Embryo transfer in horses is a reproductive technology that involves collecting a fertilized embryo from a donor mare and implanting it into a recipient mare, which then carries the pregnancy to term. This technique allows for the production of multiple offspring from a single mare within a breeding season. The procedure includes several stages: synchronization of the donor and recipient mares' estrous cycles, collection of the embryo, and its subsequent transfer to the recipient mare. Embryo transfer is utilized to enhance genetic dissemination, preserve valuable genetics, and manage breeding schedules. This page aggregates peer-reviewed research studies and scholarly articles that explore the methods, efficiency, and applications of embryo transfer in equine reproduction.
Woods GL, Steiner JV.The objective of this project was to produce multiple pregnant recipient mares during one year from each of four mares which were in athletic training and competition. Twenty-six embryos were recovered from the four mares in 38 collection attempts. Each embryo was surgically transferred to one of 20 recipient mares. Nine of 10 recipient mares which were transferred to on one occasion were detected pregnant at the one week post transfer pregnancy examination and six of eight mares which were transferred to on two occasions were detected pregnant at one of the one-week post transfer pregnancy ex...
Hyland JH, MacLean AA, Robertson-Smith GR, Jeffcott LB, Stewart GA.The removal of one of twin embryos was attempted by infusion of 24% (w/v) saline into the gestation sac in 2 mares by laparotomy. The treatment was successful in one mare (Case 1) and the untreated embryo remained viable. However, neither foetus survived in the second mare (Case 2). Plasma oestrone sulphate (E1S) concentrations fell immediately after treatment in both mares but recovered to approximately 50% of pretreatment levels in Case 1. In Case 2 plasma E1S concentrations declined steadily and were less than 1 ng/ml within 6 days of treatment. These preliminary results suggest that the me...
Iuliano MF, Squires EL, Cook VM.A 2 X 2 cross-classified experiment was conducted to investigate the effect of age of equine embryo (7 vs 8 d postovulation) and method of transfer (surgical vs nonsurgical) on pregnancy rates at 50 d of gestation. Embryos were recovered 7 or 8 d postovulation using a Foley catheter and 3 liters of modified Dulbecco's phosphate-buffered saline (PBS). Upon identification, the embryos were placed in millipore-filtered PBS containing 20% heat-inactivated steer serum and maintained at room temperature until transferred. At the time of recovery, embryos were randomly assigned to be transferred eith...
Allen WE, Goddard PJ.The uteri of 13 pony mares were examined daily by ultrasound during the first two months of gestation. The conceptus was first identified between 12 and 16 days after ovulation and the embryo was seen on the ventral surface of the conceptus after Day 21. The foetal heart could be visualised after Day 22 and spontaneous movement of the foetus occurred after Day 39. No consistent pattern was seen in the development of the foetal membranes, although attachment of the umbilical cord to the allantochorion was always on the dorsal aspect of the conceptus. Daily measurements were made of the diameter...
Kraemer DC.The procedures that are collectively referred to as embryo transfer (ET) have many uses. They were first used as research tools to study fetal-maternal physiology. Since the first successful mammalian embryo transfer in 1890, ET has been utilized for enhancement of genetic selection; diagnosis and treatment of infertility; control of infectious disease transmission; screening for genetic defects; propagation of rare and endangered species; and the study of developmental biology. Most of the embryo transfers have been intraspecific. A listing of the species includes rabbit, rat, sheep, mouse, g...
Ginther OJ.Fixation and orientation of the conceptus within the uterine lumen of 40 barren mares were examined by ultrasound daily on days 11-21 and at three-day intervals thereafter until day 48. The growth curve of the in situ conceptus had a distinct plateau between days 17-24, as determined by measurements of the width and area of conceptuses on the ultrasound images. The vesicle expanded at an average daily rate of 3-4 mm before the plateau and 2-3 mm after the plateau. Dramatic changes occurred in the shape of conceptus. The predominant shapes were approximately as follows: days 11-16, spherical; d...
Douglas RH.The number of successful embryo recoveries was 56.0% in experimental mares with no history of breeding problems but in barren mares only 34.3% of embryo recovery attempts were successful. When a recovery attempt was successful, the incidence of twin embryos was 10.7% in experimental mares and 16.7% in barren mares. Maintenance of embryos in vitro for greater than 6 h before non-surgical transfer significantly reduced pregnancy rates compared to maintenance for less than 3 h. The pregnancy rate in recipients at 100 days after ovulation was lower (P less than 0.05) than those at 24 and 30 days a...
