Genetic Disease - Who's At Risk?

Many of today's scientific advancements are based on genetic technology, and medicine is at the cutting edge of gene discovery. Equine medicine is no different. Using the very information code for life in genes, scientists are working to:

• Prevent devastating or career-ending diseases through informed breeding;

• Gain new knowledge on conditions that contribute to disease severity;

• Monitor efficacy of treatments;

• Assess fitness expression;

• Identify at-risk horses;

• Map the horse genome in order to improve the search for specific genes responsible for traits of interest; and

• Utilize gene chip technology to study disease expression.

The goal is to control, treat, and even eliminate equine genetic disorders. This article will discuss how genetic research is helping our horses live better, healthier lives.

What is a Genetic Disease?

All of the physical characteristics a horse expresses, and likely many of his mental ones, are controlled by his unique combination of genes--arrays of DNA sequence that code for specific combinations of amino acids, which form unique functional proteins. Many DNA sequence variations are fine; they just give rise to the broad spectrum of colors, sizes, and other characteristics we see in the horse population. Some variations, however, cause problems. These might range from a genetic predisposition toward a particular limb conformation to overo lethal white syndrome, which is 100% fatal.

And you can't currently cure a genetic disease. You can treat the clinical signs, but you can't change the horse's DNA to completely get rid of the problem. However, we can identify many genetic diseases via DNA testing and avoid spreading the diseases by removing affected individuals from breeding programs. Thus, much of the genetic research today is focused on identifying the gene mutations that cause particular diseases and developing tests for them. Other researchers focus on ways to improve treatment and management of affected horses so they can live more comfortable, productive lives.

Mapping the Equine Genome

"A genome map is like a road map with a set of landmarks along the chromosomes that lets a person know where on the road they are," says James D. Murray, PhD, professor of animal science at the University of California, Davis.

"Another way of looking at it is if you are looking for a house you have never been to before, you start your search by finding out where it is close to (state, town, suburb, block, etc.)," he explains. "A genome map is used in the same way. By doing a linkage mapping study of a trait we are interested in--say, a coat color in a set of families derived from stallions that produce offspring of the trait we are interested in--we can determine which highway (chromosome) the trait is on and place the trait between two markers. At that point we know the trait is between marker A and marker B, and we can then target the region in between for more markers, to get closer to the genes there that might cause the trait. In this way we can zero in and eventually identify the gene and the mutation responsible for the trait."

This same method is used for locating the defective genes that cause disease.

Dozens of laboratories and scores of researchers are engaged in this project. At this point, researchers are developing the tools needed by scientists to help them locate the genes responsible for traits. "The tools include a good linkage map and physical map to provide a source of markers whose locations are known on the map," Murray says, "and a good comparative map so that we can link from the horse genome map (which is incomplete) to the human and mouse maps (which are complete) in order to help us find what genes are present in a region of interest."

In terms of disease control, after locating the gene(s) that cause a disorder, researchers can then develop diagnostic tests to determine whether horses are clear of, carriers of, or affected with a specific disease, and breeders can use that information to select away from a disease.

The Five Least-Wanted List

The names of several genetic diseases strike fear into the hearts of horse breeders and owners. Researchers have identified the responsible gene mutations and modes of inheritance for the following five diseases:

Glycogen branching enzyme deficiency (GBED) causes abortion, stillbirths, and foal deaths in affected Quarter Horses. "All foals with GBED studied to date have died or been euthanized due to weakness," says Stephanie Valberg, DVM, PhD, Dipl. ACVIM, professor of large animal medicine and director of the Equine Center at the University of Minnesota, who is studying several genetic disorders.

The disease is inherited as an autosomal (non-sex-linked) recessive trait, meaning that an affected horse received a copy of the defective gene from each parent

Overo lethal white syndrome (OLWS) is a fatal autosomal recessive disease in which foals cannot pass feces. The disease is seen in all-white or primarily white Paint foals. Horses carrying the OLWS gene usually have a frame overo coat color pattern, although occasionally carriers can be solid-colored Paint broodstock or horses with other white color patterns. Carriers are found rarely among miniature horses, half-Arabians, Thoroughbreds, and Quarter Horses that have associated white coat color patterns. The gene mutation has been identified, and owners can test breeding stock to avoid producing affected foals. There is no means to save affected foals.

Hyperkalemic periodic paralysis (HYPP) is a muscular disorder resulting in sudden episodes of uncontrollable muscle twitching, paralysis, and sometimes collapse or death. It is seen in Quarter Horse bloodlines (and some crossbreds tracing back to affected Quarter Horses), and it is inherited as an autosomal dominant trait (any horse with even one copy of the defective gene will express the disease).

Now that a DNA test exists for identifying HYPP horses, research efforts are directed toward disease management. "We have performed research as to why some horses are more symptomatic than others, in spite of having the identical mutation," says Sharon J. Spier, DVM, PhD, Dipl. ACVIM, Professor in the Department of Medicine and Epidemiology at the University of California, Davis. Investigators found horses that are more severely affected have an increased proportion of abnormal sodium channels in their muscle. "Veterinarians have observed that heavy sedation, such as occurs with modern dentistry and general anesthesia, precipitates attacks and even death," says Spier. "We continue to investigate other muscular and neuromuscular disorders that can cause similar signs or exercise intolerance."

Management, says Jeremy Powell, DVM, extension veterinarian at the University of Arkansas, should include dietary changes to decrease potassium levels. "Feeds with relatively high levels of potassium include alfalfa hay, molasses, and wheat bran," he says. "Feeds such as cereal grains (corn, oats, barley), beet pulp, and vegetable oils contain lower levels of potassium."

Avoid electrolyte supplements containing potassium and analyze forages as potassium concentrations vary with maturity, Spier adds. Another management tool, says Powell, is to administer acetazolamide; this medication reduces episodes.

Severe combined immunodeficiency (SCID) seen in Arabian horses is an autosomal recessive disorder in which affected horses can't produce immune responses sufficient for protection against infectious diseases. Explains Lance Perryman, DVM, PhD, Dean of the College of Veterinary Medicine and Biomedical Sciences at Colorado State Univerity, "Affected foals don't produce antibodies after infection or immunization, and rarely live beyond five months of age unless the condition is corrected through bone marrow transplant."

