university to spend 5% of the total value of the endowment each year. Matching funds, if achieved, stretch the use of such dollars even further.

In trying to determine the best way to put the Link money to work for the betterment of horses, "We identified key areas in the horse industry that needed to be addressed, examined the necessary faculty and expertise, and created six programs," said Johnson. Those programs were epidemiology, colic/intestinal function, reproduction, the equine genome, nutrition, and infectious disease.

Each program has a coordinator who is responsible for oversight and for reporting results to a research advisory council set up by the university and the co-directors, Moyer and Johnson. This review process is necessary to ensure continued success. Thus, the "programs" are held up to significant scrutiny.

"Each program must show that they can use these Link resources to generate other resources," said Johnson. "The programs are not guaranteed money. Funding is based on performance and accountability."

"Success breeds success," added Moyer. "Our main mission is to produce quality research and enhance graduate education. We are also seeking ways to increase the corpus of the endowment. This program has already produced results and has had an influence on the horse world. That enhances our ability to raise money and raises our standing in the research community and the horse industry.

"Our goal is to get information to the end users--the horse owners and the veterinarians who serve them, and thus serve the intent of the will created by Ms. Link," concluded Moyer.

The cooperation between Texas A&M researchers and other universities has been "beyond what we expected," said Moyer. "The groups have worked together using expertise anywhere they can find it. The quality of science is outstanding! The future of these endeavors is only bounded by imagination."

For example, in the reproduction program alone there are collaborators from the University of Massachusetts, Colorado State University, University of Florida, University of California, University of Georgia, Louisiana State University, private equine practices in the United States, and France and Finland abroad.

"The other aspect that has happened is that of a blending of programs here at the university," he added, saying that epidemiologist Noah Cohen, AB, VMD, PhD, MDH, Dipl. ACVIM, might interact with several of the programs during projects. He said the blending of faculty and facilities improves efficiency, reduces duplication of efforts, and leads to new ideas. The ability to attract funding from organizations such as the Grayson-Jockey Club, American Quarter Horse Association, and other groups is thus enhanced. Moyer also noted that a percentage of the breakage (monies earned from gambling) from Texas racing is applied to equine research in the state of Texas. While the amount is variable, it is quite significant, thereby placing the state of Texas in a unique position regarding equine research.

Moyer and Johnson are excited about the future of equine research, not only at Texas A&M, but around the world. And that excitement was made possible in large part to a woman who wanted to better the world in which horses live.

Link Equine Reproduction Program

So much has been learned about stallion and mare reproduction in the last 20 years that the industry is not the same today as it was two decades ago. For horse breeders, this means the techniques that they learned just a few years back probably have been improved upon in the meantime, or new techniques might have made what they are using obsolete.

Plus, there are still frontiers to be crossed and progress to be made.

The Equine Reproductive Biology Program that is part of the Link research group at Texas A&M University concentrates on equine gametes--sperm and oocytes (eggs). This program is coordinated by Dickson Varner, DVM, MS, Dipl. ACT, Professor of Equine Reproduction in the College of Veterinary Medicine and Pin Oak Stud Chair of Stallion Reproductive Studies; and David Forrest, PhD, PAS, Dipl. ACAP, Professor of Animal Science, College of Agriculture and Life Sciences. It's supported by clinical investigators and basic scientists from many different departments within the university, as well as collaborations with institutions worldwide. The program is dedicated to integrating basic science with clinically applicable techniques that will benefit the horse industry in areas related to reproductive management of mares and stallions.

The oocyte group is led by Katrin Hinrichs, DVM, PhD, Dipl. ACT, Associate Professor of Veterinary Physiology and Pharmacology in the College of Veterinary Medicine. She said, "There has been little published (on oocyte maturation and fertilization), so everything we learn is new."

In fact, the first paper on the equine oocyte was published in 1982. Hinrichs said that the basic problem has been that while a lot of research has been done on cattle, sheep, pigs, and other livestock, there has been little general research in horse gamete physiology. And, she said, it is hard to "harvest" oocytes from mares.

"When we're harvesting oocytes, most are in a resting stage so they must be matured in an incubator," explained Hinrichs. This technique has taken time to perfect, but several things have been learned in the process. For example, the oocytes are better maintained at room temperature than in refrigeration during transport, and oocytes collected immediately from ovaries have a higher maturation rate compared to those removed from ovaries after storage.