Imel KJ, Squires EL, Elsden RP, Shideler RK.Embryos were recovered in 39 of 47 attempts (83%) during 1979 and in 75 of 104 attempts (72%) during 1980. The mean diameters of day 8 and 9 blastocysts were 1.00 and 2.13 mm, respectively. The injection of prostaglandin F2 alpha or prostalene on day 8 or 9 after ovulation resulted in a mean interval between embryo recovery attempts of 17.7 +/- 0.3 days. Number of embryos recovered within mares did not vary significantly with repeated attempts. Following surgical transfer, 8 of 15 recipients (53%) were pregnant at 50 days after ovulation, whereas only 4 of 15 recipients (27%) were pregnant at ...
Tischner M, Bielański A.Observations were made on 14 mares which were non-surgically flushed on one or more occasions for recovery of embryos on Days 7-9 after ovulation. Flushing alone shortened the oestrous cycle by about 2.7 days but increased the number of mares failing to ovulate. Flushing followed by an injection of a prostaglandin analogue shortened the cycle by about 4.5 days and hence increased the frequency of ovulations. From a total of 70 flushings performed, 27 embryos were recovered and of 12 mares inseminated during the first oestrus after embryo collection, 6 conceived and foaled. The other 7 mares we...
Vogelsang SG, Sorensen AM, Potter GD, Burns SJ, Kraemer DC.Embryos were collected nonsurgically on Day 7 or 8 after ovulation from 7 Quarter horse mares using a modified 30-ml Foley catheter to flush the uterine horn ipsilateral to the recent ovulation with 500 ml TCM-199 containing Hepes buffer. After collection, the uteri were infused with nitrofurazone to reduce the chances of infection due to the procedure. Eleven collections from 7 mares resulted in recovery of 9 embryos and nonsurgical transfer of 4 of these resulted in the birth of one foal. After collections, 8 oestrous cycles averages 22.75 days and 2 extended oestrous cycles were 43 and 59 d...
Allen WR.Fertilized eggs reach the uterus of the mare by the sixth day after ovulation whereas unfertilized eggs remain lodged in the fallopian tubes for many months. However, embryo removal studies indicate that the fetal message for luteal maintenance in the pregnant mare is not transmitted until Days 14-16. The equine endometrial cups comprise a series of small, ulcer-like endometrial outgrowths which are present in the pregnant horn of the uterus between 40 and 150 days of gestation. Each cup consists of a discrete colony of large, gonadotropin-secreting, trophoblast cells derived from a specialize...
Tervit HR.Recent advances in reproductive physiology offer ways for exploiting superior, female cattle and for manipulating physiological events such as parturition. The techniques involved in these advances and their associated hazards are the subject of this review.
Allen WR, Rowson LE.Surgical and non-surgical methods used for the recovery and transfer of fertilized horse eggs are described. Sixteen of the twenty-three zygotes recovered surgically between Days 1 to 6 after ovulation from thirty donor mares were transferred surgically to synchronized recipients; seven pregnancies resulted. Seven of the eleven zygotes recovered non-surgically between Days 6 to 8 after ovulation from twenty-eight donor mares were transferred non-surgically to synchronized recipients; five pregnancies resulted.
In this study, 198 donor mares of different breeds, ages, and reproductive category were inseminated with fresh, cooled and frozen or frozen and cooled semen at the embryo transfer station or in private artificial insemination centers during 10 breeding seasons. The results of this activity were retrospectively analyzed by Pearson Chi-square test and logistic regression to evaluate factors affecting multiple ovulations, embryo recovery, embryo quality, and embryo diameter. Out of the 661 cycles, 937 ovulations were recorded (mean ovulations/cycle: 1.42 ± 0.58). Ovulation rate and incidence of...
Hannan MA, Murata K, Takeuchi S, Haneda S, Cheong SH, Nambo Y.The objectives of this study were to determine the plasma profile of equine chorionic gonadotropin (eCG) and its association with the formation of supplementary corpus luteum (CL) and plasma progesterone concentrations in embryo transfer Hokkaido native pony recipient mares. Blood samples and transrectal ultrasound examination of the reproductive tract were carried out weekly from the day of ovulation until week 32 of gestation (n = 4). Plasma concentrations of eCG and progesterone were measured by enzyme immunoassays. The eCG concentration was first detectable at week 5 for 2 mares and at wee...