Perryman collaborated with other scientists to locate the defective gene, and since 1997 breeders have been able to use DNA testing to identify carrier, clear, and affected horses.

Junctional epidermolysis bullosa (JEB) is another autosomal recessive disease that causes skin lesions over pressure points of the body in newborn Belgian foals and results in large areas of skin loss. JEB is also seen in American Cream draft horses, Breton draft horses, and Comtois draft horses. The condition is untreatable, and foals die within 24 hours to 14 days after birth. A form of the disease also occurs in American Saddlebred newborns, although the mutation responsible is different from that in draft horses.

John D. Baird, BVSc, PhD, professor of large animal medicine in the department of clinical studies at Ontario Veterinary College, was part of the team that developed the test for identifying carriers of the mutation responsible for JEB in Belgian draft horses.

Still Just Suspects

There are quite a few other disorders that horse people suspect of having a genetic origin, but the specific mutations causing the diseases have not yet been unequivocally identified.

"Until you've got the gene, you just don't know for sure," says Valberg. "Research goals are to determine the underlying genetic defects, their mechanism of causing clinical signs, and the best way to alleviate those signs and make horses as comfortable and physically sound as possible," she says.

Following are seven diseases for which researchers are working to understand the genetics involved.

Polysaccharide storage myopathy (PSSM) causes tying-up in Quarter Horses, Paints, and Appaloosas; there is also a form that presents as muscle soreness, weakness, and tying-up in draft horses, draft crossbreds, and warmbloods. Horses affected by PSSM forms are managed by providing regular turnout, a gradually increasing exercise schedule that becomes a regular routine, and a low-starch/high-fat diet.

"We've done some pedigree analysis that suggests a dominant mode of inheritance for this disease," reports Valberg. Horses with PSSM should not be bred, as there is a 50% chance their offspring will be affected, she adds.

Degenerative suspensory ligament desmitis (DSLD) is a chronic, untreatable connective tissue disease that causes pain and abnormalities, primarily in limb tissues and to a lesser degree in other tissues of the body. Initially identified in Peruvian Pasos, DSLD has also been diagnosed in Thoroughbreds, Quarter Horses, warmbloods, Standardbreds, Arabians, and draft horses.

Researchers at the Universities of Kentucky and Georgia have identified eight chromosomal regions where there are differences in the marker genotypes between affected and unaffected groups of horses, reports Jeanette L. Mero, DVM, associate veterinarian at Starland Veterinary Services in Romulus, N.Y., and an activist in promoting DSLD recognition and research. More work will be done to localize the specific gene on one of these chromosomes. Additionally, various clinicians and researchers are hoping to develop diagnostics for subclinical early cases and to further clinical understanding of DSLD.

Therapies including therapeutic shoeing, medications, surgeries, bone marrow injections, and extracorporeal shock wave therapy have been ineffective for curing DSLD or relieving clinical signs, Mero reports. 

Recurrent exertional rhabdomyolysis (RER) is a form of tying-up in Thoroughbreds. Researchers have been working to identify a genetic mutation causing this disease, but have not yet located it.

RER-affected horses are managed by housing them in a quiet area of the barn, avoiding exciting situations, providing turnout, feeding a specially formulated high-fat/low-starch diet, providing warm-up exercises, and avoiding routines that promote tying-up. "In Thoroughbred racehorses," says Valberg, "this means avoid fighting to hold a horse back during galloping. In Standardbreds, limit jogging to 20 minutes per session. In riding horses, acclimate horses to new environments, horse shows, and speeds achieved during steeplechase phase of three-day events."

The disease "may be inherited as an autosomal dominant trait with variable expression," concludes a study published in 1999 in the American Journal of Veterinary Research. In other words, the appearance of clinical signs in RER-affected horses might be influenced by management--diet and exercise level/schedule. More work on the inheritance of this disease is forthcoming.

Hyperelastosis cutis (HC) or hereditary equine regional dermal asthenia (HERDA) is a rare condition occurring in some Quarter Horse bloodlines. In affected horses, the deep layers of skin are weak and thin, thus separating and tearing easily. The disorder is passed to offspring via recessive inheritance, which means that if both apparently normal parents carry the HC gene, the offspring has a 50% chance of carrying the disease without clinical signs and a 25% chance of expressing the disease. If only one apparently normal parent carries the defective gene, the foal has a 50% chance of carrying it without clinical signs, but will not express the disease.

While Cornell University's Nena Winand, DVM, PhD, searches for the defective gene, Ann Rashmir-Raven, DVM, MS, Dipl. ACVS, associate professor in the department of clinical sciences at Mississippi State University, is comparing skin biopsies, thermograpy, and biochemical studies of normal and affected foals to evaluate differences between the two.

There is no cure. "In most cases, horses are euthanatized or donated for research by the time they are four years old," Rashmir-Raven states. Occasionally, heavily managed horses with mild cases can be ridable. For them, Rashmir-Raven recommends dietary changes to include increased amounts of copper, vitamin C, calcium, and phosphorus, and good protein levels, along with sun restriction and very careful selection of saddles and pads to prevent skin rubbing and subsequent open wounds.

Laryngeal hemiplegia (roaring) is one-sided paralysis of the larynx, in which horses suffer from partial to nearly complete collapse of the airway during exercise and generate a subsequent loud "roaring" noise. In searching for the gene mutation, Elizabeth Santschi, DVM, Dipl. ACVS, clinical associate professor of large animal surgery at the University of Wisconsin, looked at mouse data to see if there was a similar gene mutation (there wasn't). She is now screening genomic material for a gene sequence difference between affected and unaffected horses. Treatment is surgical, with varying degrees of success.

Colic-induced intestinal injury refers to colic-caused damage sustained by the intestine (twisting and loss of blood supply), the most common reason for colic fatalities.

For unknown reasons, some horses' intestines do a better job of repairing themselves than those in other horses. Using genetic sequencing (figuring out the genetic code), researchers at North Carolina State University (NCSU) and Virginia-Maryland Regional College of Veterinary Medicine are searching for the genes involved in intestinal repair. With that information, they hope to learn how to turn on or off genes that aid or hinder intestinal repair.

