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cyst microscopic cropSarcocystosis is one of the most frequent infections of animals, the organisms are found in the muscles and the central nervous system! There are, perhaps, 196 species of Sarcocystis, yet the complete life cycle is only known for 26 of them. Sarcocystis require a prey-predator, 2 host, relationship to survive. Each host supports different stages of the life cycle. Hosts vary for each species of Sarcocystis.  Intermediate hosts support several species of Sarcocystis, each species use different definitive hosts.

People get sarcocystosis, two species use people as the definitive host.  Humans also serve as intermediate (or aberrant hosts) for several other species. Raw beef can infect you with S. hominis and raw pork can give you S. suihominis. Gastrointestinal symptoms are present when Sarcocystis cause disease in people  definitively hosting the parasite. Muscle pain is reported in people that serve Sarcocystis as an intermediate host. Most of the time people are asymptomatic with muscular sarcocystosis. People can get sick from an enterotoxin associated with Sarcocystis that infect horse muscles. if horse meat is uncooked.

Horses get sarcocystosis from dogs, the organism causing infections is S. equicanis (bertrami). In the US the horse-infecting organism is named S. fayeri. Donkeys harbor S. asinus, but experiments may show that this is actually S. fayeri. The visible difference in equicanis (bertrami) and fayeri is the thickness of cyst walls in muscles observed under the microscope. Our picture above is fayeri,  a thick-walled sarcocyst.  S. bertrami forms a thin-walled cyst. Dogs shed infective betrami organisms 8-10 days after eating infected muscle tissue. Horses eat the organisms from dog feces-contaminated feed.  It takes two to three weeks before signs appear in horses.  Signs include fever, neurological signs, apathy, and inappetence  a couple of months after infection. Muscle enzymes can be elevated in infected horses and increased enzymes can be measured in blood samples.

Sarcocystis fayeri cysts are found in skeletal muscles and the heart.  Ten and 25 days after infection (two waves of organisms are released from the gut) the developing parasites are found in arteries of the heart, brain, and kidney.  Muscle cysts (sarcocysts) are first seen at 55 days .  And by 77 days the muscle tissue can infect dogs to start the cycle again. We are taught that S. fayeri is only mildly pathogenic, horses develop anemia and fever after infections and some horses may have a stiff gait. More virulent isolates can cause inflammation of muscles and, in one case, the horse developed autoimmune anemia.  People that eat raw horse meat can get food poisoning, this is due to a toxin associated with the cysts.  A fetus can be infected by S. fayeri by crossing the placenta, so foals can be born with mature cysts in their muscles.

Malnourished horses show clinical muscle inflammation (myositis) and muscle atrophy that can be associated with S. fayeri. Muscle cysts are usually unassociated with clinical signs or muscle inflammation, leading to conclusions that fayeri infections are benign. Most clinicians agree S. fayeri isn’t an issue in horses.  However, there are some researchers that think S. fayeri should be considered in horses with neuromuscular disease. Scientists at UC Davis examined muscle tissues from horses with and without a history of neuromuscular disease.  They found an association, but not statistical significance, between disease and cysts.  More horses with neuromuscular disease had S. fayeri cysts when compared to the number of cysts found in horses that died from other causes.

We decided to look at the problem a different way.  While UC Davis looked at cysts, we chose to look for S. fayeri antitoxin in the serum of horses. An advantage to antitoxin analysis is that the test is performed in the live horse.  We found that 24% of normal horses had antitoxin in their serum. We also found that S. neurona antibodies were more often associated with neuromuscular disease in horses  than S. fayeri antitoxin (antibodies against the toxin released from cysts). That said, horses were more often infected with both species of Sarcocystis, not just one strain.