However, only two foals have been produced by standard in-vitro fertilization techniques, and those foals were born more than 10 years ago! No one has been able to duplicate the feat.

Preliminary studies at Texas A&M revealed that oocyte maturation medium (the substance in which it is incubated) affects fertilization rates. Although fertilization rates in the lab were low, they dramatically improved (70%) when oocytes were matured in the laboratory and then placed in the oviduct of a mare which was inseminated.

"There is very little data on how to culture horse embryos in the lab," said Hinrichs. "The best incubator by far is the mare."

Cloning is closely related to this research. The research group has had four abstracts published on the early stages of equine cloning, which they say is harder in horses than in some other species. The applications are tremendous for equine cloning--with genetically identical horses, researchers could determine the effects of genetics vs. nutrition, disease, or drugs.

"We could use cloning to look at animals with OCD lesions to see if it is because of environment or genetics," said Hinrichs. "We could ask, 'Is that stallion infertile because he was given certain drugs, or because of genetics?' "

On the stallion reproduction side, many studies are underway relating to both ejaculated sperm and testicular function. Larry Johnson, PhD, Professor of Veterinary Anatomy in the College of Veterinary Medicine and co-group leader for testicular function research, said, "The Link reproduction program was a good marriage between basic and clinical science."

He explained that in the stallion, developing (prepubertal) testes have both active and inactive stages of spermatogenesis (the process of sperm production within the testis). "But we need to understand what genes are initiating spermatogenesis in the young colt."

Basically, the group wants to know what turns on spermatogenesis in the young horse, because this information might lead to treatments for abnormal spermatogenesis in adult horses. Thus, Nancy Ing, DVM, PhD, Associate Professor of Animal Science in the College of Agriculture and Life Sciences and co-group leader for testicular function research, is investigating the genes that are turned on and turned off in the developing testes of young colts, as well as the testes of both fertile and subfertile stallions.

Another area of this research is apoptosis, or programmed cell death, which is common in all species. However, the research by the Texas A&M group was the first to describe this event in the testes of horses.

Said Terry Blanchard, DVM, MS, Dipl. ACT, Professor of Equine Reproduction, College of Veterinary Medicine, "We are interested in the production and death rates of germ cells that eventually form sperm."

These events are affected by the age of stallions, and are also impacted by many factors that damage spermatogenesis. "Research has shown that in some infertile or subfertile horses, the initial stages of spermatogenesis had a similar number of sperm-forming cells to a normal horse, but in later stages, the cells were no longer there," said Blanchard.

Varner states that, "Apoptosis is one mechanism that can explain the disruption of spermatogenesis in subfertile stallions. We want to know what to do to return subfertile stallions to normal testicular function. With this research, in the future we may have a logical way to solve some subfertility problems in stallions."

Blanchard added that this research might help to show whether a stallion's fertility problem is genetic, and thus point to the proper treatment needed for that particular horse. "But we need the influence of basic science, and we need the funding to do the research."

Varner said that this group is also investigating a variety of other laboratory tests in an effort to predict stallion fertility better, as well as improved techniques for storage of semen in both cooled and frozen states, intracytoplasmic sperm injection (ICSI), and low-dose insemination.

Horse owners and veterinarians should look for more answers to breeding questions coming from this group in the near future.

Link Equine Genome Program

This is an area of research that--like the horse industry--has its own language that is hard for an outsider to understand. Be that as it may, breakthroughs in many areas of equine wellness and disease will come through a better understanding of the genetics of the horse.

The Texas group is working with the world-wide equine genome project that is attempting to create a gene map of the horse. Since other species--such as man and mice--have been totally mapped, the work on the horse is easier, but still not simple. Loren Skow, PhD, and Bhanu Chowdhary, BVSc, PhD, are the leaders on this project at Texas A&M.

"We can make genetic material (in the lab) act as if we were breeding horses," said Skow. "That's cheaper than producing a thousand head of horses!"

"Resolution" in the geneticist's language means the ability to detect the distance between two genes. "In the mapping families used in the genome project (with living horses), you can't detect a resolution of less than 5-10%. In ours (done in the lab), the resolution is 1%."