Reports NCSU researcher Anthony Blikslager, DVM, PhD (GI physiology, associate professor in equine surgery in the Department of Clinical Sciences at North Carolina State University, "We have sequenced a group of genes involved in regulating the connection between cells that line the intestinal tract; these cells stick together and re-seal damaged junctions in between the cells that line the intestine."

But in some horses, these cells contain reduced amounts of reparative proteins. "We want to understand the cause of these reduced amounts and then figure out a way to boost their production," he adds.

Osteochondritis dissecans (OCD) is a joint cartilage disorder. Studies have found differences in the gene expression of local hormones within cartilage associated with the disease process of OCD. "The premise has been that an underlying molecular abnormality leads to matrix and cell signaling changes in OCD cartilage," says Stacy A. Semevolos, DVM, Dipl. ACVS, assistant professor of large animal surgery at Oregon State University. "These changes ultimately result in areas of thickened cartilage that is weaker than its normal counterparts, making it susceptible to biomechanical damage. Determining normal gene expression lays the groundwork for understanding what happens in disease processes like OCD and osteoarthritis."

She is also investigating how growth hormones circulating in the bloodstream affect local hormonal signaling within OCD and normal cartilage. "This project will give us further clues on how OCD cartilage may respond differently than normal cartilage during rapid growth spurts," she says.

Arthroscopy is the treatment of choice for many OCD lesions, Semevolos states. "Most horses require six weeks (hock OCD) to six to 12 months (stifle OCD) of down-time after surgery," Semevolos adds. "Gradually increasing increments of hand-walking and controlled exercise are part of post-surgical care. Conservative treatment--decreasing the nutritional plane (grass hay only, reducing or eliminating grain and alfalfa from diet), balanced nutrients in the diet, decreased exercise level, and hyaluronic acid joint injections by a veterinarian--is recommended for young horses with mild OCD lesions."

New Technologies

The way scientists study these diseases and others is continually evolving; one exciting development is the gene chip.

The next wave of modern medicine, the gene chip is a 2.5-inch x 1.5-inch cartridge with a postage-stamp-sized glass window containing some 1.3 million DNA probes representing 3,000 unique equine genes. Drop a DNA blood sample on the glass wafer, and it gives information about the individual's gene expression (phenotype).

Alicia Bertone, DVM, PhD, Dipl. ACVS, Director of the Comparative Orthopedic Research Laboratories at The Ohio State University and developer of the chip, explains, "At any given moment, you express about 5% of your DNA make-up; that's your expression phenotype, and those phenotypes will change over your lifespan."

For example, foals express many growth and development genes not seen in adults. An overweight, under-exercised horse expresses a particular phenotype, but if conditioned will produce a different phenotype--muscle enzymes that represent fit muscles, genes that are turned on to make bones denser, etc. A horse at risk for--or in the very early stages of developing--cancer, osteochondrosis dissecans, colic, or other diseases will have a gene expression profile outside the norm. A horse that has an active infection, such as herpes or EPM (as opposed to just being exposed to a virus and raising an antibody response) will express genes indicative of active infection.

Because the gene chip can view thousands of genes at once, looking at a horse's phenotype will help us make earlier and more accurate diagnoses, and providing an objective way to determine fitness.

"Most of this is still in the future," says Bertone. "We've looked at five or six key diseases in horses--that data is not published yet--and have genes on the chip that represent acute herpesvirus infection and many other infectious diseases. We're hitting the most obvious ones first. Clearly this is something with tremendous potential."

Breeders Make the Difference

With new technologies promising to expand the range of veterinary advances, along with traditional clinical trials to evaluate treatments and identify contributing causes, the tools for breeding healthier, sounder horses increase every year--but breeders must utilize those tools.

Although it's been 13 years since a DNA test for identifying HYPP in carrier and affected horses was developed, the incidence of H/H, or severely affected horses, continues to rise, meaning that people are still breeding affected horses to affected horses, says Spier. "The market has not resolved the problem, as was predicted when the test became available in 1992," she states. "Gene frequency is increasing in the breed rather than decreasing. With all of the education, publications, web sites, brochures, and presentations by veterinarians regarding this disease, it is disappointing that figures remain what they are. I would hope that breeders would have a longer-term vision for what is good, sound breeding for the health of the horse.

"The same could be said for breeding horses with poor hoof conformation or soundness issues," she adds. "We can manage these for the short term (and shortened equine career), but should breeders continue to promote these horses as the best of the breed? I believe the problems that occurred in the Quarter Horse breed and other breeds over the past 20 years have been due to the over-specialization and over-use of inbreeding to produce 'carbon copy' images of what is considered ideal for that event."

Spier says there should be more emphasis placed on hoof and leg conformation, temperament, and soundness rather than bloodlines when breeding. "I wish breeders would take more chances and breed horses outside of their specialty to see if we can maintain what makes the Quarter Horse so popular--a healthy, versatile horse."

Santschi finds that attitudes about breeding decisions are sometimes ambivalent. "They all think genetic disease should be as limited as possible," she says, "but some get weaker-hearted when it involves their horses. It's also about money: The more valuable breeds are, the less interested some breeders are in talking about genetic defects."

It's hard to determine the percentage of breeders who are willing to make the tough decisions when it comes to the overall well-being of the breed versus the breeding career of their prized stallion or mare--and the financial well-being of the breeding operation. Obviously, although some breeders are reluctant to embrace the big picture, others are willing to move forward, particularly when it involves diseases that result in foal death or young horse loss.

Along those lines, Baird notes, "The JEB test is now accepted as a useful tool by draft horse breeders, as it prevents the financial losses associated with the birth of a foal with a lethal genetic disease."

Additionally, organizations including The American Quarter Horse Association, Morris Animal Foundation, Grayson Jockey Club Research Foundation, Oak Tree Racing Foundation, and University of Minnesota Equine Center contribute to and support genetic research. "Research into genetic diseases in horses is highly dependent on the support of breeders through research funding and cooperation with veterinarians to identify and allow affected and unaffected horses to be tested," states Valberg. "The cost of research is such that it would be a rare instance that herds of horses of sufficient size could be kept by universities to study these diseases.

