Basics for breeding mares with cooled or frozen semen (Proceedings)

Advanced reproductive technologies such as cooled semen, frozen semen, embryo transfer and gamete inter fallopian tube transfer (GIFT) have given horse owners choices and freedom. Mares can be bred at home with semen collected from stallions that live anywhere in North America, Europe or Australasia. Stallions can compete during the breeding season while mares are bred with their previously frozen semen. Embryos can be collected from performance mares between competitions. The genetics of valuable infertile mares can be maintained through GIFT, a procedure that involves the aspiration of eggs from follicles on the donor mare's ovaries. The eggs are then placed in a fertile recipient mare's oviduct after which she is inseminated with semen. But as with all choices, these technologies come with a price. To be successful, a higher level of veterinary expertise is needed, mares need to be examined more often, pregnancy rates are lower and costs are higher. Equine semen does not tolerate cooling, freezing and the manipulation needed for processing as well as other species. So, more money will be spent to obtain somewhat lower pregnancy rates then that seen with natural breeding or breeding with fresh semen by artificial insemination. So before you sign a breeding contract for the stallion of your dreams, read the following so that you do not naively enter into the breeding game.

The ins and outs of breeding mares with cooled semen

To breed mares successfully with cooled semen all parties involved, mare owner, stallion manager and veterinarians, need to cooperate when coordinating the semen shipments with the timing of the mare's ovulation. Before shipping semen, the owner should clarify several points with the stallion manager.

     • The cost of stallion collection

     • The cost of preparing the semen for shipment, the number of collections provided gratis (if any), the cost of shipping semen tanks by air, and when and how the semen tanks must be returned.

     • The days of the week the stallion is collected

     • Times during the breeding season when the stallion will not be available

     • The number of days notice that the stallion manager needs before the semen shipment

     • The latest time one can call to obtain semen (for example-one must call by 9am to receive semen by the next day)

     • The longevity of the semen-does it live in the tank for 12, 24 or 36 hours

     • First-cycle conception rate of the stallion

     • The method of air transport used (same-day air or overnight shipment.

     • Number of times the mare can be bred if she does not conceive (is the contract limited to 1, 2 or 3 years)

     • The breed registry requirements, and the number and timing of post-insemination clinical (pregnancy) examinations must be established.

Management of mares for breeding has changed in the last 15 years as more mares are bred on farms where there are no means for teasing the mare to determine where she is in her estrous cycle. Stabling a mare at a veterinary clinic or farm where the veterinarian visits daily, saves money on veterinary travel fees. Furthermore, many of these facilities have a stallion to tease the mare to determine when she is in heat, thereby, limiting the number of examinations. Once the mare is in heat, she will need to be examined at least every other day and bred within 24 hours of ovulation.

Pregnancy rates are highest when mares are bred within the 24 hours that precede ovulation using semen of high fertility. The quality of the semen is of paramount importance: stallions with low fertility usually have much lower first cycle pregnancy rates than those with high inherent fertility. In addition, the handling of the semen is critical; failure to prepare it correctly as well as poor subsequent handling at the mare end can make the process very disappointing. Timing of the breeding with the ovulation can be difficult especially if the stallion is collected only 3 times a week. Ovulation can be induced with drugs such as hCG or deslorelin, however the window from injection of the drug to ovulation varies. Mares may ovulate as quickly as 24 hours, as late as 48 hours after administration of hCG or they may not respond at all. The window from injection of deslorelin to ovulation is tighter than that of hCG with most mares ovulating between 42 and 48 hours, however, it costs about 2.5 times more than hCG.

There are standards that semen needs to meet to be considered of adequate quality. There is controversy on what is an adequate semen dose for shipment. The old rule of thumb was a dose of semen should contain a minimum of 500 million progressively motile sperm with at least 30% of the sperm being progressively motile. However, some stallions have acceptable pregnancy rates with doses of 250 million progressively motile sperm. Many farms are now shipping a dose of 250 million sperm/bag. The minimum dose will vary between stallions so it is prudent to contact stallion managers if one is experiencing low first cycle pregnancy rates in reproductively normal mares. Each time the mare is bred with cooled semen, it should be examined carefully after it has been warmed for a minimum of 3 minutes. If it is of poor quality the stallion manager or veterinarian for the stallion should be notified. In a study from Texas A and M, reproductive records from all mares bred with cooled semen were reviewed. The group showed that first cycle pregnancy rates were approximately 65% if the mare was bred with a minimum of 500 progressively motile sperm. However, first cycle pregnancy rates dropped to less than 30% if mares were bred with a lower number of motile sperm. We recommend that the stallion manager place at least 1 billion progressively motile sperm in the package before shipment so that 24 hours later when the mare owner receives the semen, there is at least 500 million progressively motile sperm in the dose.