Horses infected with S. neurona and S. fayeri were more likely to show disease. We looked at inflammation using CRP (C-reactive protein) and found that CRP was detected in horses with and without apparent disease. We did find that significantly more horses with neuromuscular disease had an elevated CRP when compared to normal horses (p= .0135).  However, the data we needed to link the UC Davis study and ours was missing.  We needed to show  the relationship of antitoxin found in horses (our test) with sarcocysts found on histopathological slides (the gold-standard test for fayeri-sarcocystosis).

We examined three tissues (muscle tissue from the heart, esophagus, and skeletal muscle) from thirty-two horses (that’s a big number in horse studies) and measured the presence of antitoxin.  Our results confirmed that our serum test was almost the same as the results you would get if a full post-mortem exam was conducted. Also, in our study the infected horses were asymptomatic.

The conclusions thus far are that S. fayeri can be detected pre-mortem and should be considered in horses with signs of sarcocystosis or neuromuscular disease that has no other cause.  Also, horses with neuromuscular disease are more likely to have an elevated CRP.  In our S. fayeri study horses did have elevated CRP values but the inflammation was associated with other gastrointestinal parasites, not EMS (equine muscular sarcocystosis).

Why are we still interested in S. fayeri? Certainly our results support the strain of S. fayeri  infecting our 32 horses was non-virulent and support the view of most clinicians that S. fayeri may not be an issue in most horses, just some of them.

For us, it’s about the toxin.  The toxin is an actin-depolymerizing factor (ADF).  Another Apicomplexan, Toxoplasma gondii, has a very similar ADF. The TG-ADF can protect lab animals against T. gondii infections. Interestingly, some animals with lethal S. neurona infections also were infected with T. gondii. The difference between protection (mice) versus no-ADF protection (sea otters) could be stage of infection (with TG), ,strain-virulence, or something we didn't think of yet.

Can S. fayeri-ADF protect against S. neurona infections in horses? Is ADF species specific? Is ADF a virulence factor or even a survival factor for the organism? Are there cyst-producing S. fayeri organisms that don't produce ADF? Does S. neurona produce an ADF? And, can a S. neurona be incorporated into a fayeri sarcocyst? These are all questions to be answered by experiment.

Here is our opinion. It is worth monitoring S. fayeri infection in horses. It is worth considering S. fayeri infection as a cause of neuromuscular disease in horses showing weakness, muscle atrophy, and inflammation for which there is no other explanation. The disease EMS should be considered in old and debilitated horses.  Sarcocystis fayeri should be considered in horses with chronic inflammation. When  horses showing weakness do not have antibodies against S. neurona, EMS should be considered. Horses that provide meat that is fed to dogs should be monitored for S. fayeri.

If you have questions about S. fayeri, give us a call.

Acetylcholine_binding_proteinOur business is consulting.  That means we dispense advice.  We don’t look into a crystal ball or read tea leaves. We base our opinions on an analysis of your case and immunodiagnostic testing.  Analyzing a case involves looking at laboratory values, the case history, and physical examinations done by licensed veterinarians.  The underpinning of our opinion is immunodiagnostic testing associated with neurological diseases and understanding the parameters and limits of each diagnostic test we use.

We are not on the sidelines when it comes to the literature on sarcocystosis. Our works are published so other experts, and you, can critically review our approach.  We rely heavily on works of others to develop new theories that are tested by direct observations.  Our hypotheses’ are scrutinized by our Statistician, he has a PhD in statistics, years in the pharmaceutical industry, and is considered one of the best in the industry.  So when we are asked a question, we answer with our objective opinion with statistical significance.

Recently we were asked, “Can you clarify your comment that the “titer” is an indication of duration of infection (vs the degree of disease) and explain the difference between a negative IFAT test and the SAG titer of 8?” It seems to be a simple enough question.  To get to the answer, one has to A) know how the organism behaves in the horse, B) correlate antibody levels with clinical disease in horses, C) understand the critical differences in quantitating antibody between test formats.

