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BREEDING WITH FROZEN SEMEN

Difference between Frozen Semen and Chilled Shipped Semen
Chilled shipped semen refers to semen which is cooled to approximately 4°C but not frozen. Frozen semen is frozen in liquid nitrogen to -196°C. Chilled semen is usually fertile 1-3 days after collection, while frozen semen has indefinite “shelf life” until thawed. Breeding mares with frozen semen can yield successful results but it is not for every mare. It requires thoughtful mare selection, extremely careful mare management, attention to detail, and proper handling of good quality semen. This article will discuss the use of, and concerns faced by, the mare owner choosing to breed their mare with frozen semen, hopefully helping you decide if it’s suitable for your mare and pocketbook. If you’d like more information on chilled shipped semen, please refer to our article on Breeding Your Mare with Shipped Semen.

Advantages of Frozen Semen
Breeding mares with frozen semen has all the advantages of artificial insemination (AI) with chilled shipped semen, namely: large selection of high quality individuals without the problem of shipping mares over great distances. Breeding to overseas stallions is readily available with frozen semen but not with chilled semen. Additionally, breeding with frozen semen eliminates some of the availability problems encountered with chilled semen: when there are more requests than semen available, a stallion’s competition schedule taking them away from breeding shed, stallion illness, problems with overnight shipping, or death of the stallion. Frozen semen does however come with its own set of challenges. Some of these challenges arise from the basics of frozen semen itself, others from how stallions are selected as sires and of the marketplace which has developed around this industry.

Fertility of Frozen Stallion Semen: Stallion Perspective
One challenge of frozen semen is that fertility rates may be somewhat lower in comparison to AI of fresh or chilled semen and live cover. There are many possible causes for the lower fertility of frozen semen.

From the stallion prospective, not all stallion semen survives the freezing process equally. As a very rough example, 25% of the stallion population freeze very well, 50% freezes acceptably, and 25% freeze very poorly or not at all. This isn’t a problem with the cattle frozen semen industry. When bulls are chosen as sires for frozen semen programs they are selected both on the basis of the potential they can pass to their offspring and how well their semen survives freezing. If the bull’s semen does not freeze well they are culled regardless of their genetic potential. Stallion selection on the other hand does not take the “freezability” of the stallion’s semen into much consideration. If a stallion performs well (for whatever criteria is deemed in demand) their semen will probably be frozen if permitted by the breed registry.
As a consequence not all frozen semen is of high quality. This makes it important to know the post thaw characteristics of semen prior to purchase. This topic is addressed below under "Questions to Ask the Stallion Manager."

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“Breeding Dose”
Another challenge of frozen equine semen is that the breeding dose- or the minimum number of progressively motile sperm that must be used to reasonably ensure pregnancy is not standardized industry wide.
With chilled shipped semen the most commonly accepted breeding dose is 500 million progressively motile sperm. Progressively motile sperm are the sperm moving in a forward direction - the ones generally accepted as being the only ones capable of making an embryo. This is in contrast to sperm swimming in circles, or just wiggling without forward movement.

The breeding dose for frozen semen reported has varied widely from 300 million to 1 billion total sperm, or 250 to 600 million progressively motile sperm. Besides just the magnitude of difference in numbers (250 million vs. 1 billion) these are very significant differences, because total sperm refers to all sperm both progressively motile and infertile dead or circling sperm. So, if a breeding dose contains 1 billion total sperm, but it only has 10% progressive motility, then the number of sperm actually capable of fertilization is only 100 million progressively motile sperm - below all current recommendations. Currently the only progress being made in this area appears to come from government regulation in The Netherlands where it’s required that a breeding dose of frozen semen contain a minimum of 300 progressively motile sperm.

Now bear in mind, pregnancy has been achieved with as little as 1 million sperm using specialized AI techniques discussed later (deep horn insemination or video hysteroscopic insemination), but the likelihood of success is below what many would consider acceptable in light of the costs.

Mare Management Perspective
Careful selection of fertile mares for frozen semen programs is often more important than for other methods of breeding because of the price of frozen semen and mare management. Greatest success is usually seen in mares less than 6 years old. In older mares, between 6 to 16 years of age, success is more likely when a history of reproductive problems is not present. Mares 16 or older should be reproductively evaluated prior to entering into a frozen semen program. Older mares may have defective eggs or may have more of a problem clearing the uterus of excess fluid following breeding. This delay in uterine clearance can be a more common aftereffect following AI with frozen semen. Problem” or infertile mares may require several cycles and breedings to become pregnant. From a management perspective, breeding with frozen semen may require intense mare management: up to 3-4 mare exams per day when nearing ovulation or careful attention to timing of exams, breeding, and timing of ovulation induction (discussed under AI protocol below).

Mare Examination and insemination: Palpation and Ultrasound
Breeding mares with frozen semen involves ultrasound and rectal palpation exams to follow a mare’s cycle until she has a mature follicle. At this stage the breeding management is dictated by the AI protocol decided upon as discussed below. Exams continue every 6 to 8 hours or timed with insemination 24 & 40 hours following administration of ovulation induction agents.

Induction of Ovulation
Pregnancy is best achieved by breeding mares as few times as possible within a few hours of ovulation. In order ensure ovulation occurs close enough to AI, mares are injected with HcG (human chorionic gonadotropin) or deslorelin, hormones which will stimulate a mature follicle to ovulate. HcG will usually result in ovulation by 48 hours, deslorelin by 40 hours.

