WHAT IS EMBRYO TRANSFER?

Embryo transfer is the procedure in which a number of embryos (fertilized eggs) are removed from one nigh-value female (the embryo donor) and transferred into many less-valuable females (the embryo recipients). The recipients complete the pregnancies and raise the offspring of the valuable donor. The donor will return to normal estrus (during the breeding season) and will rebreed, or can be used again to produce embryos in about 6 weeks.

Normally, the embryo donors are brought into a synchronized heat using progesterone hormones. Hormones are also given to increase the number of unfertilized eggs that are ovulated during the heat (superovulation). The donor is bred and the eggs become fertilized and recovered using surgical procedures about a week later – 6 day old embryos. These embryos then must be transferred into recipient animals whose uterus is at the same hormone stage as the uterus of the donor that provided the embryos – about 6 days following a previous heat. The embryos trick the recipient into believing she is pregnant from her own heat - which prevents the next heat from occurring and causes a normal pregnancy to develop and so carrying the donor’s embryos to tern.

 WHY TRANSFER EMBRYOS?

There are many reasons to transfer embryos from a donor to a group of recipients. In most cases its because the donors are more valuable than other sheep and we use ET to increase the numbers of offspring that the valuable donor could produce in a given period of time. Reasons that a donor might be considered more valuable include: health, production record, pedigree, availability (genetics that cannot be purchased in any other way) etc. Embryos are also the most efficient method for moving genetics over long distances.

Properly managed, embryos are free of production diseases common to sheep and goats. The International Embryo Transfer Society ( IETS) specifies handling procedures that are recognized by all regulatory agencies that, if followed correctly, reduce the risk of disease transmission using ET. These include washing the embryos in enzymes during the transfer procedure. Unfortunately, the one disease that we are not yet certain of is scrapie – but evidence is growing that scrapie is not transmitted using ET. So embryos, cleaned by washing and then transferred into disease free recipients produce kids or lambs of high health status. ET is effective to "clean up" positive flocks, and to move embryos between farms, countries or continents where disease control is a concern.

Because of the clean nature of embryos, no health testing of donors is required before the embryos are recovered. This means that genetics that could not be moved as semen (because the bucks or rams did not pass the health testing) can still be marketed.

By increasing the normal number of ovulations (superovulation), ET can be used to greatly increase the number of offspring born per donor in a short period of time. This can be particularly useful to recover investment in valuable animals. Donors can undergo ET repeatedly, easily two or three times a season, and then return to normal breeding. We have many examples where over 30 offspring have been born to one donor in a single year.

ET can be useful to maintain production from better animals that can no longer raise kids or lambs, for example, that older ewe that has developed mastitis or doe with advanced CAE.

By freezing embryos, ET can be used to store embryos for future use and to move genetics between farms and regions.

 Embryos have been an effective method of marketing cattle genetics - as demonstrated by the international demand. Most potential purchasers don't want to wait for embryos to be produced. By having embryos available "on-the-shelf" and promoted internationally, we should be able to increase genetic sales without seriously altering our breeding programs - great value added to normal production.

WHAT WOULD BE THE TYPICAL SUCCESS OF ET?

To accurately evaluate an ET program, it is necessary to evaluate each of the procedures that make up the program: superovulation program, embryo recovery surgery, the number of transferable-quality embryos recovered, the number of recipient pregnancies established and the number of offspring produced per embryo. Ultimately, the only thing that matters to the producer is the number of offspring born, but it is important to be able to identify what parts of a program have worked well or failed.

Superovulation. Superovulation is done using twice-daily injections of hormones for three days in advance of the donor's heat. The success of superovulation is determined during the surgery – ovulation sites on the ovaries (Corpora Lutea - CLs) are counted. Results range from none to over 30 with a typical response being about 12 in prolific breeds and about 8 in terminal sire breeds of sheep and about 8 - 10 in most North American breeds of dairy and meat goats. Results are affected by age, season, body condition, breed, nutrition, lactation and stress. Variation in superovulation is one of two great unknowns when starting an ET program.

Embryo recovery. The typical success of the surgical recovery of embryos is about 75% of the CLs counted, but can range from less than 40% to almost 100 %. Embryo recovery is affected by donor age, health of uterus, number of lambings/kiddings and experience of the surgical team.

