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Early Embryonic Mortality

by Patrick J. Meyers
Meyers Equine Clinic
Hillsburgh, Ontario, Canada

What is Early Embryonic Mortality – EEM?

In order for us to talk about early embryonic mortality (EEM) in mares let’s first of all define what it is. In the strictest of terms, EEM refers to the loss of an established pregnancy during a period of gestation known as the embryonic period. Synonyms for early embryonic mortality include early embryonic death and embryonic loss but not fetal death, fetal mortality or abortion. Pregnancy loss is an all encompassing term that includes the loss of the embryo or fetus regardless of when during gestation it occurs. An embryo is defined as the product of a fertilized egg (the result of a successful mating) from conception to the completion of the stage of organogenesis (when all of the embryo’s major body organs have differentiated). In the equine, the embryonic period includes the first 40 days of gestation. After the embryonic period, the embryo becomes a fetus until it is born. Therefore, early embryonic mortality in mares is defined as the loss of the embryo between fertilization and day 40 of gestation.

How is early embryonic mortality detected?

EEM has been proposed as a factor involved in reduced fertility in mares since the 1940s but much of the recently revived interest in embryonic loss is due to the introduction of the ultrasound machine (in the early 1980s) for early pregnancy detection. EEM can be expressed as a percentage of embryonic loss in mares in which the embryo is detected or present on the first given day and dead or absent on the next. Prior to the use of diagnostic ultrasonography in mare reproduction, studies on the incidence and time of embryonic loss in the mare relied primarily on per rectal palpation of the mare’s reproductive tract. Although presumptive diagnosis of pregnancy (increased tone and tubularity of the uterus and a tight elongated cervix) could be made in this manner by an experienced veterinarian as early as days 18 to 22 after mating, initial detection of the embryonic vesicle (pregnancy “bulge”) could not be made until day 25 to 30. Therefore, embryonic losses before day 20, could not be differentiated from failure of fertilization except by inference when the mare became pseudopregnant. Recent studies of fertilization rates in both normal and subfertile mares indicates that fertilization failure represents a very small proportion of fertility losses. Detection of the embryonic vesicle can be made as early as day 10 with transrectal ultrasonography. Accuracy of embryo detection at day 11 can be 98% with a high frequency (5 MHz) ultrasound scanner. Embryonic loss before day 10 can not be detected unless embryos are recovered via embryo collection procedures. Some work has shown that a significantly lower number of embryos are recovered from subfertile mares compared to mares of normal fertility.

What clinical signs (symptoms) do we see with EEM?

With EEM in mares the most common clinical sign that an owner sees is infertility or a syndrome called “repeat breeding”. Repeat breeding simply describes a mare that consistently returns to heat on regular 21 day cycles after mating. Repeat breeding could, in this situation, be due to the failure of fertilization but more likely is due to early embryonic mortality. If an ultrasound examination is not performed on the mare between day 10 to 20 after mating the owner will never know if his/her mare ever conceived. The mare may have lost the pregnancy during this time.

In the mare, early embryonic mortality is very seldom seen visually, due to the rapid uterine expulsion or the intra-uterine resorption of the small ovoid embryonic vesicle. Death of the embryo and complete resorption of embryonal remnants and fluids prior to the time of maternal recognition of pregnancy (MRP), at day 14 of gestation, allows release of prostaglandin, destruction of the corpus luteum, and the mare’s return to heat at the expected time of her next cycle. Clinical signs of disease are usually inapparent in the mare. An examination of your mare by a veterinarian usually reveals no abnormalities. A vulvar discharge may sometimes be indicative of an endometritis (uterine infection). Endometritis may have been present prior to mating or the infection may have been acquired at breeding. Endometritis provides a hostile uterine environment which is toxic to the embryo or causes enough irritation of the endometrium to cause the release of prostaglandin.

Pseudopregnancy (“false” pregnancy) often results when the dying embryo or its remnants, in the absence of endometritis, successfully blocks the release of prostaglandin at the time of MRP, around day 14. The mare will not return to heat at the expected time but her cycle will be prolonged due to the persistence of a corpus luteum in the absence of an embryo. The owner will think that the mare was pregnant because she doesn’t tease in or shows no signs of heat until after her expected return to heat; usually from a period of days to a week or two. In this scenario irregular estrus cycles may be a clinical indication of EEM.

Slow impending loss of the embryo beginning around the time of release of eCG from the endometrial cups (day 36), while the conceptus is still called an embryo, may cause pseudopregnancy of a different kind. This time the mare will not return to heat for months until after the endometrial cups are gone. The owner who does not have the mare checked for pregnancy by palpation during this time will have lost most of the breeding season.

What does the vet see with EEM?