What that means is that they are better able to compare where a gene should be on the horse genome, using human or other completed genomes as their guides. However, they said that some traits are unique to the horse and are better studied in the horse. For example, coat color genetics can be studied using mice. But, if you are interested in what genes control a horse's response to a specific infectious disease, then you need to use horse genetics.

There are many things that can cause genetic disorders, from missing chromosome pieces to rearranged or defective genes.

"We want to know what genes are affected when a horse is exposed to a disease, and how different horse genes respond and control disease resistance," said Skow. "We want to learn the degree of genetic involvement in lameness. I've always been interested in soundness and performance genes, but they are harder to get a handle on because of the environmental influence."

In their cooperative work with the reproduction group, the genome group's research plays a role in identifying a genetic basis for infertility and subfertility. This work mostly was done with stallions in the past, but more studies now are being conducted with mares.

One of the eye-opening facts about genetics is that while the genome might have three billion bases (places where genes reside), the active genes only are a small part of the whole--one-half of one percent. The rest of the DNA contains non-functional genes. When comparing horse genes to other species, "equids' genes look like they were reshuffled," said Skow. "We'd like to know what happened to the equid genome (during evolution) and why they are rearranged."

Some of the answers that genetics has given to the horse industry already include being able to pinpoint genes related to the deadly SCID (severe combined immunodeficiency) in Arabian horses, HYPP (hyperkalemic periodic paralysis) in Quarter Horses, and overo lethal white syndrome in Paint horses. What is interesting is that these particular diseases are located in the same area on the equine genome as on the human genome.

"Once the horse and human genomes are aligned for traits shared, it will be easier to find responsible genes," said Skow.

One of the next steps in the genome project will be moving from a description of the normal horse genome to the abnormal. "How do we get that material? What problems will we face with confidentiality?" questioned Skow. "Some breeders don't want to allow an abnormal genetic horse to be studied. But we could probably do this with hair or blood and bank those samples.

"In four to five years, we hope to have enough material to study disease in horses," Skow continued. "With blood typing, if we saved serum there would be enough DNA to be able to pick up and amplify DNA and recover a gene of interest. We need more of that type of resources."

Link Equine Infectious Disease Program

A study done at Texas A&M University estimated that Texas' approximately one million horses directly generate more than $3 billion annually, and that medical expenses are second only to feed costs for horse owners. Respiratory disease is the most common cause of disease and death in Texas foals.

The coordinator of the infectious disease research program at Texas A&M is Ronald J. Martens, DVM, PhD. Throughout his career in private practice and working in university settings, Martens has had an interest in the pathogen Rhodococcus equi, which is the cause of most severe and deadly foal pneumonia world-wide.

The bacterium lives in the soil, so it is nearly impossible to eradicate. Research has shown that it is not possible, based on soil cultures for R. equi, to determine whether a farm is at increased risk of promoting disease caused by the bacterium. Research has also shown an association between the following factors and the presence of R. equi infections on horse breeding farms: Large-acreage farms, farms with large numbers of mares and foals, farms with high densities of transient mares and foals, and farms that house foals in stalls with dirt floors.

R. equi pneumonia is perplexing in foals, said Martens, because they get the infection early but don't show clinical signs until about three months of age. "That resembles TB (tuberculosis) in many ways," said Martens. "The R. equi grows insidiously over time. The foals clinically appear normal, then suddenly they have respiratory disease."

Other similarities between R. equi pneumonia in foals and TB in humans include the type and extent of lung damage, with significant damage occurring before signs of illness are readily apparent; the difficulty in establishing an early and accurate diagnosis so that specific therapy can be started; the need for long-term treatment; and the lack of an effective vaccine to prevent infection.

Texas A&M has collaborators in researching R. equi in Canada, Japan, Ireland, Argentina, Israel, and in the United States. Research by Dr. Shinji Taki in Japan has proven that some strains of R. equi are virulent (can cause disease), and other strains avirulent (don't cause disease). Learning to differentiate the strains is part of the research, as is learning how to assess factors that might be associated with an increased risk of a foal contracting R. equi pneumonia.

A controversial paper published by Martens and his colleagues suggested that foals are infected with R. equi in the first few days of life, which disputed the previously held belief that foals are infected at about one month of age when maternal antibodies are waning.