"Progress," she adds, "will depend on the degree to which breeders work with the researchers and breed associations to develop these genetic tests." And to the degree that breeders utilize them.

Take-Home Message

There have been many advances in genetic research, some of which have resulted in genetic tests to help breeders avoid passing on disease-causing genes. However, there is still a lot to learn, and once learned, breeders must put the tools into practice in order to breed healthier, happier horses.

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Unlocking Genetic Secrets: How Research Works

How do scientists uncover the secrets of genetic disease? The search begins by looking for evidence that suggests the disease is hereditary, says Stephanie Valberg, DVM, PhD, Dipl. ACVIM, professor of large animal medicine and Director of the University of Minnesota Equine Center. One indicator is if the disease is common in some families or lines, but not in others.

From there, approaches vary. Valberg describes one process:

• Develop a clinical diagnostic test to identify normal or affected individuals, then work with breeders and owners to test as many related individuals as possible in multigenerational families, classifying them as normal or affected. This could suggest the mode of inheritance--whether offspring need to inherit the defective gene from only one parent or from both to be affected, and whether the gene is passed from either or both sexes.

• Collect blood or tissue samples from all family members to isolate DNA and perform a comparative review of scientific literature to find any similar diseases described in other species. "If these comparative diseases have known genetic causes, this suggests a possible 'candidate' gene for the equine disease. An assay can be done to see if the gene is abnormal, then the gene would be sequenced to see if there is a mutation in the genetic code. If found, the mutation would be checked to ensure that it is consistently present in all the affected horses and consistently absent in the normal horses."

• If no candidate gene is suggested from other animal species, perform a genetic linkage analysis to look at the alleles (individual characteristics) of specific markers spread throughout all the equine chromosomes. "We look to see if alleles of key markers are consistently associated with the disease and not associated in normal horses. This provides an approximate location of the disease gene on the equine chromosomes."

• Study the human genome map, examine corresponding chromosomes in humans, and see if there are defective genes that could produce the clinical signs seen in the affected horses. "These genes are then selected and the genetic code sequenced to find a mutation. The equine genome map is now developed to the point that linkage analysis can be performed in horses, and several groups are using this approach for PSSM, RER, and HERDA."

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Dominant, Recessive, What's the Difference?

When you hear that a trait is dominant or recessive, does that confuse you? You're not alone. But it's not too complicated, and knowing how these diseases are passed can help you make breeding decisions that sidestep the problems entirely.

Many traits are controlled by a pair of genes; each single gene can be either dominant or recessive. If a dominant gene is present, it will be expressed (i.e., you will see that characteristic) regardless of whether the other gene is dominant or recessive. A recessive gene, however, is not expressed unless both members of the gene pair are recessive. The gene types are normally noted with capital letters for dominant traits and lowercase letters for the recessive version.

For example, let's say a brown eye color (written as "B") is dominant over blue (written as "b"). A horse would have brown eyes if he had either a BB (homozygous dominant, or having two copies of the dominant gene) or Bb (heterozygous, or having one dominant and one recessive gene) genotype, but he would have blue eyes if both copies of that coat color gene were recessive (bb, homozygous recessive).

The mating diagrams below show the outcomes of various crosses with recessive and dominant traits.

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RECESSIVE TRAITS
(example: Overo lethal white syndrome, or OLWS)  

MATING TWO CARRIERS (LlxLl)   L l L LL Ll l Ll ll 25% normal (LL) 50% chance of carriers (Ll) 25% chance of an affected foal (ll)

MATING A CARRIER TO A NORMAL HORSE (LlxLL)   L l L LL Ll L LL Ll 50% chance of a normal foal (LL) 50% chance of carriers (Ll)

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DOMINANT TRAITS
(example: Hyperkalemic periodic paralysis, or HYPP)  

MATING AFFECTED HORSES (NHxNH)   N H N NN NH H NH  HH 25% normal (NN) 50% chance affected (NH) 25% chance of a severely affected foal (HH)

MATING AN AFFECTED HORSE TO A NORMAL ONE (NHxNN)   N H N NN NH N NN NH 50% chance of a normal foal (NN) 50% chance affected (NH)

It was two years ago--in April of 2004--that this magazine published the announcement that the Poco Bueno Quarter Horse sire line had been identified as carrying the recessive gene that causes hyperelastosis cutis (HC, also known as hereditary equine regional dermal asthenia or HERDA). This is an affliction involving the skin that very often carries with it a death sentence for the affected horse. During the past two years, a good deal more has been learned about HC, but at the same time, it remains an elusive entity as researchers seek to pinpoint its genetic location. This would be a major step in developing a DNA test that would identify carriers. (The gene is characterized as being autosomal recessive, which means that both parents must pass it on for their offspring to be affected.)

University of California-Davis researchers have reported progress in their genetic investigations of this disease, but have not yet identified the specific gene. They have identified the chromosome on which the HC gene is located and have narrowed the field as to which gene might be involved, but have not reported identifying the gene itself.

A new urine test can be administered to foals to determine if they are affected, but there is not yet a DNA test to determine which horses are carriers and can pass the condition to offspring. And, as knowledge expands, there is a growing concern that HC might involve more than just the skin of an affected horse.

What is HC?