When your vet palpates a mare that has undergone EEM, at day 16 to 18 after breeding, the mare will “feel” as if she is coming back into heat. The uterus will feel as if it has no tone or tubularity to it. This of course will depend on when exactly the embryo has started to die and undergo loss. If the mare is pseudopregnant (as described above) remnants or fluids from the embryo may be present and prevent the mare from returning to heat right away. Pregnancy may be difficult to assess in this case and a second examination may be required in a couple of days.

With the ultrasound the identification of the normal or dying embryo is readily identified. Normal embryos are spherical and produce a wellÄcircumscribed black image indicative of the embryonic vesicle, a collection of embryonic fluid. The embryonic vesicle grows very rapidly during day 10 to 15, therefore, measurement of a vesicle too small for the stage of gestation may indicate impending loss. Movement of the embryonic vesicle throughout the uterus, as seen with the ultrasound from day 10 to 15, ceases at day 16 to 17 when it fixes in one spot of the uterus. Continued movement of the embryo beyond this time of gestation usually indicates impending embryonic loss. The embryo itself, other than the fluid, is not visible with the ultrasound until day 21. The embryo’s beating heart can be noticed at approximately 24 days. Embryo disorientation, disruption of the spherical shape of the embryonic vesicle, and a decrease in vesicle size are all signs, viewed ultrasonically, of loss of the embryo.

How much EEM is there?

Until recently few reports on the incidence of EEM are available. Most studies of this problem include statistics on all pregnancy loss. These investigations were performed in the United States, Europe, Australia and New Zealand. Early studies of all pregnancy loss, including both embryonic and early fetal loss in the mare, estimated an incidence of 7 to 16 percent between days 20 to 90 of gestation. The incidence is even higher for known subfertile mares. Greater than 75% of all pregnancy losses detected by palpation per rectum occur during the embryonic period. The ultrasound has helped us pinpoint the majority of detectable pregnancy loss to the embryonic period, specifically between days 11 to 15 of gestation. A higher incidence of EEM may even exist prior to the earliest limit of detection with the ultrasound. Lower embryo recovery rates from known subfertile mares between day 7 to 10 post-ovulation support this theory. Loss of embryos from subfertile mares between day 2 or 4 and day 14 post-ovulation is estimated to be greater than 60 percent.

What causes EEM?

Many factors have been implicated as the causes of EEM. These factors can be broadly categorized as intrinsic or mare factors, extrinsic (environmental or management) factors or embryonic factors.

Intrinsic factors include things such as endometrial disease, progesterone deficiency, maternal age, lactation or foal-heat breeding.

Endometritis and periglandular fibrosis are two examples of endometrial disease. Breeding, foaling, and manipulation of the reproductive tract during a veterinary examination are procedures which normally challenge the defenses of the uterus against infection. An increased susceptibility to endometritis has been observed in certain chronically infected (subfertile) mares. Endometritis results in the loss of the embryo in three possible ways. Firstly, the increased number of inflammatory cells in the uterus, initiated by infection, may cause destruction of the embryo. Secondly, the organism causing the infection (usually a bacteria) may have a direct effect on the embryo. Thirdly, uterine irritation caused infection may cause the mare to return to heat, through the release of prostaglandin.

Periglandular fibrosis (scarring of the endometrium) is another endometrial disease that may cause EEM. The degree of fibrosis has been inversely related to the likelihood that a particular mare can carry a foal to term. Some studies have shown an increase in embryonic mortality in mares with severe fibrosis. The severity of fibrosis is a function of the mare’s age, the number foals she has produced over her lifetime, and the level of uterine infection she has been exposed to. Fibrosis essentially chokes-off endometrial glands, thereby preventing adequate nutritional support of the embryo. These types of mares may be good candidates for embryo transfer if another foal is desired.

Progesterone, as we stated above is important for pregnancy maintenance. Decreased progesterone in early pregnancy may be due to endometritis, failure of MRP or primary insufficiency of the corpus luteum. Although it has not been proven, primary luteal insufficiency is the reason some reproductively normal mares, who habitually experience EEM, are placed on supplemental progesterone therapy. Progesterone supplementation can be an expensive treatment regimen, therefore all other causes of EEM should be ruled out before a mare is placed on progesterone.

The age of the mare may be an indirect cause of EEM. Studies have shown that the incidence of EEM increases with increasing age. Generally, when a mare becomes older, she is less able to fight off uterine infection after each foaling or mating. Endometrial fibrosis is also more common in older mares. There is no critical age when these changes take place and certainly there are a lot of mares older than 10 to 15 years of age that produce foals fairly regularly.

Some studies of EEM have detected higher losses in lactating than dry mares. It has been theorized that the hormones involved with lactation or stress after foaling cause an upset of the normal endocrine pattern. More likely though, management of the mare including foaling management and nutrition after foaling play more of a role.

Mares bred at foal heat have been associated with a higher embryonic loss rate. There are however, conflicting reports on this subject. Obviously, poor foaling management leads to a poor uterine environment and increased chances of infection. Endometritis will definitely lead to higher EEM. Again the difference in observed embryonic losses is most likely due to management.