Witt MC, Bollwein H, Probst J, Baackmann C, Squires EL, Sieme H.The aim of the present study was to investigate the effects of a gonadotropin treatment to induce superovulation on ovarian and uterine blood flow and its relationship with steroid hormone levels and ovarian response in mares, using color Doppler sonography. Each of six mares were examined sonographically in five cycles for 3 d (t1 to t3) during the follicular development phase (FDP) beginning at a follicle size of ≥ 22 mm, and for 4 d (D-4 to D-1; D0 = Ovulation) in the preovulatory phase (POP). After each examination, total estrogens (E(tot)) and progesterone (P(4)) levels were determined ...
Clérico G, Taminelli G, Veronesi JC, Polola J, Pagura N, Pinto C, Sansinena M.Oocyte vitrification is considered experimental in the horse with only three live foals reported. The oxidative conditions induced by vitrification could in part explain the poor results and melatonin, a powerful antioxidant, could stimulate ROS metabolization and restore mitochondrial function in these oocytes. Our objective was to determine the oxidative status of vitrified equine oocytes and to analyze the effect of melatonin on mitochondrial-specific ROS (mROS), oocyte maturation, ICSI embryo development and viability. Immature, abattoir-derived oocytes were held for 15 h and vitrified in...
Cochran R, Meintjes M, Reggio B, Hylan D, Carter J, Pinto C, Paccamonti D, Graff KJ, Godke RA.In vitro fertilization in horses has been less successful than anticipated owing to: (i) the inability to collect large numbers of good quality oocytes; (ii) alterations in the zona pellucida that occur during in vitro maturation of equine oocytes; and (iii) inadequate preparation of equine sperm cells. In addition, studies in humans, mice and cattle have indicated that high concentrations of glucose in culture media may inhibit embryonic development in vitro and this may also be a problem for development of equine embryos in vitro. The aims of the present study were: (i) to achieve fertilizat...
McCue PM, Matthews PM, Prell MJ, Bellone RR, Allen H.Genetic testing is required for the registration of foals of most equine breeds. Objective: To describe two clinical cases of marked delayed embryonic development or delayed fertilisation in pregnancies generated by embryo transfer. Methods: Case report. Methods: Donor mares were inseminated with semen from one stallion during one oestrous cycle and semen from a different stallion on the subsequent oestrous cycle. Embryo(s) were collected 8 days after ovulation during the second oestrous cycle and transferred into synchronised recipient mares. Genetic testing was performed to determine paren...
Squires EL, McKinnon AO, Carnevale EM, Morris R, Nett TM.For embryos collected from mares 7 days after ovulation, embryo recovery for single-ovulating mares was 53% compared to 106% for double-ovulating mares. Pregnancy rates 50 days after surgical transfer were 68 and 129%, respectively. Concentrations of LH were similar during the periovulatory period for cycles which included single or double ovulations. Horse pituitary extract given for 5.5 days resulted in greater than or equal to 2 ovulations (mean 3.8) in 26 of 28 mares and 2.0 embryos were recovered per donor compared to 0.65 for controls. Non-surgical pregnancy rates for embryos collected f...
Bergfelt DR, Meira C, Fleury JJ, Fleury PD, Dell'Aqua JA, Adams GP.A regimen of progesterone plus estradiol (P&E) was used as a standard for ovarian synchronization to test the efficacy and evaluate the commercial application of ultrasound-guided follicle ablation as a non-steroidal alternative for ovulation synchronization in mares. Recipient mares at a private embryo transfer facility were at unknown stages of the estrous cycle at the start of the experiment on Day 1 when they were randomly assigned to an ablation group (n=18-21 mares) or to a P&E group (n=20-21 mares). In the ablation group, mares were lightly sedated and all follicles > or = 10...