When a horse has HC, there is a lack of cohesion within the dermis, the deep layer of skin, due to a collagen defect. Collagen is the protein constituent of bone, cartilage, tendon, and other connective tissue that in a manner of speaking, plays the role of glue in keeping the layers of skin together. However, when a horse has HC, the glue is inferior and the layers separate. When the horse is ridden or suffers trauma to the skin, the outer layer often splits or separates from the inner layer. It can also tear off completely. When the wound heals, there very often is a disfiguring scar. New damaged areas arise continuously, often without obvious trauma. Some affected horses have lived to old age, but for the most part, they have been the recipients of tender, loving care that prevented skin trauma. Sun damage is also a concern, and affected horses must be kept out of direct sunlight as much as possible. Ann Rashmir, DVM, MS, Dipl. ACVS, a researcher who heads up the HC research program at Mississippi State University, says that the affliction continues to strike with varying degrees of severity. With some, it is so bad that portions of the horse's skin literally fall off its body. With others, the horse is affected only mildly and can often be ridden in the performance ring if handled correctly, she says. However, one thing has not changed: There still is no cure for HC. For most horses, it is a death sentence, with the average life span being in the two- to four-year range. Much has been learned about handling affected horses, Rashmir says, with overall quality care and proper nutrition playing valuable roles. There normally are 24 affected horses in the herd at MSU that are being continually studied in an effort to learn more about HC and how to deal with affected horses. Unfortunately, says Rashmir, the often-virulent nature of the disease means that there is a fairly rapid turnover of animals in the herd. For example, two affected yearlings that had been donated didn't reach their 2-year-old years before progress of the disease and its side effects made it necessary that they be euthanatized. When a DNA test is developed, it could reveal if a sire or dam carry the recessive HC gene. The genetic law of averages indicates that if both parents are carriers, 50% percent of the offspring will be carriers, 25% will have HC, and 25% will be normal. When a horse with HC is crossed with a carrier, the odds go up. Under that scenario, the genetic law of averages indicates that 50% of the offspring will be carriers, and 50% will have the disease. When a horse with HC is crossed with a normal horse, 100% of the offspring will be carriers. Thus, one could reason, if the DNA test reveals that both stallion and mare have the HC gene, the wise thing is to avoid breeding them to each other. As a practical matter, however, that will not happen. Some of the HC gene carriers are talented stallions and mares in the cutting and performance horse arenas. Just being a carrier has no known negative effect on the horse's ability to perform. One breeder, who chooses to remain anonymous, says that he has bred three of his mares to a known carrier stallion that has a reputation for siring winning cutting horses. The mare owner does not know whether his mares are carriers, but says he was willing to "roll the dice," gambling that they are not. Even if they were known carriers, he says, he perhaps would have bred them to the stallion anyway, gambling that the offspring from the two carriers would be in the 50% group that were carriers or the 25% group that were normal. If one or more of the offspring from those matings fell within the 25% that were HC-affected, he would simply accept his loss, he says. New Test for Foals One of the more significant accomplishments to date at MSU involves development of the urine test mentioned earlier that can identify HC-affected horses when they are foals. There are changes in the horse's urine similar to what is seen in humans with Ehlers-Danlos type VI. (In this disease, tissue fragility and easy bruising are seen.) The prime researchers involved in developing the test were Rashmir; Cyprianna Swiderski, DVM, PhD, assistant professor in the department of clinical sciences at Mississippi State; and Marzia Pasquali, PhD, associate professor of pathology and director of the Biochemical Genetics and Supplemental Newborn Screen at ARUP Laboratories at the University of Utah. The test, says Rashmir, leaves some questions unanswered, even though it identifies affected foals. "It does tell you that this is a horse you will have to worry about because it has HC," she says, "but it doesn't tell you how seriously the horse is affected. You don't know whether it will turn out to be a mild case or one that is so severe that it results in the horse being unusable."

HC/HERDA UPDATE: OWNERS OF CARRIER STALLIONS NOTIFIED Approximately 100 Quarter Horse stallion owners have received, or will receive, a telephone message from Ann Rashmir, DVM, MS, Dipl. ACVS, associate professor of surgery and head of the Hyperelastosis Cutis (HC) Research Program at Mississippi State University (MSU), that contains unwelcome news. The message is that the stallion is a carrier of the recessive gene that causes HC, also known as hereditary equine regional dermal asthenia (HERDA). In dramatic cases, the skin can split along the back and even roll down the sides, with the horse literally being skinned alive. Generally speaking, she says, the average lifespan for an HC horse is two to four years. Here is a horse showing classic signs of HC/HERDA on the body. The hock of a horse with HC/HERDA. Skin lesions on an HC-afflicted research horse at MSU. The lesions rarely heal without disfiguring scars.  Rashmir shows how easily the layers of the skin separate on a horse with HC. The back and side of a horse with HC. Cindy Lyles of Haslet, Texas, bred her mare and did an embryo transfer. When flushed, the mare produced two viable embryos and each was implanted in a surrogate mare. Two foals were born (one is shown above). Both of them demonstrated outward signs of HC early in their lives. Further analyses, including biopsies, proved they were afflicted. Lyles has donated the two colts to the HC research herd at Mississippi State University. TOP SEVEN IMAGES: COURTESY ANN RASHMIR, DVM, MS, DIPL. ACVS. BOTTOM TWO IMAGES (PAINTS): COURTESY CINDY LYLES

More Than Skin Deep?

There are other unanswered questions concerning HC. A number of stallions with HC are cryptorchids, Rashmir says. Is it merely the result of continued inbreeding, or did HC play a role?

A number of HC foals also suffer what sometimes develops into a life-threatening infection. This appears to be happening with too much frequency to be coincidence, she says.

Does this mean that HC horses are different from other horses in more ways than just defective collagen involving the skin? Nena Winand, PhD, a geneticist at Cornell University who has been researching HC, thinks that might be the case. "This is not just a skin disease," she says. "There is an impact on other tissues that needs to be studied."

Winand says, for example, that she has seen evidence that HC might have an orthopedic impact. Her investigation into that possibility is continuing.

There also is the question of whether there is a connection between vision problems and HC. MSU researchers are working on this and suspect that there might be, Rashmir says, but as yet do not have definitive answers.

Interestingly, the two researchers who authored the first paper on HC in 1978 also mentioned that one of the two affected horses studied had a reported vision problem. The two researchers were Dana J. Lerner, DVM, MS, an equine veterinarian in Monticello, Ill., and Malcolm McCracken, DVM, PhD, a professor in the College of Veterinary Medicine at the University of Tennessee.

The two horses featured in that first paper were brought to the clinic at the University of Illinois in 1976 and ultimately were diagnosed as being afflicted with HC, according to the researchers' published report in the Journal of Equine Medicine and Surgery. In the report, they described what had been found in a 2-year-old Quarter Horse colt and a 5-year-old Quarter Horse mare. The researchers suspected that genetics might be involved because both horses had the same grandsire. The grandsire was not identified.