Transferable quality embryos. Not all embryos recovered are "transferable" - capable of producing pregnancies. There is no sense in using good recipients to carry poor quality embryos. As a result, embryos are graded for their probability of producing a pregnancy/offspring. Embryos are graded based on their stage of development ( are they at the developmental stage they should be 6 days after breeding?) and their "quality". Quality is affected by the amount of physical damage found – some degenerate or are damaged by the handling procedures. Generally, about 80% of embryos recovered are at the correct stage of development and are of an acceptable quality. The most common reason for classifying embryos as not transferable is finding them unfertilized. In some cases only a few of the embryos may fail to be fertilized. In other cases none may be fertilized. Fertilization failure is one of the most common causes for failure of an ET program and will be explained further.

Pregnancy and lambing rates. Generally, transferring good quality fresh embryos into good recipients should result in a pregnancy rate of about 70%. This can range from none to almost 100%. Most programs put two or three embryos into each recipient. Not all pregnancies will be carrying live lambs from all embryos received. Generally, with fresh embryos, the chance of each good-quality embryo transferred to good recipients producing an offspring is 70%, but again can range well below to over 80%. Generally, one should be able to expect an average of between 3 to 7 live offspring born per donor flushed when good embryos are transferred into well-managed recipients. .

 IF EMBRYO RECOVERY AND TRANSFER ARE SURGICAL, IS THERE RISK TO MY ANIMALS?

Yes, there is risk in any surgical procedure. But the risk is very small.

Embryo recovery requires complete general anesthetic and full abdominal surgery. The uterus is exposed and punctured at two sites. Embryos are transferred laparoscopically. This is a minor-surgical procedure but still requires anesthesia and abdominal invasion. So the risks include anesthetic death, damage to the reproductive tract (future fertility) and injury to other abdominal organs.

But when the animals are properly selected and managed and with good equipment and facilities and an experienced team, the risk is very small.

In the past three years, we have been involved in almost 500 embryo recovery surgeries. We are aware of (and people do tell us) two donors that failed to breed following the surgery - which frankly might have been caused by other reasons. We experienced two tragic anesthetic deaths - both in Boer goats, who we now know have a familial-breed reaction to the anesthetic used at that time. In the same period we have transferred embryos into thousands of recipients - both sheep and goats. We had two die of clostridial disease and four from other abdominal injuries - both appear to be associated with diet and improper starvation prior to the surgery.

We have learned from these experiences and those reported by others. As a result, what little risk there is, is reducing all the time - better equipment, better instructions to producers, newer anesthetics, more experience. Still, insurance can be considered by producers.

We have more experience with ET in these species than any other group we are aware of in North America.

Results from ET programs vary greatly. Some programs are complete failures while others result in success beyond that expected. Most fall within the ranges and averages described. Failure of superovulation and failure of fertilization are the two reasons most programs fall below expectations. About 25% of donors fail to respond to superovulation. Some of those will again fail to respond when repeated. Others will respond normally. This high variability in results also occurs in cattle ET programs and is a priority for many research programs.

Successful fertilization requires close attention to breeding superovulated donors. The donor’s blood hormone levels are abnormal due to the large number of ovarian follicles activated by the superovulation drugs. Those hormones alter signals of heat and can affect the transport of semen through the cervix to the site of fertilization. In addition, the ram or buck is often expected to fertilize 20 or more eggs - instead of the normal 2 to 5. There is a strong relationship between the success of superovulation and fertilization failure: donors that successfully produce many CLs (20 or more) have a higher proportion of those as unfertilized.

Donor breed. Prolific and maternal-line breeds respond better to superovulation drugs than terminal sire breeds.

Donor age. As with normal breeding, more ovulations are produced during the peak years of reproduction – typically 2 to 5 years. Younger or older donors generally produce fewer embryos. Ewe lambs or young does have been reported to produce embryos with lower survival rates.

Season. There is a seasonal reduction in embryos obtained from superovulated donors and from pregnancies established in recipients. One can estimate generally, about a 10% drop – mostly in the months of April and May. As well, season has a great effect on the ability of the bucks and ram to fertilize superovulated donors.

Stress. All goat and sheep farmers should know that stress reduces fertility. Stress will reduce the success of an ET program as well. The times most seriously affected by stress are at breeding of the donors, the time of estrus in the recipients, and the first two weeks following the ET in the recipients. Goats are far more affected by stress than are sheep. Disease creates a chronic stress than can reduce results. Moving donors for breeding is not recommended. Moving donors for ET is not affected by stress - as the pregnancy is already established and the eggs are to be removed. Moving sheep recipients is routinely done, but moving goat recipients may reduce pregnancy rates.