How does a Sarcocystis behave in an animals that it infects?  The authority would be Dr. Edith Box.  Throughout the 1980’s Dr. Box infected budgies with S. falcatula.  It is historically important that budgies were lethally infected by S. falcatula. An aside to the question posed above is the 1995 hypothesis by researchers from the University of Florida.  They presented molecular evidence that S. falcatula and S. neurona were the same organism!  Their hypothesis was refuted by demonstrating that S. neurona does not infect budgies at all. Horses are infected by S. neurona, budgies not. Budgies, but not horses, are infected by S. falcatula. Until proven otherwise, another study conducted at UF is the law of the land.  Their horse-infection study indicates that horses are not susceptible to S. falcatula infections. 

How does a Sarcocystis behave in a natural host? S. falcatula in budgies is the model. Box showed that the organism is distributed throughout the organs from the gut (the end result is a muscle cyst).  New flushes of organisms are not lingering in organs outside the gut.  New infections didn’t spring up from muscle cysts either. Box showed that some birds did get infections that went to the brain. (We always suspected S. neurona wasn’t special in a proclivity for brain tissue).

Wait, you say. What about organisms that are not in the natural host? The horse is not a natural host for S. neurona.  Usually Sarcocystis are highly host specific. Host specificity is how UF used biological studies to correct and retract their claims that neurona and falcatula were the same. When based on molecular data they were pretty darn similar.  When tested in animals, not so. Other studies support that horses with an immune system quickly remove organisms just as the bird model shows for falcatula. Interestingly, horses with abnormal white blood cells do not clear the organisms at all…and paradoxically the immune-deficient horses do not show clinical signs.  Heads up!  The immune response to the organism caused the clinical signs, not the organism.

And now back to the basis for answering the original question.  We did it by animal experiment, first developing an infection model in the horse. We infected 75 horses with S. neurona using white blood cells to mimic a “natural” infection. We didn’t evaluate the horses ourselves, we brought in EPM-experts from across the country (at least 2 for each study) to examine the horses as the disease progressed.  They were unaware of which horses were infected and which ones were not so we “blinded” the experiment.  Experts saw the horses before infection and then every month until the experiment was terminated four months later. They scored the disease using a meticulous neurological exam.  Yes, they documented the horses got worse over time.

The horses’ blood and CSF fluid was taken every month at the same time the exams were conducted.  We used a SAG 1 ELISA  test and also sent duplicate samples to a Kentucky laboratory. Both laboratories tested the samples for the presence of antibodies.  That way there was no bias in testing.  We used only one test because a SAG 1 strain of S. neurona was used for the infections.  In natural infections, there are three serotypes of S. neurona, SAG 1, SAG 5, and SAG 6.  Each SAG is mutually exclusive-only one SAG is present in an organism. Multiple serotype infections are possible in the field because an opossum can be infected by all three serotypes.  Three different infections. Our experiments were limited to a single serotype. We sent the KY lab a sample of SAG 1 to run in their test making sure they had the same marker that we had on our test. Thus, any difference between the tests would be a consequence of ability to detect and interpret a single protein.

Our Statistician crunched the numbers.  He concluded that as time progressed, the clinical signs got worse and that was statistically significant.  Also, as time progressed, antibodies increased, but the antibody level did not statistically correlate with the degree of clinical signs observed by the clinicians. 

And there was individual variation in the antibody response.  Some really sick horses had lower antibodies when compared to not-so-affected horses that had high antibodies. The Kentucky test was less likely to identify infected horses.  Twenty-five percent of the test results differed. That means that had we conducted the experiments only using the KY lab, the interpretation of the study would be different. That is why different scientists have different views of immunodiagnostics.  We base our opinion on study results.

Subsequent experiments showed that horses that had never experienced an infection produced less antibody and dropped the antibodies quickly when compared to “experienced” horses.  Experienced horses got a measurable antibody response quickly (the scientific word is amnestic response) and retain those antibodies for up to 10 months.