Martens pondered the question of why foals are exposed to R. equi but don't initially show clinical signs. He said that some research has suggested that the bacteria are intracellular inhabitants, which makes it difficult to impossible for macrophages (certain immune system cells) to get to the R. equi and kill it. Research has shown that R. equi can pass through an adult horse's GI tract unaffected, but when it is ingested by a foal, the bacterium is amplified.

The only known way to protect foals at this time is to vaccinate adult horses with R. equi and give hyperimmune plasma to foals (adult horses don't get R. equi pneumonia). In some challenge studies, the foals given the plasma were protected from disease. However, creating hyperimmune plasma and administering it to foals are expensive and labor-intensive.

"This is the only way we know to prevent the disease," said Martens. "But we don't know when the opportune time is to give the hyperimmune plasma. I think it might be at one to two days of age."

Martens also noted that R. equi can commonly cause problems in other parts of the foal's body, including synovitis (joint infections) or uveitis (eye infections).

Future research should help veterinarians pinpoint which foals are at greater risk of R. equi infection and discover new ways to protect foals from the illness caused by the bacterium.

Link Equine Epidemiology Program

"Epidemiology is a science that can be applied to just about any disease," said Cohen, epidemiology program coordinator. "It's the science of studying the pattern or distribution of disease and determinants (risk factors) of disease in populations or groups."

Cohen said that by nature, epidemiology is a collaborative effort, but it hasn't historically been used in studying horses. There are three principal thrusts at the moment, said Cohen, but the program is far from limited to those areas. The three are colic, performance horse medication, and foal pneumonia caused by R. equi (as discussed previously).

Epidemiology also is unique in that it allows university scientists to collaborate with private practitioners. For example, the colic surveys that have been done have depended on surveys of, and reports from, veterinarians in the field. That original colic survey (published in The Horse of May 1995) identified dietary and management practices that owners could use to help prevent colic. Epidemiology also helps put to rest theories or wives' tales that have no basis in science, for example that pelleted feeds caused increased colic. On the contrary, studies showed that some extruded pelleted feeds reduced the incidence of colic.

Epidemiologic research at Texas A&M done in conjunction with researchers in California showed that certain testing practices for the bleeder medication furosemide (Salix, formerly Lasix) were more accurate than others. That led to the proposal of guidelines that could be adopted by all racing jurisdictions for uniform, accurate regulation of furosemide.

Other research is geared toward determining risk factors for injury in racehorses. For example, Link-funded studies documented evidence that horses with abnormalities of the suspensory ligament that are detectable to veterinarians performing pre-race inspections were predisposed to a variety of injuries of the affected limb. One of the conclusions was that in Thoroughbred racehorses, pre-race veterinary exams are valuable tools in preventing injuries.

But on a cautious note, epidemiologic studies have shown that extrapolating results among racing jurisdictions can be misleading. In California, injured horses accumulated more high-speed exercise during the two-month period prior to injury than did control horses. In contrast, horses in Kentucky tended to be exercised less prior to injury. This points to the need for recognizing and accounting for regional differences in training practices and other conditions that predispose a racehorse to injury.

Another research focus will be epidemiologic studies of developmental orthopedic disease (DOD) in horses. The emphasis of these studies will be on management factors--particularly nutrition--predisposing horses to these conditions.

Cohen also is working with the state of Kentucky and private practitioners on an epidemiologic survey of problems in the spring of 2001, including pericarditis (heart problems), early fetal loss, and late-term abortion.

Link Gastrointestinal Research

Allen J. Roussel Jr., DVM, MS, Dipl. ACVIM, is head of the Links gastrointestinal (GI) research group. Following along on what Cohen reported on colic, other GI research continues with factors related to colic, and looks at gastric emptying and stomach ulcers. (In humans, heartburn often means the person has delayed gastric emptying, and some people with irritable bowel syndrome sometimes have accelerated gastric emptying, explained Roussel in comparison.) The research group also would like to look at antibiotic and prokinetic (drugs that make the bowel move contents faster) effects on the GI tract, and postoperative ileus (failure of contents to move through the digestive tract after surgery).

