HOW TO TREAT THE SUB-FERTILE MARE


set time after gonadotropin administration. Typi- cally, the gonadotropin stimulation is given on the day that the dominant follicle reaches receptive di- ameter (based on mare history or typical receptive size for breed) or appearance on ultrasonography. A workable schedule is to give the gonadotropin stimulus in the afternoon of Day 1, aspirate the follicle 24–28 hours later on the afternoon of Day 2, and immediately ship the oocyte by courier to the laboratory where it is subjected to ICSI the morning of Day 3. These steps are difficult to organize to a set schedule and so may result in oocytes being recovered when a laboratory will not receive them or when they may result in significant after-hours charges for handling and micromanipulation. Although the above drawbacks exist for utilization


of the dominant stimulated follicle, oocyte recovery from this follicle is much easier and initially more rewarding than is aspiration of immature follicles, so practices just starting oocyte recovery may want to start by performing preovulatory follicular aspiration.


Oocyte Transfer


Because OT involves standing flank laparotomy on a recipient mare, it is best conducted when a single oocyte of high viability is available. Thus, OT is most suitable for use with the in vivo-matured oocyte recovered from the dominant stimulated follicle.3 It is possible to use OT with multiple in vitro– matured oocytes (such as after collection of imma- ture oocytes from TVA of all immature follicles, or recovery post-mortem);4 however, the pregnancy rate per oocyte transferred is very low because: 1) the maturation status of the oocytes is unknown at the time of transfer (the cumulus is left on so that the oocytes will “stick” in the oviduct, thus staging the meiotic status by viewing the polar body is not possible), and 2) the developmental competence of in vitro-matured oocytes is lower than that for in vivo- matured oocytes recovered from the stimulated pre- ovulatory follicle. In my opinion in the case of multiple oocytes, it is best to transfer all oocytes (maximum, 12 per oviduct) to one or two recipient mare(s), then to flush the uterus for embryo recov- ery—this has been performed successfully 9–10 days after OT in a commercial program5—and per- form standard embryo transfer (ET) to secondary recipients with any recovered blastocysts. This has the drawback of potential inefficiencies at uterine flush and at ET, but is preferable to trying to per- form a surgery for each of multiple oocytes, with a low chance of pregnancy per surgery. All procedures for OT are workable within a well- equipped practice. Oocytes may be recovered from the stimulated preovulatory follicle with a needle placed through the flank,1 eliminating the need to purchase and learn to use a TVA system. Recovery rates with this system are similar to those with transvaginal aspiration.5,6 If the oocyte is har-


156 2016  Vol. 62  AAEP PROCEEDINGS vested close to the time of ovulation, it can be held


for a few hours without a CO2 incubator or need for gassing, if medium with a nonbicarbonate buffer system such as in M199 with Hanks’ salts with 10% fetal bovine serum (FBS), is used. The oocyte may be placed in this medium in a capped tube in a water bath. The transfer procedure is straightforward5,6 and is performed via standing flank laparotomy. The OT process is robust; within broad limits


there is little effect of variation in factors such as time from donor gonadotropin administration to fol- licle aspiration, time from oocyte recovery to trans- fer, or time of recipient follicle aspiration in relation to insemination or transfer.5 The main drawback to OT is the labor entailed. Recipients must be exactly synchronized and must have their preovulatory follicle successfully aspi- rated (or be in anestrus or early estrus and be hor- monally treated);7,8 the donor mare must be monitored for follicle activity and have her follicle successfully aspirated, the recipient must be insem- inated (thus good-quality semen is required), and then the surgery must be performed and the recipi- ent cared for afterward. When OT is used clinically with old, sub-fertile donor mares, the foaling rate is only approximately 25% per oocyte transferred5,9,10 therefore 75% of the time this labor will not result in a foal. However, it is a viable method for producing foals from mares that cannot produce foals via nat- ural conception or ET.


Somatic Cell Nuclear Transfer (Cloning)


Cloning is an effective method to produce a young, fertile filly with the genetics of the original donor. This filly can then be bred throughout its life— including via assisted reproductive technologies—to produce foals carrying the original donor mare’s genetics. Cloning typically involves transfer of the donor


mares’ genome (via the nucleus of one of her somatic cells) into a host oocyte obtained from another mare. Host oocytes are typically obtained from ovaries re- covered from an abattoir. The mitochondria of the resulting foal will be that of the host oocyte, not of the original donor mare. It is unknown whether this has any effect on the cloned offspring but the client must be informed of this, and educated to understand what it means. If this is important to your client, this can be avoided by performing nu- clear transfer on oocytes recovered from mares that are maternally related to the donor.11 Cloned foals and their progeny are not able to be registered with many breed registries. If registra- tion is important to your client, then they should check with their breed registry to determine the regulations regarding clones. Cloning is many times more expensive than is one


cycle of ICSI or OT; however, once the cloned filly is produced, the expense of achieving foals from her is that of standard reproductive procedures. Thus, over the cloned filly’s life span, her ability to produce


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