The experiments in 75 experimental infections allow us to say that in horses the antibody response to SAG 1 will increase over time and linger longer in an “experienced” horse and antibodies are more a reflection of duration of infection, not degree of disease.

A couple of things help explain part C of the question quantitating the difference between two tests.  It is important to recognize that testing formats are different.  That is to say what is measured (antibody against a protein) and how the protein is prepared for testing are different.  It has been known for a long time that reducing conditions change how antibodies recognize a protein.  Without going into the fascinating field of protein chemistry, suffice it to say, a protein is a string of amino acids.  The amino acid sequence is the primary structure of the protein and that is determined by the DNA that codes for the protein. 

The secondary structure of a protein is determined by folds in the backbone of the protein.  This means side groups don’t interact in determining the secondary structure.  The overall tertiary structure of the protein, its 3D structure, is determined by the reactions and bonding between side groups.  The charge of individual amino acids, the affinity or aversion to water, and special bonds are important in tertiary folding of a protein. The amino acid cystine has very strong disulfide bonds to contribute to 3D structure. I digress to point out that Sarcocystis surface proteins characteristically have a lot of cystines and it can be expected these amino acids give important tertiary structure to the SAG’s. One would speculate, without any other knowledge about the protein, that the cystines give the protein its functionality.

A horses’ immune response is to the tertiary structure of the SAG proteins making antibodies to the 3D molecule, not the primary structure.

Think of a protein as a rope, twisted it up upon itself into its 3D structure.  Now mentally dip that twisted up protein into a vat of dye-lets imagine red.  You have tie-dyed your protein!  When it is unfolded into its primary (linear) structure you will see where areas of the protein, that were far apart, touched when folded.  Red marks are distributed seemingly randomly along the molecule and there are long areas between the marks that have no dye.  The scientific term for the areas that touched are “conformational epitopes” and the areas that are side by side are called  “linear epitopes”. An “epitope” is a section of  amino acids that induce an antibody reaction in a host.  You can correctly guess the amino acids do not have to be side by side to make the host react, in fact you expect the 3D structure to be more important in inducing a response.  Conformational epitopes are important in immunodiagnostics.

Some testing formats use chemicals to stretch out the proteins into linear molecules.  Other formats retain the 3D conformation of the protein.  You can correctly guess that each test would measure antibodies that recognize the same protein, but by different markers.  We have always favored conformational epitopes and run our tests without protein-straightening chemicals. Our thoughts are not original. It is long known that Sarcocystis infections are more recognizable when diagnostic tests that use conformational epitopes are selected.

We evaluated SAG 1 and recognized a relationship between breaking tertiary bonds and the concentration of chemicals used to break those bonds. It was dose dependent. That means that the KY lab that tested samples from the experimentally infected animals looked for linear epitopes. and depleted the reactions to SAG 1. We keep the SAG 1 for detection folded in its happy state. 

You correctly point out that an IFAT test uses the whole organism.  Are the surface proteins altered in the IFAT test?  It depends.  It matters how the organisms are prepared for the test. More important to the argument is that commercial IFAT tests are run on SAG 1 strains.  Great for experiments using SAG 1 organisms to cause infections, but not so great in field infections that are caused by SAG 5 and SAG 6 strains, as well as the more common SAG 1 strain.

There are other complicating issues: Sarcocystis are known to selectively stop displaying some proteins during infection, different testing labs use different dilutions of the serum, some tests are measured by machine and some are subjectively evaluated by lab technicians. All things considered, an IFAT reported as negative at a 1:40 dilution tested on a SAG 1 strain is not comparable to a 1:8 dilution (titer) on a SAG 5 surface protein.

We take these things into consideration for you as we consult on your case.

 

lab opossum

Neurologic deficits are seen in horses with abnormal gaits, changes in behavior or signs that are limited to the cranial nerves.  These horses are difficult to diagnose because the list of etiologies that result in neurologic deficits is long. The list of disease-causing possibilities is long because animals have a short list of responses to infection and injury.