Body condition. Donors in particular should be in good body condition – ideally 3 to 3.5. Recipients must also be in good condition

Diet. Well-conditioned animals do not benefit from nutritional flushing. Poorly conditioned animals will benefit. Flushing should focus on energy not protein. Very high protein diets are associated with reduced embryo quality and increased early embryonic death in recipients. Diets high in legume might also contain phytoestrogens (plant estrogens) that are also associated with reduced fertilization and increased embryonic death. In my view most people get carried away with nutritional concerns – trying to insure a successful program. Selecting well-conditioned animals, in good health and on a sensible diet of good quality forage with a suitable grain supplement and adequate mineral is best.

Donor health. ET can be used to salvage genetics from donors affected by disease. Results suffer as body condition declines. Serious illness drastically reduces superovulation results.

Breeding program. Normally we recommend one ram or buck to one or two superovulated donors. Superovulated ewes come into heat earlier than normal within 18 to 24 hours of pessary removal. Males are introduced for 36 hours or are hand bred during that period. GNRH is administered to help to insure successful ovulation. Laparoscopic AI is often used to overcome reduced fertilization. Laparoscopic AI has shown most benefit when young donors and/or young males are used in an ET program.

Drug handling. Mishandling easily alters drugs used for superovulation – particularly temperature. Results will suffer. Directions must be followed closely. Drugs must be administered every 12 hours for three days and must be given subcutaneously (in the fat under the skin).

Number of embryos transferred. Pregnancy and embryo survival rates are highest when two or three embryos (in contrast to one) are transferred to each recipient.

Recipient selection and care. Selecting good recipients is critical to success. It is a mistake to believe that any old ewe or doe will do. Recipients should be young (2-4 yr.), reproductively proven, in good condition and in good health. They are the ones doing all the work of accepting the embryos and carrying the pregnancy. They should be free of diseases of concern to insure the birth of clean offspring.

Producers must have realistic expectations and understand the risks inherent in embryo programs. Many of the common causes of low success can be prevented by good management.

WHAT IS EMBRYO TRANSFER?

Embryo transfer is the procedure in which a number of embryos (fertilized eggs) are removed from one nigh-value female (the embryo donor) and transferred into many less-valuable females (the embryo recipients). The recipients complete the pregnancies and raise the offspring of the valuable donor. The donor will return to normal estrus (during the breeding season) and will rebreed, or can be used again to produce embryos in about 6 weeks.

Normally, the embryo donors are brought into a synchronized heat using progesterone hormones. Hormones are also given to increase the number of unfertilized eggs that are ovulated during the heat (super ovulation). The donor is bred and the eggs become fertilized and recovered using surgical procedures about a week later – 6 day old embryos. These embryos then must be transferred into recipient animals whose uterus is at the same hormone stage as the uterus of the donor that provided the embryos – about 6 days following a previous heat. The embryos trick the recipient into believing she is pregnant from her own heat - which prevents the next heat from occurring and causes a normal pregnancy to develop and so carrying the donor’s embryos to tern.

 WHY TRANSFER EMBRYOS?

There are many reasons to transfer embryos from a donor to a group of recipients. In most cases its because the donors are more valuable than other sheep and we use ET to increase the numbers of offspring that the valuable donor could produce in a given period of time. Reasons that a donor might be considered more valuable include: health, production record, pedigree, availability (genetics that cannot be purchased in any other way) etc. Embryos are also the most efficient method for moving genetics over long distances.

Properly managed, embryos are free of production diseases common to sheep and goats. The International Embryo Transfer Society ( IETS) specifies handling procedures that are recognized by all regulatory agencies that, if followed correctly, reduce the risk of disease transmission using ET. These include washing the embryos in enzymes during the transfer procedure. Unfortunately, the one disease that we are not yet certain of is scrapie – but evidence is growing that scrapie is not transmitted using ET. So embryos, cleaned by washing and then transferred into disease free recipients produce kids or lambs of high health status. ET is effective to "clean up" positive flocks, and to move embryos between farms, countries or continents where disease control is a concern.

Because of the clean nature of embryos, no health testing of donors is required before the embryos are recovered. This means that genetics that could not be moved as semen (because the bucks or rams did not pass the health testing) can still be marketed.