What the two researchers reported after examining the two affected horses would be repeated over and over in the future as more and more cases came to light. Here, in part, is what they had to say about the colt: "A black colt was presented in late February with an 18-month history of abnormal skin. The horse had recently been put into a training program when several abnormal areas in the saddle path appeared to be unusually sensitive. Previous treatment for this had included oral griseofulvin, which was ineffective. Skin lesions were present on the left foreleg below the elbow and on the left lateral thorax at the level of the 18th rib. These areas appeared dry and were hairless. They varied in diameter between 4 and 6 centimeters. Additional skin lesions were noted over the back, between the 13th and 18th ribs (two in number) and one on the right rump over the gluteal musculature. Each of the latter areas was very hyper-elastic. With tension, the center of each lesion was readily elevated 3 to 5 centimeters off the body. No resentment was evidenced by the animal. Immediately after the tension was released the skin returned to the original site. Extreme thinness was readily appreciated..."

With that horse, the researchers reported, there was no evidence of abnormalities within the musco-skeletal or ocular systems. However, with the second horse examined-the 5-year-old mare--the owner had reported a vision problem.

The concern among horse owners and researchers alike is that the HC problem is continuing to grow because of continued inbreeding that concentrates the gene pool. Horses carrying Poco Bueno blood, in many cases, are prized for their athletic ability and, in the minds of some breeders, if a little Poco Bueno blood is good, a lot is better.

There has been some speculation that many of the cutting horses carrying Poco Bueno blood have skin that is more sensitive and flexible and, as a result, are more apt to be affected than their counterparts who are not cutting horses.

As of a couple of months ago, according to Rashmir, the number of horses positively identified as HC carriers stood at 465, an increase of about 300 over the known number of carriers two years ago.

There can be little doubt that the 465 horses identified as HC carriers represent the tip of the iceberg. So, one might wonder, how will this all play out? Will a definitive DNA test in the future result in fewer carriers being mated to carriers, or will breeders knowingly continue to "roll the dice?" Where does responsibility lie in identifying carriers? Many of the horses offered for sale at the prestigious auctions during the NCHA Cutting Horse Futurity have the potential to be carriers. Are sale companies responsible for alerting the public as to a horse's HC status?

No, says Rashmir to the last question. "A sales company's job is to sell horses, not guarantee their health status. It comes down to the owners. We can hope that someone will guarantee a horse to be a not a carrier or not affected and, as a result, will get more money for it. That would encourage others to test their horses (when a DNA test for carriers becomes available) or test the foals with the urine test to guarantee that they are not affected."

At least one case involving a breeding that resulted in a pair of HC foals via embryo transfer has been in the court system, and the result of that suit might also have a bearing on how the industry handles disclosure of HC carrier status.

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A cryptorchid, also called a ridgling, is a male horse in which one or both testicles do not descend into the scrotum. In the developing fetus, the testicles are formed within the abdomen. As the fetus nears term, the inguinal rings and inguinal canal (passage through which the testicles descend) expand to allow for descent of the testicles into the scrotum. This canal contains blood vessels, nerves, and the cremaster muscle, which raises or lowers the testes in response to temperature change or threat of injury.

Based on fetal dissection studies, descent of the testicles is usually completed during the last 30 days of gestation. Testicles in the fetus are quite large, perhaps due to hormonal influences from the dam. They become smaller as the fetus matures. If they stay too large, they might not be able to come down through the inguinal canal.

If migration down into the scrotum is inhibited (if the canal fails to expand or if the testicle is too large to pass through), it is trapped within the abdomen. In these instances, the horse is a true abdominal cryptorchid; the testicle failed to enter the inguinal canal before closure of the internal inguinal ring (where the canal meets the abdominal wall). If it gets trapped somewhere below this ring but above the scrotum, the horse is an inguinal cryptorchid.

One or both testicles might remain high in the flank, stuck in the inguinal canal, and the horse is called a ridgling or high flanker. If the testicle is in the abdomen or very high in the flank, it makes the horse impossible to geld by routine castration. The undescended testicle must be located and removed via abdominal exploratory surgery. Horses with a retained testicle at the external inguinal ring (where the canal meets the scrotum) are called low flankers, and these are easier to remove.

Tom Yarbrough, DVM, PhD, a veterinary surgeon practicing in northern California, explains that a small structure (a cord called the gubernaculum testes) is an important player in the descent of the testicles. In a normally descended testicle, this little cord becomes a scrotal ligament, anchoring the testicle in the region of the scrotum--although it can still be pulled up closer to the body for warmth when necessary. If this structure is weak or absent (an inherited weakness), one or both testicles might remain above the scrotum or up in the abdomen.

Incomplete Gelding

Sometimes a cryptorchid is gelded, removing the one descended testicle and leaving the other. If only one testicle is found, some people assume the horse is a monorchid (a rare instance in which the horse only has one testicle) and geld him, not searching for the retained one. Or, the intention is to do surgery later to find the missing testicle--but the horse gets sold in the meantime and the new owner assumes he's bought a gelding.

Even if the properly descended testicle has been removed, the horse will still act like a stallion. He will try to breed mares, even though he is not fertile. The sex gland retained within the body is small and undeveloped compared to a normal testicle, and it is unable to produce viable sperm because body heat is too high to allow sperm to be created and survive. But it is still producing male hormones. The retained testicle must be removed to rid the horse of those tendencies.

Yarbrough says, "We also worry about testicular torsions, because the testicle is not tethered down. It can become twisted or become the focus of entrapment for a section of small intestine. These are the possible long-term health risks for the abdominal cryptorchid." He said the horse owner needs to be aware of these risks.

Ahmed Tibary, DVM, PhD, associate professor of theriogenology at the Veterinary Teaching Hospital at Washington State University, says colic caused by a retained testicle might not be discovered until the horse goes to surgery. "Most of the time this would be an incidental finding when the horse is opened up for colic surgery," says Tibary. "You would not suspect this, unless you know the horse is a cryptorchid."

Another problem might be a tumor that has developed because of a retained testicle.

"The most common thought as to why tumors develop is that because the testicle is retained, it has a different temperature environment," says Tibary. "Cells that are programmed to function under a certain temperature now have a higher temperature, which may trigger abnormal growths. Another theory is that the testicle may have been retained because it was abnormal to begin with. That can help explain the tumors called teratomas, which are made up of misplaced embryonic tissue."