Long ago lowly slime molds and paramecia selected and perfected a group of chemicals that allowed them to achieve movement and the ability to communicate with other organisms. The chemical signaling mechanism that parasitic protozoa and animals use today is based on these evolutionary successes. Redundancy is built into the chemical structures and the way signals reach the target tissues. Redundant molecules also are used to turn off these systems. The basics of the pathways are common to all living things and they are primal.

Signaling pathways are also common to many tissues and they are ubiquitous throughout the body. An infectious agent will set off  a protective primal response in an animal. That response is called “innate immunity,” and this immunity results in signs that are non-specific to a single agent.  Vertebrates have many checks-and-balances regulatory pathways that control these common systems most of the time.  Sometimes the pathways become unregulated. As animals evolved to efficiently use the chemicals (cytokines) involved in protection against infections, parasitic microbes exploited the same systems. These highly successful disease-causing organisms use some pretty nifty  methods to evade or even hijack the innate immune signaling cascade.

The result is that an animal with neurologic disease has a limited repertoire of responses to several insults and they all look the same to the clinician.  We have accrued a vast amount of scientific knowledge about some diseases.  The diagnosis of these diseases is verified by  objective methods.  Some diseases have classically presenting signs that allow a veterinarian to easily rule them in or out. Of course, field experience is valuable in recognizing some classic signs. The diagnostician thins the list of possible causes of disease to the short list.

The short list for neurologic diseases that are found in horses often don’t have definitive diagnostics and that leads to using exclusion to help diagnose the cause. A diagnosis of exclusion  is a diagnosis of a medical condition reached by a process of elimination, which may be necessary if the presence cannot be established with complete confidence from the history, examination or testing. Such elimination of other reasonable possibilities is a major component in performing a differential diagnosis.

Diagnosis by exclusion tends to occur where scientific knowledge is scarce, specifically where the means to verify a diagnosis by an objective method is absent. As a specific diagnosis cannot be confirmed, a fall back position is to exclude that group of known causes that may cause a similar clinical presentation.  Polyneuritis equi (PE) is such a disease.  We have, along with our regulatory partners, put together a list of definitive tests and some that are exclusionary to pinpoint the tests most likely to give us the target population for our polyneuritis equi study.
http://pathogenes.com/w/polyneuritis-equi-is-an-overlooked-disease-part-1/
http://pathogenes.com/w/polyneuritis-equi-is-an-overlooked-disease-part-2/

The PE horse has evidence of neurologic disease that can be identified by neurological exam and those that are treatable have serum antibody against some specific proteins.  These are inclusionary criteria for our study.  The horse may or may not have antibodies against parasitic protozoa. To keep our study uncomplicated, we exclude horses with antibody against S. neurona, a parasitic protozoa that causes equine protozoal myeloencephalitis (EPM). Other exclusionary criteria are no recent history of trauma, no  recent respiratory infection (or a current vaccination for EHV-1 can satisfy this one).  Vaccination for rabies will exclude rabies as a cause.  Of course,  a horse would need to have a normal vitamin E level ( > 1.5 micrograms/ml serum).

Some treatments will exclude horses from our study.  They include anti-inflammatory agents within 3 days and anti-protozoal agents within a time frame that the anti-protozoal is expected to exert its effect.  Each expected effect from licensed anti-protozoal medication is different, some 30 days and some beyond 90 days.

Diagnostics are a major frustration for owners of horses with suspected EPM.  Owners spend many dollars on exclusionary diagnostics and don’t have an answer at the end of it.  Unfortunately, that is the nature of EPM. It is also the nature of Sarcocystis infections to trigger innate immune inflammatory cytokine responses that produce the signs associated with neurological diseases.  That means the horse can have two active pathological processes!  Because the nature of these diseases and the repair mechanisms that are associated with them use similar pathways, it is unlikely that there will be definitive diagnostics.  A good neurological examination by a veterinarian familiar with neurological disease and judicious use of diagnostics to form a short list  is the most successful path to health.
See: http://pathogenes.com/w/the-biology-of-sarcocystis/

crystal ballThe orbuculum, or crystal ball, was invented about 3000 BC according to Wikipedia.  Mystical orbs were used in numerous cultures to communicate with the gods or learn of future threats. Wouldn’t it be nice to identify horses that are genetically pre-disposed to get sarcocystosis?  A genetic EPM-crystal ball.