AI Protocol for Frozen Semen
Choices for breeding mares with frozen semen are influenced by semen packaging (0.25, 0.5, 5.0 ml straws), semen characteristics (total number of sperm, percent progressively motile sperm), the number of straws (1, 4, 8) recommended by the producer as a breeding dose, and costs of semen and mare management. Protocols usually involve breeding with one dose (“traditional” method below) or two doses (timed insemination with half or full doses), all yielding similar fertility results, but with significantly different requirements for mare exams. Depending on the above we offer the following successful breeding protocol:

Timed insemination using 2 half doses at specific timed intervals.
  • Requires daily mare examinations (when approaching ovulation)
  • 1 full breeding dose packaged as multiple straws per dose
  • 400 million progressively motile sperm per half a breeding dose
 
Timed insemination using 2 full doses at specified timed intervals.
  • Requires daily mare examinations (when approaching ovulation)
  • Use of 2 full breeding doses
  • Sufficient progressively motile sperm per breeding dose
 
“Traditional” frozen insemination technique a few hours after ovulation
  • Requires much closer mare management: exams every 6-8 hours when approaching ovulation
  • 1 full breeding dose
  • Sufficient progressively motile sperm per dose

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AI Techniques: Deep Horn, Video Hysteroscopy
Insemination techniques have changed with the use of frozen semen. Breeding with chilled shipped semen entails deposition of the semen into the body of the uterus. This is also done with frozen semen but techniques have evolved using much smaller volumes deposited at the very tip of the uterine horn on the oviductal papillae (intersection of the oviduct and uterus) closer to where the egg is released from the follicle. This technique referred to as Deep Horn insemination has also been accomplished with the use of a flexible video endoscope (video hysteroscopy). Hysteroscopy allows the visualization of the papillae and deposition of very small volumes of semen at the site.

Deep Horn AI is favored by many when using a small volume breeding dose or with the use of specialized “AI guns” when the breeding dose contains multiple straws. Fertility has been reported in some reach to be higher with Hysteroscopy than deep horn AI with frozen semen containing reduced sperm numbers.

Deep horn and hysteroscopic AI has resulted in pregnancy with breeding doses as low as 50 to 150 million progressively motile sperm. This low numbers were stallion dependent and it was not possible to improve pregnancy rates of stallions with frozen semen of poor fertility.
For more information read Endoscopic Assisted A.I.

Frozen Semen Marketplace: Smaller Breeding Doses
Although it is has been shown that it’s possible to breed mares with as little as 1 million sperm, success is more likely when using 800 million progressively motile sperm. Previous to the introduction of frozen, stallion breeding rights were sold with a live foal guarantee or ability to rebreed to the stallion without additional cost aside from collection and shipping fees. The goals for mare and stallion owner were the same, a foal with a minimum number of breedings.
Unfortunately, with frozen semen the goals of mare and stallion owner are not the same anymore. Mare owners still want the same, but stallion owners want as many breeding doses from a single semen collection as possible. Because pregnancies have been achieved with very low sperm numbers, and frozen sperm is usually sold by the breeding dose, the result has been a trend to get as many breeding doses from a semen collection and for breeding doses to become smaller. This trend will undoubtedly continue until the marketplace changes.

Questions to Ask Stallion Management
Sources of frozen semen include both farms where the stallion is standing at stud as well as semen brokers. It is important to inquire about the stallion’s fertility statistics with frozen semen (conception rates, pregnancy rates etc.) as well as post thaw semen motility. At the very minimum post thaw progressive motility should be known.

Other relevant questions to ask are: number of mares bred and number of mares pregnant, number of breedings per cycle, number of cycles per pregnancy. The answers to many of these questions can be difficult to compile because many of the inseminations are not under direct control of stallion management. At the very least these statistics can indicate how closely a farm is monitoring the fertility of their frozen semen, at best it will give you some idea of how successful you may be with a particular stallion.

If the farm does not have any firm statistics have your veterinarian open a line of communication with the farm manager or veterinarian before you purchase the semen or choose another stallion.
For more information please see Questions to Ask the Staliion Manager.

EVA Status of Stallion (and Mare)
Equine Viral Arteritis (EVA) is a disease which can be transmitted by live cover, and AI of both chilled and frozen semen. For more information see click here.

It is fairly simple to control with the following protocol. Ask the stallion owner about the EVA status of the stallion. Stallions can be vaccinated against this disease, but must test negative before vaccination. If breeding to an EVA positive stallion, vaccinate your mare at least 30 days before breeding. These mares should be tested prior to vaccination and should be isolated from direct contact of non vaccinated horses. If breeding to an EVA negative stallion no action is necessary. If EVA status of the stallion is unknown the risk of this serious disease to your mare is unknown.

Frozen semen allows you to choose from many stallions without the trouble, expense, and stress of shipping your mare as well stallions overseas or maybe even deceased. Breeding with frozen semen requires more intensive mare management and frequently more expense. Unlike other breeding scenarios it is usually sold on the per dose basis rather than live foal guarantee. It’s a great advance for horse breeding but requires experience, knowledge, and tremendous attention to detail. If you would like more in depth information on equine reproduction, chilled and frozen semen, please don’t hesitate to contact our office.

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Friedman Veterinary Service & The Oregon Equine Reproduction Center
P.O. Box 695  |   Lake Oswego, OR 97034  |  (503) 675-0757  |   ron@oregonequine.com
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