Roussel said that a retrospective study showed that ileus is the most serious problem following colic surgery, and the number one cause of death. It also showed that about 20% of surgical colic cases have post-operative ileus, with older horses more likely to have the complication than younger horses. Several other factors are known to increase ileus, such as length of surgery time, but on a positive note, it was found that enterotomy (opening the large intestine and removing the contents) reduced post-operative ileus by half.

Researchers are looking for a non-invasive means of measuring gastric emptying, such as with scintigraphy, blood samples, and breath analysis. They also have disproven several hypotheses in the process of working on the GI tract studies. For example, it long had been thought that an indwelling stomach tube delayed gastric emptying, which is not true.

One reason gastric emptying is of importance to the horse owner is that the horse cannot vomit, so if the stomach doesn't empty properly, it can rupture and kill the horse. Collaborating researchers in Florida and Scotland are trying to find a way to measure breath content and relate specific contents to how fast the stomach empties.

Roussel said that exciting research into ulcers is using sucrose absorption as a marker for gastric ulcers. "If the mucosa (lining the stomach) is not damaged, sugar should not be absorbed," he said. "If there is intact sucrose in blood or urine samples, then it leaked through damaged mucosa, indicating ulcers. This really looks promising as a relatively non-invasive way to diagnose ulcers."

Link Nutrition Program

"Most of our research involves understanding the athletic horse's physiology and nutrition," said Gary D. Potter, PhD, PAS, Dipl. ACAN, leader of the Link nutrition group. He is interested in strategic feeding and training of the equine athlete to extend the distance a horse can maintain his speed at the end of a race, and in learning more about bone quality.

Researchers already know that young horses in race training have a decrease in bone density about 60 days into training, said Potter, which is when bone is weakest during remodeling. That coincides with the time trainers ask for speed work. Thus, the young horse is at greater risk for injury.

During the bone demineralization period and subsequent remineralization, the horse loses mineral matter from the skeleton that must then be replaced. But nutritional requirements to provide for normal bone growth and the additional requirements for bone remodeling are poorly understood.

Thus, Potter and his group are working to define the nutritional requirements of the juvenile horse in race training. They want to define the effects of training on calcium, phosphorus, and magnesium absorption and retention, and refine existing recommendations for intake of those minerals in the diets of young horses in race training.

The second area of research is to determine the usefulness of several biochemical markers of bone activity in blood and urine to monitor changes in the bone of juvenile horses in race training.

Three papers from this project were presented at the 2001 Equine Nutrition and Physiology Society meeting. Potter reported the following:

"In summary, data from mineral balances, bone density, and biochemical markers of bone activity confirmed previous observations that young horses in training undergo a process of bone demineralization and subsequent remineralization in the first four to five months of training. This process resulted in decreased absorption of calcium, phosphorus, and magnesium during the demineralization phase.

"Even in the presence of extremely efficient retention of calcium (Ca) and phosphorus (P), the highest amounts of minerals consumed (150 mg Ca/kg/day; 60 mg P/kg/day) were not adequate to maximize retention of calcium and phosphorus during the bone demineralization phase. This was due not only to reduced absorption, but also to an increase in urinary excretion, particularly of calcium.

"Magnesium retention was maximal at a daily intake of 40 mg/kg/day. During the remineralization phase, calcium retention was maximized at a daily intake of approximately 125 mg/kg, while the highest daily intakes of phosphorus and magnesium (60 and 45 mg/kg, respectively) were not sufficient to maximize retention of those minerals. Thus, recommendations for mineral intakes for young horses in training must be adjusted.

"Further research is needed, but workable recommendations can be made from these data."

Potter reported that the next phase of research focuses on determining copper, zinc, and manganese requirements for juvenile horses in training, and continued effort to develop useful markers of bone modeling and remodeling. Other related work will attempt to clarify absorption of different chemical forms of minerals, management systems to enhance bone development, and the role of omega-3 fatty acids in the athletic horse.

"Success breeds success," commented Moyer. "Our main mission is to produce quality research and enhance graduate education."

About the Author

Kimberly S. Brown

Kimberly S. Brown was the Publisher/Editor of The Horse: Your Guide To Equine Health Care from June 2008 to March 2010, and she served in various positions at Blood-Horse Publications since 1980.

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