Thirteen years ago, we participated in studies that sought to identify cell markers unique to horses with equine protozoal myeloencephalitis, EPM. The idea was that cell markers or the “gene expression signature” unique to EPM would be found in immune cells circulating in the blood. The differences in gene expression between animals with and without clinical evidence of EPM would be analyzed using several statistical measures. Genes that showed statistically significant differences in clinically positive horses were compared to those that were clinically negative, and the genes that showed a significant difference (those significantly up-regulated) would constitute the EPM-gene signature.

Controlled laboratory studies testing the hypothesis that a gene signature could be found and  be useful in the diagnosis and treatment of EPM were undertaken. A controlled infection (induced stress) study was used to accurately know the day of exposure. Sarcocystis neurona oocysts were administered to 20 stressed horses to elicit disease. Blood samples were taken 10 times over 28 days to collect RNA, the measure of a turned on gene. The up-regulated genes (identified by the RNA analysis) were assayed on a custom microarray for determining gene expression (the specific array was patented, but not by us). In this blinded study (veterinarians didn’t know which horses were infected) clinical exams were performed,serum and CSF were tested, and post-mortem exams were conducted to ensure that clinically ill horses did get EPM.

This experiment was eventually published with the infection data, but the gene analysis data was not reported. Remarkably, the horses that showed signs didn’t have organisms that could be demonstrated in the brain tissues. Inflammation was considered diagnostic of successful infection. Scientists conducting this study identified a gene signature. Success! There were differences between the up-regulated genes in the clinically ill horses that were infected and those that were not infected, control horses.

Time to test the gene signature. Field cases of suspected EPM were used in a second study. The gene expression from horses with suspected EPM, those that had serum and CSF analysis to be as sure as possible the horses fit the diagnostic criteria at the time, came from clinical cases. It took 6 weeks to process the samples and get a result because the assay is technical. An obvious down side of the endeavor was cost. The hundreds of dollars that the eventual assay would cost, and the six-week turnaround time, made it a clinical non-starter. More importantly, the assay didn’t diagnose chronic disease (disease that was present after 28 days from the initial infection). The cases presented to veterinarians are chronic. The acute gene signature did not identify field cases that the veterinarians diagnosed.

In a third study, 13 animals were used in a merozoite challenge model that did not use stress. The horses were randomly assigned to a group, 8 were challenged while 5 were sham challenged. This study ran 90 days to detect an acute and chronic gene signature. Cells were assayed for gene expression at 28 days (acute) and 90 days (chronic). If the acute gene signature was the same in both models, an accurate marker between the two studies could be identified to identify acute, possibly current, disease. Likewise, the chronic markers, significant expression of genes at 90 days should match the field study and identify horses with long term disease even if the organisms were eliminated.

The cumulative results of the controlled studies identified 31 genes that were highly statistically different at day 28 between animals that developed clinical EPM and those that did not. An EPM index score calculated for the gene signature, identified in the first controlled study, was successfully used to identify some, but not all, of the horses with acute disease in the third, controlled study.

Horses with chronic EPM, day 90 of the third study, were not identified using the signature developed in the first study. Further, chronic EPM could not be identified in clinical field samples using the gene signature developed from acute disease, day 28, in either the stress model or the merozoite model. Because many horses present with suspect EPM after having had clinical signs for weeks or months, the value of a genetic signature was doubtful.