Extrinsic factors associated with EEM include stress, nutrition, climate, breeding management and techniques, and sire factors.

Stress has been shown to decrease progesterone levels in the mare, which in turn lead to increased embryonic losses. There are many factors that could contribute to stress and most of these are difficult to assess in the overall stress level of the mare. Stress may be related to pain, foaling, breeding, nutrition, transport, weather etc. A study conducted at Colorado, demonstrated that extended transport of the mare immediately after breeding had no significant effect on the level of embryonic loss.

Poor mare nutrition and body condition influence the rate of embryonic loss. A critical time of embryonic loss appears to be between day 25 to 30 for mares receiving poor nutrition.

The influence of climatic factors (season, light, temperature) definitely affect a mare’s reproductive cycle. However, the effect of these factors on embryonic loss is unknown. Heat stress has been demonstrated to be an important factor associated with high embryonic loss rates in cows in southern summer climates.

Breeding management is a very important factor associated with EEM. Breeding a mare either by natural cover or artificial insemination (AI), at the optimum time insures the proper timing of insemination in relation to when the mare ovulates. A good teasing and palpation program, to pinpoint imminent ovulation, goes a long way towards decreasing the number of times a mare is mated during her heat. Repeated matings without regard to when a mare ovulates greatly increases the degree of bacterial contamination the uterus is subjected to. This is especially important to chronically infected mares with an increased susceptibility to acquiring endometritis. The use of hormonal preparations to ensure ovulation, and semen extenders containing antibiotics to prevent bacterial overgrowth in the uterus, are also important in obtaining a successful mating in these mares. While AI generally provides a cleaner mating environment, it must be handled properly. Improper handling of collected semen, by exposing it to light, cold, heat, chemicals etc., prior to insemination will annihilate sperm. Aged and damaged sperm although capable of fertilizing an egg, may be a factor in early embryonic death. The use of frozen semen in mare breeding programs is limited by restrictions of certain breed registries. The effect of frozen semen on EEM is not known but approximately 50 % of live sperm numbers are lost after thawing. Again damaged sperm could possibly have an effect on EEM but no studies have been reported.

Embryo transfer to reproductively sound mares is a useful procedure to produce foals from donor mares with a history of EEM, especially due to endometrial fibrosis. However, studies have shown that embryo recovery attempts from subfertile mares may yield lower embryo numbers and smaller damaged embryos compared to mares of normal fertility. The transfer of healthy normal embryos to mares of normal fertility does not seem to increase the rate of embryonic loss, although data on large numbers of mares is not available.

Although it has never been demonstrated clearly, the use of certain sires may be associated with a higher rate of embryonic loss. This may be the result of infection of the mare at the time of breeding or the poor quality sperm that do not have much longevity in the mare’s reproductive tract.

Embryonic factors causing EEM are the result of abnormalities of the genetic makeup of the embryo. The incidence of genetic abnormalities of equine embryos has not been established but may be related to the improper timing of insemination resulting in fertilization of aged sperm and eggs. An older mare may produce an increased number of embryos with genetic abnormalities.

Prevention and management of EEM

Prevention of EEM begins with the manipulation of the intrinsic and extrinsic factors that are in the control of the veterinary practitioner. Improper teasing and breeding procedures and erroneous pregnancy diagnosis may contribute significantly to an apparent increase in EEM. However, there is little that can be done to prevent EEM due to embryonic factors.

Performance of a complete breeding soundness exam prior to the breeding season identifies barren mares with endometritis and periglandular fibrosis. While fibrosis is not amenable to treatment, therapy for endometritis and/or surgical correction of a mare with poor vulvar conformation could decrease her susceptibility to infection.

Proper management of foaling mares, i.e. providing a large comfortable clean stall and supervision at foaling, minimizes the risks of the mare acquiring an infection. Closure of a previously opened Caslick’s suture immediately after foaling prevents the aspiration of air and contaminants into the uterus. A mare that has had any difficulty foaling or has had a retained placenta should probably not be rebred on foal-heat. Also any mare with an abnormal discharge in the post foaling period, indicative of infection, should not be bred.

Once the breeding season has arrived, strict attention to hygiene at breeding whether it be by natural cover or AI, again helps to reduce contamination. The use of good teasing practices, regular palpations, induction of ovulation by hormonal therapy, and good breeding technique ensures timely mating. Proper timing of mating with ovulation is especially important in situations, such as a subfertile mare, where only one cover is desired.

Regular pregnancy examinations with an ultrasound at 14 to 21 days and 22 to 28 days ensures embryo viability and rules out pseudopregnancy and EEM. This allows early recognition of EEM in order that these mares may be rebred. Mares with a history of EEM but no identifiable problems may maintain an early pregnancy with supplemental progesterone therapy.

Attention to body condition of the mare and level of nutrition at breeding and especially between 20 to 30 days post breeding will help prevent EEM due to malnutrition.