By increasing the normal number of ovulations (super ovulation), ET can be used to greatly increase the number of offspring born per donor in a short period of time. This can be particularly useful to recover investment in valuable animals. Donors can undergo ET repeatedly, easily two or three times a season, and then return to normal breeding. We have many examples where over 30 offspring have been born to one donor in a single year.

ET can be useful to maintain production from better animals that can no longer raise kids or lambs, for example, that older ewe that has developed mastitis or doe with advanced CAE.

By freezing embryos, ET can be used to store embryos for future use and to move genetics between farms and regions.

 Embryos have been an effective method of marketing cattle genetics - as demonstrated by the international demand. Most potential purchasers don't want to wait for embryos to be produced. By having embryos available "on-the-shelf" and promoted internationally, we should be able to increase genetic sales without seriously altering our breeding programs - great value added to normal production.

WHAT WOULD BE THE TYPICAL SUCCESS OF ET?

To accurately evaluate an ET program, it is necessary to evaluate each of the procedures that make up the program: super ovulation program, embryo recovery surgery, the number of transferable-quality embryos recovered, the number of recipient pregnancies established and the number of offspring produced per embryo. Ultimately, the only thing that matters to the producer is the number of offspring born, but it is important to be able to identify what parts of a program have worked well or failed.

Super ovulation. Super ovulation is done using twice-daily injections of hormones for three days in advance of the donor's heat. The success of super ovulation is determined during the surgery – ovulation sites on the ovaries (Corpora Lutea - CLs) are counted. Results range from none to over 30 with a typical response being about 12 in prolific breeds and about 8 in terminal sire breeds of sheep and about 8 - 10 in most North American breeds of dairy and meat goats. Results are affected by age, season, body condition, breed, nutrition, lactation and stress. Variation in super ovulation is one of two great unknowns when starting an ET program.

Embryo recovery. The typical success of the surgical recovery of embryos is about 75% of the CLs counted, but can range from less than 40% to almost 100 %. Embryo recovery is affected by donor age, health of uterus, number of lambings/kiddings and experience of the surgical team.

Transferable quality embryos. Not all embryos recovered are "transferable" - capable of producing pregnancies. There is no sense in using good recipients to carry poor quality embryos. As a result, embryos are graded for their probability of producing a pregnancy/offspring. Embryos are graded based on their stage of development ( are they at the developmental stage they should be 6 days after breeding?) and their "quality". Quality is affected by the amount of physical damage found – some degenerate or are damaged by the handling procedures. Generally, about 80% of embryos recovered are at the correct stage of development and are of an acceptable quality. The most common reason for classifying embryos as not transferable is finding them unfertilized. In some cases only a few of the embryos may fail to be fertilized. In other cases none may be fertilized. Fertilization failure is one of the most common causes for failure of an ET program and will be explained further.

Pregnancy and lambing rates. Generally, transferring good quality fresh embryos into good recipients should result in a pregnancy rate of about 70%. This can range from none to almost 100%. Most programs put two or three embryos into each recipient. Not all pregnancies will be carrying live lambs from all embryos received. Generally, with fresh embryos, the chance of each good-quality embryo transferred to good recipients producing an offspring is 70%, but again can range well below to over 80%. Generally, one should be able to expect an average of between 3 to 7 live offspring born per donor flushed when good embryos are transferred into well-managed recipients. .

 IF EMBRYO RECOVERY AND TRANSFER ARE SURGICAL, IS THERE RISK TO MY ANIMALS?

Yes, there is risk in any surgical procedure. But the risk is very small.

Embryo recovery requires complete general anesthetic and full abdominal surgery. The uterus is exposed and punctured at two sites. Embryos are transferred laparoscopically. This is a minor-surgical procedure but still requires anesthesia and abdominal invasion. So the risks include anesthetic death, damage to the reproductive tract (future fertility) and injury to other abdominal organs.

But when the animals are properly selected and managed and with good equipment and facilities and an experienced team, the risk is very small.

In the past three years, we have been involved in almost 500 embryo recovery surgeries. We are aware of (and people do tell us) two donors that failed to breed following the surgery - which frankly might have been caused by other reasons. We experienced two tragic anesthetic deaths - both in Boer goats, who we now know have a familial-breed reaction to the anesthetic used at that time. In the same period we have transferred embryos into thousands of recipients - both sheep and goats. We had two die of clostridial disease and four from other abdominal injuries - both appear to be associated with diet and improper starvation prior to the surgery.

