Embryo Transfer (ET)

History of ET

The first embryo transfer calf was born in 1951, but ET as we know it was not done on larger scales until the 1970’s, where surgical collection and implantation were used. In the late 1970’s non-surgical techniques were pioneered and the ET industry developed into what we know today.

Why Use ET – Pros

♠ ET allows access to elite genetics from other folds that you would otherwise not gain access to (breeders are not going to sell these animals, nor are they likely to sell the progeny of these animals). You can also rapidly multiply the number of offspring possible from your own elite female.

ET Allows the Use of Elites Dams & Sires

♠ ET allows you to maximise genetic improvement over a short time period with the number of calves that can be produced.
♠ It can utilise potentially superior genetics on both sire’s and dam’s side, from other folds (including overseas), compared with just the sire side with AI.
♠ You can potentially get a lifetime of calves on the ground from a particular female by the time she is 2-3 years old.

Why Not to Use ET – Cons

♠ Flushing can yield quite variable results (you might get 18 embryos, or you might get none – the average is 4-5 per flush).
♠ Complications for the cow such as lost production (for 6+ months) because she is getting flushed and this takes time, sometimes poor fertility/conception rates for the donor cow after flushing, raised tail head post flushing and very occasional serious complications (deaths have occurred, uterine infections).
♠ You need to carry a number of recipient females that will not end up with an ET calf inside (although some companies will provide a service that gets around this need).
♠ Abortion & calf losses can be as high as 10% (over & above natural losses).
♠ Although a number of ET animals will be superior animals if you have selected wisely, not all progeny from elite sires and dams are elite themselves. You still need to use selection pressure & cull those animals that are not up to standard. We eat imported embryo animals if they are not of adequate quality.

What Exactly Happens with ET?

► Selecting an elite cow is not just choosing the cow with the best production figures or the most show ribbons, they also need to be amongst the most fertile in your herd. They must never have taken more than two inseminations to fall pregnant & consistently had calves 365 days apart or less. Heifers can also be used here, remembering that they are unproven, and that the cervix is a little narrower and can present difficulties getting through in some instances.

► The cow is ‘superovulated’, meaning that they are programmed to come on heat (with the use of hormone injections and implants) and are stimulated to produce more than the usual one egg with the use of the a drug called FSH (follicle stimulating hormone). So long as nutrition is managed carefully, this process can be repeated every 45-60 days.

► When the first sign of standing heat is noticed, the cow is inseminated 12 & 24 hours later (& sometimes 36 hours later). Seven days later the flushing procedure occurs (fertilized embryos being around 7 days old). These are removed with the aid of an epidural local anaesthetic to prevent straining in the rectum and 5-10 flushes of a special fluid are used in each uterine horn, and the embryos drain into a collecting receptacle.

► The embryos are then graded according to an international grading system and can be used fresh or can be frozen in liquid nitrogen. Like semen straws, if they are stored in liquid nitrogen, they will last for decades (with a marginal drop in pregnancy rates once frozen).

Table of Various Grades of Embryos and the Affect on Conception rates:

Grade of embryos	Proportion of embryos produced	Condition/Quality of embryos	Freezable?	Conception Rate
A	55%	excellent	Yes	60-70% *
B	10%	good	Yes	40-50% *
C	5%	fair	No	10-30% (fresh only)
Degenerate	}40%	useless	No	0%
Unfertilised		useless	No	0%

* Conception rates for frozen embryos (expect 10-15% above this if using fresh embryos).

► The recipients are synchronised in the same way as the donor cow, and  if fresh embryos are to be put in, then they are synchronised on exactly the same days. The ET vet will palpate their ovaries 7 days after they have cycled and determine which ones have had strong enough heats to support a pregnancy. Approximately 25% of recips that are programmed will not be used because their heat was not strong enough –  some may not cycle at all or in the 24 hour window allowed, and others may not have had a good heat and therefore may not sustain the pregnancy. So out of the 10, you might end up with 7-8 that can have embryos put in. With a 60-70% conception rate, you would then end up with 4-5 pregnancies, after starting with 10 recips!