We identified drugs that selectively reduce the expression of some of the upregulated genes that were stimulated during acute and chronic disease. Some drugs returned horses to normal, removal of the drugs allowed the horse to again show signs of disease. Surprisingly, some drugs we tested made horses worse! We found that the innate immune response and the genetic signature of host cells are the key to disease associated with sarcocystosis.

The data was useful. The upregulated genes included MHC Class II receptors, chemokine receptors, IgG molecules, natural killer cells, several interferon-induced proteins and a handful of others.

More studies are undoubtedly in the pipeline. As those studies are completed and in a few years published, they may be compared to the work done in 2005. Or perhaps the EPM-gene signature is already relegated to the cutting room floor, the genes frozen in time, yielding space in the freezer for newer endeavors and lost to analysis. Our experience showed us there are disease-signatures present in the blood samples and these are markers that can effectively direct treatment for horses with disease and identify horses that are resistant to disease. We don’t think there will be a crystal ball that predicts which healthy horse will be come sick when exposed to Sarcocystis, sadly it isn’t that simple.

Horses with clinical signs of equine motor neuronprzewalski-1972728__340 disease (EMD) or vitamin E deficient myopathy require supplementation with vitamin E.  Horses that graze green grass should be fine without supplementation.  Supplements are expensive, here is a primer to guide you.

The critical factors associated with vitamin E are: determining that your horse is deficient (solution: test the serum levels); deciding what supplement is most appropriate (solution: determine what are you treating); delivering the dose efficiently to the horse (formulation and dose); and when to discontinue treatment (test the serum levels!)

Supplementing with the intention to increase vitamin E in the central nervous system (CNS) of a horse with neurologic disease requires a different protocol than supplementing for diet deficiency in a normal horse (a horse that has no access to green grass).  Studies show that some supplements do not increase the levels of vitamin E in brain tissue.  Studies show that vitamin E supplements do not increase the levels of the vitamin in muscle tissue.  And some supplements are active at five to six times other formulations!

There are no studies describing toxicity in horses from too much supplementation.  Vitamin E can be toxic because it is stored in fat (lipid) and is not excreted like water soluble vitamins are. It is possible that vitamin E inhibits vitamin A, another fat soluble vitamin although there are no published studies. In other animals, including humans, neurologic complications result from overdosing vitamin E!  There is no reason to suggest toxicity won’t occur in horses.

Horses showing no clinical signs of vitamin E deficiency

Supplementing with vitamin E can be expensive and can put a horse at risk for toxicity. If you suspect a deficiency you can easily test the serum concentration.  Pathogenes offers a discounted program for testing vitamin E in association with our clinical trials.  Call us for more information and be sure to send the sample with our Test submission form.

Is it time to stop supplementing?  A simple test will tell you.  Because there is a rapid decline of serum levels after discontinuing some forms of vitamin E, it is best to wait a week after stopping vitamin E dosing before sending a serum sample for testing a long time supplemented horse.

Clinically ill horses

Horses showing clinical signs of equine motor neuron (EMD) disease or vitamin E-deficient myopathy can benefit from treatment.  Often suggested, but not proven, vitamin E is supplemented in  cases of active equine protozoal myeloencephalitis (EPM), with or without measured low levels.

Determine the base line levels of serum vitamin E before supplementing.  In diseased horses requiring supplementation, 5000 IU/day of a soluble, natural form is useful.  Not all horses respond the same way to supplements- there is individual variation!  After two weeks of supplementing the serum level should be assessed and adjust the dose accordingly.  A tapered regime with a gradual transition to a natural powder form of vitamin E is  appropriate.  The natural power form of vitamin E will return a horse to a normal serum value in 7 weeks but  normal CSF levels are not achieved with this supplement-form.

Levels of vitamin E

Normal serum levels of vitamin E in horses are greater than 2.5 µg/ml.  A level that is considered adequate is a range between 1.5-2.4 µg/ml.  Horses with serum levels less than 1.5 µg/ml are deficient.