The testicle's reproductive tissue provides ideal conditions for this misdirected tissue growth. In these instances, a teratoma (usually a grapefruit-size mass of fetal material that can contain hair, teeth, and connective tissue) forms in the abdominal cavity where the testicles originally develop. This type of tumor starts early, in the fetus, and might continue to grow during the life of the horse. The tumor can become life-threatening if it becomes large and puts pressure on other organs, or if it becomes malignant.

"Normally, testicular cancer in horses is not malignant, but there have been cases where these tumors have metastasized," says Tibary.

Diagnosis

If you are uncertain whether a horse is a cryptorchid, he can be checked by a veterinarian. Careful palpation of the scrotum and the tissues above it, and rectal palpation to check the pelvic area, can give clues. But a retained testicle is sometimes hard to palpate in the standing horse. If it's in the abdomen, it is usually suspended by a fold of peritoneum (the membrane that lines the abdominal cavity).

According to Tibary, rectal ultrasound can often help locate it, but not in all cases. The testicle is somewhat mobile within the abdomen and might be hard to spot if it has become mixed up with coils of intestine or lies beside the bladder. In some cases, it adheres to the wall of the abdomen or to an internal organ (such as the spleen) and can be hard to identify. A retained testicle within the abdomen is usually small, flabby, and undeveloped, but it can be very large if it becomes tumorous.

"There is a test that can be used to differentiate between a completely gelded horse and one that has a retained testicle in the abdomen," says Tibary. "What you look for in the test is the response of the horse to an injection of HCG (human chorionic gonadotropin) to know if he still has some testicular tissue. Blood samples are taken before the administration of HCG and one to two hours after. If you see a two- to three-fold increase in testosterone, you are dealing with a horse that has a retained testicle. There are other hormonal tests, but I like this one because it is very simple and works well. If interpretation of the testosterone level is not clear enough with the HCG, then the other tests can be done."

Some fully castrated males still have some stallion-like behavior. If you buy a gelding and wonder if he is a gelding or a cryptorchid, the hormone tests are useful.

Yarbrough says, "Any time we are faced with a case in which there is questionable history on the horse--if we don't know if he was completely gelded or not--we run a hormone profile. If the hormone profile comes back indicating that there's a testicle present, we go in with a laparoscope (used to view within the peritoneal cavity; similar to an endoscope) to explore for it."

If only one testicle was removed in an earlier castration, you don't know which side it was on. Using a laparoscope makes it easier to find the missing one. And if a stallion has only one testicle in the scrotum, you don't know whether this might be a rare case of monorchidism (the horse had only one testicle to begin with). A hormone assay can tell you whether to go looking or not, says Yarbrough.

Surgical Removal

Most owners of a cryptorchid want the retained testicle removed. A testicle caught in the inguinal canal is easiest to address. The veterinarian reaches into that canal to search for the missing testicle and works it out through the canal. "These can often be palpated, or located by placing the ultrasound on the scrotal area," says Tibary.

Yarbrough says the standard inguinal approach to search for the missing testicle is the oldest method. The horse is on his back, anesthetized. "Most practitioners will cut very close to or immediately over the external inguinal ring and go down the canal looking for the inguinal extension of the gubernaculum," says Yarbrough. "They follow it to the internal ring. If the horse is a high flanker, you would encounter the testicle before you get to the abdomen, but otherwise you'd follow the canal right on into the abdomen, to the internal inguinal ring. Once you have identified that structure, you can nip the internal sheath and tease the testicle out."

A testicle retained in the abdomen is usually very soft and smaller than normal. Therefore the inguinal ring doesn't have to be opened very much to get it out. "If you do have to open it, you must make sure none of the small intestine comes out through that hole," says Yarbrough. "If that happens, you must suture the external ring closed."

In earlier years, surgery to find and remove an abdominal testicle could be risky for the horse. If the veterinarian didn't find the testicle within the flank, the incision was enlarged enough to reach a hand into the abdominal cavity, perhaps as far as into the area where the testicles were originally formed. Many of these retained testicles have moved within the abdomen, however, and it could take a lot of searching to find them. The farther up in the abdomen (and the more difficult to find), the more risky the surgery. It might take the veterinarian several hours to find the retained testicle.

With laparoscopy, this is easier and safer, and very helpful in cases where the testicle is difficult to find. A laparoscope can be inserted through a small incision in the abdomen, into the peritoneal cavity, and the testicle can be brought out through this small incision after it is located. "It's not as invasive, and it's visual," says Tibary. "You don't have to open the abdominal cavity, as was done in the past.

"Our surgeon here at WSU, Dr. Claude Ragle, is very good at laparoscopy and does quite a few of these surgeries," continues Tibary. "He gets a lot of referral horses, particularly show horses and performance horses where the owner does not want to leave much scar. In some cases, the procedure can be done with the horse standing, just sedated--which makes for less stress on the horse."

Yarbrough says laparoscopic surgery is a bit more time-consuming (45 minutes to an hour) than a traditional surgery (15-20 minutes) if the horse is on his back and anesthetized. The table is tilted so the abdominal contents fall forward, away from the area to be explored, and the abdomen is inflated like a balloon for the laparoscopic examination.

If the laparoscopy is done standing, the time and cost might be more comparable to regular surgery because you won't have the added cost of anesthesia. As a rule, however, a laparoscopic procedure will cost a bit more due to the cost of the equipment being used. But in a complicated case, the laparoscope can save time and stress on the horse. "Some veterinarians do all their cryptorchid surgeries with a laparoscope, but most of us use it simply for optimal case management--in cases where traditional surgery would be more difficult or risky," says Yarbrough.

"If a veterinarian had already explored the horse inguinally and could not find the testicle, or maybe removed one testicle and came back later to do a hormone profile to see if there is another one up inside and has proven that there is, that's usually when we go in with the laparoscope," he says.

If the horse has both testicles retained, or has already had the descended testicle removed, it's easy to do the laparoscopic surgery standing. If there is still a down testicle to deal with, however, it's usually safer to have the horse anesthetized and on his back, because standing castration can be risky, says Yarbrough.