We have learned from these experiences and those reported by others. As a result, what little risk there is, is reducing all the time - better equipment, better instructions to producers, newer anesthetics, more experience. Still, insurance can be considered by producers.

We have more experience with ET in these species than any other group we are aware of in North America.

Results from ET programs vary greatly. Some programs are complete failures while others result in success beyond that expected. Most fall within the ranges and averages described. Failure of super ovulation and failure of fertilization are the two reasons most programs fall below expectations. About 25% of donors fail to respond to super ovulation. Some of those will again fail to respond when repeated. Others will respond normally. This high variability in results also occurs in cattle ET programs and is a priority for many research programs.

Successful fertilization requires close attention to breeding super ovulated donors. The donor’s blood hormone levels are abnormal due to the large number of ovarian follicles activated by the super ovulation drugs. Those hormones alter signals of heat and can affect the transport of semen through the cervix to the site of fertilization. In addition, the ram or buck is often expected to fertilize 20 or more eggs - instead of the normal 2 to 5. There is a strong relationship between the success of super ovulation and fertilization failure: donors that successfully produce many CLs (20 or more) have a higher proportion of those as unfertilized.

Donor breed. Prolific and maternal-line breeds respond better to super ovulation drugs than terminal sire breeds.

Donor age. As with normal breeding, more ovulations are produced during the peak years of reproduction – typically 2 to 5 years. Younger or older donors generally produce fewer embryos. Ewe lambs or young does have been reported to produce embryos with lower survival rates.

Season. There is a seasonal reduction in embryos obtained from superovulated donors and from pregnancies established in recipients. One can estimate generally, about a 10% drop – mostly in the months of April and May. As well, season has a great effect on the ability of the bucks and ram to fertilize superovulated donors.

Stress. All goat and sheep farmers should know that stress reduces fertility. Stress will reduce the success of an ET program as well. The times most seriously affected by stress are at breeding of the donors, the time of estrus in the recipients, and the first two weeks following the ET in the recipients. Goats are far more affected by stress than are sheep. Disease creates a chronic stress than can reduce results. Moving donors for breeding is not recommended. Moving donors for ET is not affected by stress - as the pregnancy is already established and the eggs are to be removed. Moving sheep recipients is routinely done, but moving goat recipients may reduce pregnancy rates.

Body condition. Donors in particular should be in good body condition – ideally 3 to 3.5. Recipients must also be in good condition

Diet. Well-conditioned animals do not benefit from nutritional flushing. Poorly conditioned animals will benefit. Flushing should focus on energy not protein. Very high protein diets are associated with reduced embryo quality and increased early embryonic death in recipients. Diets high in legume might also contain phytoestrogens (plant estrogens) that are also associated with reduced fertilization and increased embryonic death. In my view most people get carried away with nutritional concerns – trying to insure a successful program. Selecting well-conditioned animals, in good health and on a sensible diet of good quality forage with a suitable grain supplement and adequate mineral is best.

Donor health. ET can be used to salvage genetics from donors affected by disease. Results suffer as body condition declines. Serious illness drastically reduces superovulation results.

Breeding program. Normally we recommend one ram or buck to one or two superovulated donors. Superovulated ewes come into heat earlier than normal within 18 to 24 hours of pessary removal. Males are introduced for 36 hours or are hand bred during that period. GNRH is administered to help to insure successful ovulation. Laparoscopic AI is often used to overcome reduced fertilization. Laparoscopic AI has shown most benefit when young donors and/or young males are used in an ET program.

Drug handling. Mishandling easily alters drugs used for superovulation – particularly temperature. Results will suffer. Directions must be followed closely. Drugs must be administered every 12 hours for three days and must be given subcutaneously (in the fat under the skin).

Number of embryos transferred. Pregnancy and embryo survival rates are highest when two or three embryos (in contrast to one) are transferred to each recipient.

Recipient selection and care. Selecting good recipients is critical to success. It is a mistake to believe that any old ewe or doe will do. Recipients should be young (2-4 yr.), reproductively proven, in good condition and in good health. They are the ones doing all the work of accepting the embryos and carrying the pregnancy. They should be free of diseases of concern to insure the birth of clean offspring.

Producers must have realistic expectations and understand the risks inherent in embryo programs. Many of the common causes of low success can be prevented by good management.