Normal levels decline significantly in just 18 days in horses that are not allowed access to grass and are fed a pelleted ration that is not supplemented.  Considerations here are horses stalled due to colic surgery, metabolic syndrome, or other similar conditions.  Horses don’t have green grass in northern climates during the winter, something to consider.

Vitamin E levels in serum
Normal 2.5 µg/ml
Adequate 1.5-2.4 µg/ml
Deficient less than 1.5 µg/ml

It is interesting to note that in a controlled study there were no differences in the mean concentration of CSF vitamin E in un-supplemented (normal levels were present) or supplemented horses (all supplements).  Or in deficient horses, before and after supplementation!  There was a significant (linear) correlation between serum and CSF concentrations, the higher the serum level the higher the vitamin E in the CSF of most horses.  It is possible there is a limit to the amount of vitamin E that can be measured in the CSF (does it all go to the cell membranes) or it is a fault in the testing protocol (little correlation of values with test results).

Types of supplements

Vitamin E is available as an injection, usually in conjunction with selenium, and is a form that is by prescription for use by a licensed veterinarian.  Some serious and life threatening reactions can occur with intravenous or intramuscular injections of vitamin E-selenium.  The injectable form bypasses the inhibition seen in some oral formulations. Oral synthetic and natural vitamin E preparations are available.  The synthetic has eight stereoisomers (the molecular shape of the molecule and its rotation).  Animals have a preference for only one.  Natural vitamin E comes in only one isomer, the one preferred by the liver.  There are two synthetic forms of acetate, the powder is twice as available to the animal as the pelleted form, the powder increasing serum concentrations in about two months.  The water soluble, liquid form, is five to six times as available for uptake by the horse and increased concentrations are accomplished in 12 hours.  Thus the acetate forms elicit a gradual increase when supplemented.

What?  Who said sulfur inhibits vitamin E?

Vitamin E and selenium are intertwined with sulfur metabolism.  There is a relationship between selenium and vitamin E overcoming sulfur-induced depletion in the body.

What the vitamin E Guru’s suggest

Horses that have no clinical signs of deficiency can be supplemented with the less expensive acetate forms at 10 IU/kg body weight per day over months to achieve normal serum vitamin E levels.

The acetate form isn’t a good choice in horses with clinical signs of EMD or vitamin E deficient myopathy.  These horses require an immediate increase in serum and CSF vitamin E concentrations.  The veterinarian can use an injection to rapidly increase levels and the treatment can be repeated at 5-10 day intervals.  This form is labeled for selenium-tocopherol deficiency syndrome that presents clinically as rapid respiration, profuse sweating, muscle spasms and stiffness accompanied by an increased SGOT (liver enzyme).

Levels can be restored to normal by giving 5000 IU/day of the soluble vitamin E and then tapering the regime to transition to 5000 IU/day of the oral acetate. This protocol resulted in horses with a prolonged increase in CSF concentrations 8 weeks after beginning supplementation.

Sulfur in the digestive track can inhibit vitamin E uptake, in sulfur-inhibition resulting in deficiency, an injectable form is preferable.

What we suggest

Test the serum  vitamin E levels before supplementing this essential nutrient and again after 7 weeks of supplementation. After 7 or more weeks, discontinue the supplement for 7 days and then test.  It may be wise to re-evaluate serum vitamin E levels after several weeks on the acetate form to ensure concentrations remain within a normal range. It may be of value to determine vitamin E levels in horses suspected of EPM.  A diagnosis of the neurologic disease EMD can be supported by measuring a low serum vitamin E concentration.  Test these horses when they are tested for suspected EPM and once on therapy, 7 weeks later.  Horses with equine degenerative myeloencephalopathy (EDM) will not respond to vitamin E.  EDM is an inherited condition that prevents uptake of vitamin E early in life.  Once neurological signs are present they usually don’t get worse…or better in these EDM horses.