"The laparoscope allows you to do a much greater exploration, and the area where you remove the testicle is at less risk for evisceration (prolapse of the intestine through the incision)," explains Yarbrough. "The incision is smaller, and in a structure you can suture nicely. If the horse is standing, you pull the testicle out through a region low in the flank. If he's on his back, you pull it out through the dense muscle and fascia, which you can suture--making a more solid closure. The inguinal ring and inguinal canal structures (that you go through to find the testicle in a traditional surgery) are more risky to suture."

When to Do It

Sometimes people wait to geld a cryptorchid, trying to give him time for the undescended testicle to come down. A few young horses do have temporarily retained testes that are caught in the inguinal canal below the abdomen. This occurs most often in ponies. The pony colt might have one small testicle that does not descend when the normal one does. The smaller one might eventually grow larger and descend into the scrotum by the time the animal is two or three years old.

If you check a young horse and cannot feel the testicles, how long you should wait before you determine the colt is truly a cryptorchid?

Tibary says the testicles should both be in the scrotum at birth or within a few months. "However, there are reports of some stallions being two years old before the testicles descend," he says.

If a testicle is caught partway up the inguinal canal, there is a chance it will come down when the horse matures. Enlargement brought about by puberty, and increased weight of the testicle, helps pull it down. But body heat in the flank might inhibit proper growth of the testicle and it might not enlarge and descend.

"Once it has been determined that a colt is a cryptorchid, it is generally recommended that the retained testicle be removed by the time he is about 12 months old," recommends Tibary. "If the horse is still a high flanker or abdominal cryptorchid, it's time to do something about it."

Yarbrough says that if the testicle has not descended through the internal inguinal ring by the time the ring starts to contract (at about six months of age or earlier), it will not get out and will be permanently trapped in the abdomen. "It is possible for a high flanker to have a testicle not make it all the way down to the scrotum until two years of age, but that's very unusual," says Yarbrough. "If you know the colt is a cryptorchid, there is no real advantage to waiting. It's actually easier to do an inguinal approach on foals because they have less inguinal fat, and the blood vessels are smaller, so the exploration is easier."

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CRYPTORCHIDS AS STALLIONS 

Keeping a cryptorchid as a breeding stallion is very controversial, and most horse owners and veterinarians advise against it. There is a genetic component to this problem, and offspring might have a greater chance of having the same problem. Some breeds allow cryptorchids to be stallions; others don't. Cryptorchids are more difficult (and expensive) to geld, and a cryptorchid stallion can be at a disadvantage for breeding. If anything happens to his one functional testicle, he is out of business as a sire. He will still be fertile with only one testicle, but will not produce as many total sperm as he would with two testicles, and he won't be able to breed as many mares.

"For a testicle to develop and function normally, it must be able to thermoregulate," says Tom Yarbrough, DVM, PhD, a veterinary surgeon practicing in northern California. "It needs to be at a temperature a little below normal body temperature. That's why the cremaster muscle will draw the testicle up or let it down, depending upon whether the air temperature is warm or cold. So a testicle retained within the abdomen will not be producing viable sperm, and cannot function normally."

This genetic defect is not as harmful to the horse as some other inherited faults, since the affected animal can live and function normally (but he might have aggressive behavior that could put handlers at risk). "We castrate a high percent of these animals, and as long as horse owners don't mind the added expense to geld the progeny of a cryptorchid stallion, cryptorchidism may be an acceptable inconvenience if the horse has other qualities that are desired," says Yarbrough. "It's a matter of whether the horse owner wants to put up with the inconvenience, and whether we want to take on a bigger financial burden when gelding the offspring."

Ahmed Tibary, DVM, PhD, associate professor of theriogenology at the Veterinary Teaching Hospital at Washington State University, says leaving a horse a cryptorchid can be dangerous for people handling him. "Some people will tell you that cryptorchid stallions can become a lot more dangerous than normal stallions, and I have seen this in Thoroughbreds," states Tibary. "The crankiest Thoroughbreds I have seen were cryptorchids."

If a tumor develops in that testicle, it can also change the horse's behavior--by altering the hormone balance, triggering a change in personality.

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INCIDENCE AND HERITABILITY 

Cryptorchidism occurs in most mammals, perhaps more often in domestic animals than in wild ones. We have bred horses selectively for traits that suit our purposes, and sometimes in our efforts to choose breeding stock with characteristics we desire, we overlook others. Thus we might perpetuate or increase the incidence of certain problems.

Cryptorchidism in horses is thought to be inherited, although the actual mechanism of inheritance is not yet fully understood. Ahmed Tibary, DVM, PhD, associate professor of theriogenology at the Veterinary Teaching Hospital at Washington State University says, "It has been confirmed in sheep and swine to be hereditary, and we assume it is in horses. Some authors think it's inherited as a dominant autosome (in a pair of chromosomes), and others think it's a recessive gene (that must be inherited from both parents to show up in the offspring). In my experience, I've seen a lot of cryptorchids in Akhal-Teke horses, a breed with a limited gene pool."

Within all breeds, there are some bloodlines that produce a greater number of cryptorchids than the average for that breed, and some breeds have a higher incidence. A study that looked at more than 5,000 cryptorchid horses (documenting breed incidence, among other things) found the highest number in Percherons, Quarter Horses, Saddlebreds, and ponies, says Tibary.

"From that study, it was also surmised that the breeds that don't have many cryptorchids are the ones that have tried to eliminate this trait by not allowing them to be used for breeding," he adds. "This lends credence to the idea that it's inherited, but at this point we're just using the experiments from other species to assume it is hereditary in horses," he says.

"In that study, it was also found that the left testicle is more commonly retained than the right in horses that are abdominal cryptorchids," notes Tibary. "One speculation as to why this occurs is because of the way the kidneys are placed. One kidney is higher than the other. So it has to do with position of the kidneys and testicles while the fetus is developing."

In ponies with complete abdominal retention, the left testicle is retained as often as the right, but in horses, left side abdominal retentions outnumber the right side cases two to one. Sometimes both testicles are retained in the abdomen (bilateral cryptorchidism), but this is rare. More often just one is retained.

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