Alternatively, some artificial breeding companies have a service where they have access to bulk recipient females and can do all this for you & supply you with a confirmed pregnancy (9 to 12 weeks). There is a cost for this service of course, but it certainly reduces the need to carry all those recipients right through.

How to Maximise the Success of an ET Program?

Similar to AI, there are many steps that are required to be performed appropriately, otherwise the expected success rate (pregnancies achieved) will decrease a little to a lot with each link in the chain that is not well performed. The major determinant of success here are:

A) Embryo Quality
♠ Donor cow condition, health & nutrition
♠ Handling & selection of embryos by ET vet
♠ Environmental temperatures

B) Recipient Suitability
♠ Condition, health & nutrition
♠ Recip temperament is critical
♠ Selectiveness of ET vet (need to have first signs of standing heat within 24 hrs either side of 7d, and ideally 12 hours.)

1)     Condition of the donor cow – not underweight but at least as big a problem is being over weight. Body Condition Score of 3.0 ideal, but not < 2.5 or >3.5 to maximise success.

2)     Nutrition of donor cow – they need to be on a rising plain of nutrition over the 4-6 weeks prior to flushing (meaning slightly more feed each week leading up the that date), but interestingly, too high a protein level is not a good thing. This means that free access to lush pasture is not ideal. A better system is to lock them up and hand feed a known amount of average quality hay and 1-2kg of lower protein pellets (~12% protein) each day. High legume (clover and lucerne) intake should be avoided not only because of the protein content, but also because they can contain variable amounts of oestrogens which can potentially interfere with hormone programming.

3)     Vitamin & mineral supplementation – Vitamins B12, A, D & E & selenium should be given to every donor (& recipient) 4-5 weeks prior to programming. While not every animal needs this, on average better results are seen with supplementation.

4)     The skill of the ET vet. Here experience certainly counts in the flushing process as well as the implantation of the embryos, or more importantly, the selection of which recipient animals should receive an embryo.

5)     The choice of recipient – these cows need to be of a large enough frame & have adequate milk production (dairy cross animals are ideal) but most importantly they need to be quiet. Nervous or fractious recipients will have a 5-10% lower conception rate. We find black Angus heifers ideal (easy to get, cheap & easy to move on afterwards).

6)     Nutrition of recipients – needs to be exactly the same as for the donor cow – not too fat & not too thin & on a rising plain of nutrition, and watch the protein levels in the feed here to.

7)     The first 9-12 weeks of gestation – the recipients need to be put down in the back paddock and not stressed for this period. There is an increased rate of abortion of around 10% over and above normal losses in this period, so we minimize stressful events, until they are preg tested at 9-12 weeks.

8)     Fresh vs frozen embryos – using fresh embryos will increase the conception rate by 10-15% over frozen embryos, but it is not always possible to use fresh embryos.

9)     Pestivirus infection – a growing problem in Australia that can affect flushing of the donor cow, as well as a cause infertility in recips. Blood tests should be performed on all donor and recip animals prior to commencing the program.

10) Freezing in glycerol versus ethylene glycol – glycerol in the right hands may give a marginally improved conception rates but experience in thawing is imperative, otherwise poorer conception rates are found. Generally, most commercial embryos are preserved in ethylene glycol and can be inseminated directly in a similar fashion to doing an AI.

11) Environmental temperatures – we have had personal experience with poor flush results when very hot days occur at the time of insemination or in the week after this, prior to flushing. Very hot days and Highlands do not go that well together. We have heard of Highlands not flushing very well in below zero temperatures.

The Full Costs to Produce an ET Calf

There are many hidden costs in getting an embryo calf on the ground & while we can not give exact costs that are relevant for every situation, an embryo calf will cost from $6-$9,000. This includes flushing or purchase price of embryos, cost of purchasing recips, drugs for recips & ET vet transfer costs, feed costs for donor & recips, and should ideally take into account a figure for your time involved in all the steps.