After insemination, the reproductive tract of the mare should be examined daily until she ovulates. If she does not ovulate within 24 hours she should be bred a second time. Age of the mare may affect her ability to get pregnant. Mares that are more than 10 to 12 years old when they are bred for the first time may retain fluid in the uterus after breeding because their cervix does not open properly. When semen is deposited in the uterus, the mare responds with an inflammatory response because the semen is recognized as a foreign substance. Reproductively normal mares clear the fluids that are associated with the inflammation within 12 to 18 hours of mating, those that do not, accumulate fluid. The inflammatory products in the fluid are harmful to the uterine lining as they have a low pH (acid). Your veterinarian can identify if your mare has a cervical problem before she is bred or at the time of breeding. Mares that have an incompetent cervix should have their uterus lavaged with saline or lactated ringers 4 to 8 hours after breeding to remove the fluid before it causes significant damage to the uterine lining.

Trials and tribulations of breeding with frozen semen

Breeding a mare with frozen/thawed semen can result in a lower first-cycle pregnancy rate than if the mare was bred with either fresh or cooled semen. Pregnancy rates of 0 to 70% /estrous cycle have been reported with an average pregnancy rate/cycle of 35 to 40%. Pregnancy rates are highest when a full dose of frozen semen from a stallion of known fertility is placed in a young mare of known fertility. Mares that are more than 14 years of age when they are bred with frozen semen have a mean reported pregnancy rate of about 20%. It is more costly to breed mares with frozen semen because the mare needs to be inseminated within 8 hours of ovulation. Therefore, many of these mares need to be examined more than twice daily IF they are to be bred with only one dose of semen after ovulation.

Points that must be clarified before buying frozen semen:

     • What is the first cycle conception rate of the stallion when breeding with the frozen semen

     • What is the cost of semen

     • What constitutes a dose- number of straws

     • Is semen sold by the dose or by the straw

     • How are the straws to be thawed?

     • Are nitrogen tanks available to store the semen

     • What is the rental fee for the dry shipper

     • What paperwork does the breed registry require

Semen is now typically frozen in one half ml straws. A dose will range from 4 to 8 straws containing a total of 200 to 400 million progressively motile sperm and is dictated by the fertility of the horse. It is not uncommon for owners to request that a partial dose be used for insemination so the mare will need to be inseminated after and within 8 h of ovulation. Semen can be sold by the dose OR by the straw. As the price for a straw is less than that of a full dose, many owners elect to buy only a few straws. However, in most cases one is more likely to get a mare pregnant on the first cycle using a full dose of semen than with a partial dose. Most mares are now bred with flexible, long, insemination pipettes deep in the horn ipsilateral to the dominant follicle.

Mares can be bred once before and once after ovulation if a full dose (divide the dose in half) or more than one dose is provided. If a partial dose or only one dose is available mares are bred directly after ovulation. The number of reproductive examinations conducted each day during estrus varies among practitioners. Some veterinarians still evaluate the reproductive tract every 6 to 8 h however, management protocols that include the use of either hCG or Deslorelin enable the veterinarian to achieve good pregnancy rates by palpating the reproductive tract only once every 12 to 24 hours. The ovulatory agent is administered after the follicle reaches 35 mm in diameter and is medium soft. If hCG is used to induce ovulation, the mare is first bred 24 hours after she was given hCG and then a second time at 36 hours after hCG, if she has ovulated by 36 hrs. If she has not ovulated at 36 hours, the veterinarian may elect to breed after ovulation has been confirmed. In the latter instance the mare should be examined every 8 hours from hour 36 (time 0 = time when hCG was given) to ovulation. If deslorelin is used as the ovulatory agent, the mare is first bred at 36 hours and then again at 48 hours. For example, the mare is examined at 5 pm when it is first noted that the follicle has reached 35 mm in diameter. She is then given hCG and bred at 5 or 6 pm the next day. The following morning the mare is re-evaluated at 6 am to determine if she has ovulated, if she has ovulated then the 2nd dose of semen is inseminated. A similar protocol would be followed with Deslorelin except that the mare is first bred 36 h after administration of the drug. This author prefers using Deslorelin as the ovulatory agent because the window in which the mare ovulates after administration is smaller than after hCG, 40 to 48 h versus 24 to 48 h

Some mares will exhibit an acute, persistent intra-uterine inflammatory response after being bred with frozen semen. Because of this consequence, many veterinarians will lavage the uterus 4 to 8 h after the mare is bred. This author will evaluate the mare approximately 8 h after she is bred to determine if the mare has ovulated and if there is either an abnormal degree of uterine edema or free intra-uterine fluid. If either exists the uterus is irrigated with lactated ringers solution. If the mare requires a second insemination, she will be bred 2 to 4 h after the uterine irrigation.