The Future for the ET Industry

Sexed embryos

This was used a lot in the 80’s & 90’s, but less so now. It was established for the dairy  ET industry as bull calves have nearly no purpose. Only certain ET technicians can perform this, as it is a delicate and tedious operation. There is a cost associated with it because of this and the conception rates with sexed embryos will be marginally lower because of the test requires some cells to be removed from the embryo. Because of the damage to the embryos, they are not of exportable quality to most countries. The accuracy rate is around 95%, and needs to be performed prior to freezing. This could be a very promising technique for Highland breeders, who mostly want heifers as well.

Using Female Sexed Semen

Sexed semen is becoming more common place since 2020 and many bulls are collected for both conventional (non-sexed semen) and female sexed semen. The latter is quite a process and doesn’t always work in every collection, and conception rates are slightly lower. This semen can be used in embryo transfer programs to give you a 95% guarantee that the calves produced from embryos flushed will be heifers.

Embryo Splitting

Can be done but is of questionable value, economically anyway. You need twice as many recipients and the pregnancy rate is lower, so it is possible but not commonly utilised.

DNA Analysis on Embryos

In the same way DNA tests can be performed on tail hair samples, a plug of cells from an embryo (normally 4-6 cells) can be replicated to create endless amounts of DNA. This DNA can then be analysed for various characteristics to know what type of calf will be produced should that embryo result in a calf on the ground. Currently only certain characteristics can be accurately determined via DNA analysis (eg sex, coat colour, marbling) but more values will become available over time.

Use of Ultrasound

Skilled operators can detect pregnancies as early as 4 weeks gestation which allows early reuse of recips rather than waiting for the traditional pregnancy testing at 8-9 weeks at the earliest. Ultrasound can also be used to sex calves at 8-10 weeks gestation which is really only of use when recips are sold in calf commercially.

Cloning

It is possible to clone cattle (or embryos) but the process is still in its burgeoning stages. There are still likely to be some short comings in the life of animals produced by this process (we all remember Dolly the sheep). Because of the expense and the moral dilemmas here, it is not likely to be a technique used in the near future in cattle breeding.

 

IVF (‘Test-tube calves’) This is a specialist area within the ET industry. Eggs are collected from the donor cow with a needle guided by an ultrasound image, all through the vagina. They are fertilised with the semen of your choice in the laboratory and then incubated in various fluid media until the embryo is 7 days old. Potentially, donor cows can be any age at all, and can even be up to 3-5 months pregnant. They do not need to be programmed to superovulate and on average 7-10 eggs are recovered from each cow/heifer, but this collection process can be repeated twice a week for 2-3 weeks. On average, around 60% of eggs collected end up as usable embryos, but there appears to be more variation in these figures than there is with conventional embryo flushing. Sometimes large numbers of eggs can recovered from one cow on one occasion – as many as 40-50, but as for larger numbers retrieved with conventional flushing, the quality of these eggs can often be poorer compared with eggs from a normal collection of 7-10. Conception rates have improved with some newer techniques that have been developed in the last 2-3 years and are approaching those achieved with regular superovulation and flushing – around 50-60% for fresh and 40-50% for frozen embryos. Where poorer quality embryos are recovered (as for difficult breeders, older cows or with extremes of environmental temperatures), these figures can drop significantly. Pregnancy rates can be as low as 10-30% in many cases. Lost pregnancies in the first 3-6 weeks were higher in the past but these rates are said to be similar to conventionally produced ET pregnancies with the newer techniques and improved embryo quality. Costs are 50-100% higher per embryo, and you still need a similar number of recips per embryo. The main positives are that no drugs are required and hence complications for the donor are rare, and there is flexibility of the animals that can be used for egg donation. Another benefit is that you can get by with only a fraction of a semen straw (one straw can be used across the eggs of 10-20 cows), so this is a great technique to use with rare or very expensive semen. As further advancements are made, and costs come down, it is easy to believe that some day IVF may replace the tradition method of creating embryos.

Referrences:

1) Training Manual for ET– Excellent training manual for embryo transfer in cattle (considerable detail)

2) Factors Affecting Success in ET – Know how on improving pregnancy rates in traditional ET programs.

3) The Real Costs of ET – Factors involved in calculating total cost of an ET program.