Tozaki T, Ohnuma A, Nakamura K, Hano K, Takasu M, Takahashi Y, Tamura N, Sato F, Shimizu K, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Hamilton NA....The creation of genetically modified horses is prohibited in horse racing as it falls under the banner of gene doping. In this study, we developed a test to detect gene editing based on amplicon sequencing using next-generation sequencing (NGS). We designed 1012 amplicons to target 52 genes (481 exons) and 147 single-nucleotide variants (SNVs). NGS analyses showed that 97.7% of the targeted exons were sequenced to sufficient coverage (depth > 50) for calling variants. The targets of artificial editing were defined as homozygous alternative (HomoALT) and compound heterozygous alternative (ALT1/...
Binder H, Jakob C, Bucher P.Successful application of embryo transfer (ET) has become common practice in cattle, horses, sheep, goats and a variety of other species held in captivity. Yet in cattle only has the technique been established commercially. In 1994 more than 100,000 bovine embryos have been transferred in European countries. Important progress in transvaginal ovum pick up (OPU), in vitro production (IVP) and cryopreservation have further improved the applicability of ET. Direct transfer simplifies the procedure considerably allowing individual transfers and eliminating the need of synchronizing recipients. In ...
Ball BA, Little TV, Weber JA, Woods GL.The estimated embryonic loss rate between Days 4 and 14 after ovulation for young, normal mares (9%) was significantly lower (P less than 0.01) than the estimated embryonic loss rate for aged subfertile mares (62%). Fertilization rates, which were based on the recovery of embryos at Day 4 after ovulation, were 96% and 81% (P less than 0.1) for normal and subfertile mares, respectively. Day-4 embryos were collected from the oviducts of normal and subfertile donors mares. These embryos were transferred to the uteri of synchronized, normal recipient mares to test the hypothesis that the high inci...
Rossini JB, Rodriguez J, Bresnahan DR, Stokes JE, Carnevale EM.The clinical use of intracytoplasmic sperm injection (ICSI) in horses usually involves the transfer of embryos into recipient mares, resulting in substantial cost increases. This is essential when subfertile mares are oocyte donors; but some donors are fertile, with ICSI compensating for limited or poor-quality spermatozoa. Fertile oocyte donors could carry pregnancies, eliminating the need for a recipient. We assessed the potential of using oocyte donors as recipients for their own ICSI-produced embryos during the same cycle. Donors in oestrus and with large dominant follicles were administer...
Iuliano MF, Squires EL, Cook VM.A 2 X 2 cross-classified experiment was conducted to investigate the effect of age of equine embryo (7 vs 8 d postovulation) and method of transfer (surgical vs nonsurgical) on pregnancy rates at 50 d of gestation. Embryos were recovered 7 or 8 d postovulation using a Foley catheter and 3 liters of modified Dulbecco's phosphate-buffered saline (PBS). Upon identification, the embryos were placed in millipore-filtered PBS containing 20% heat-inactivated steer serum and maintained at room temperature until transferred. At the time of recovery, embryos were randomly assigned to be transferred eith...
Cran DG.Many tens of thousands of calves resulting from artificial insemination (AI) have been born worldwide after XY sperm separation and commercial production is underway in several countries. Accuracy of sex selection is some 90% and can be achieved both in research facilities and at AI studs in rural locations. Most facilities sort X- sperm which have also be utilised for superovulation and embryo transfer projects and for in vitro fertilisation (IVF) as well as AI. Sort rates of some 15 x 10(6) sperm/h are currently achievable and are used for low dose insemination, generally at 2 x 10(6) frozen...
Gil L, Olaciregui M, Luño V, Malo C, González N, Martínez F.In recent years, there has been an increased interest in new preservation techniques that facilitate sperm storage and distribution, with freeze-drying (FD) having been proposed as an alternative method for sperm preservation and maintenance of genetic resources in different animal species. FD is a method in which frozen material is dried by sublimation of ice, thereby involving a direct transition from a solid (ice) to a vapour (gas) phase. One of the main advantages of FD is that nitrogen and dry ice are no longer required for the storage and shipment of frozen sperm, which can be stored at ...
Woods GL, Steiner JV.The objective of this project was to produce multiple pregnant recipient mares during one year from each of four mares which were in athletic training and competition. Twenty-six embryos were recovered from the four mares in 38 collection attempts. Each embryo was surgically transferred to one of 20 recipient mares. Nine of 10 recipient mares which were transferred to on one occasion were detected pregnant at the one week post transfer pregnancy examination and six of eight mares which were transferred to on two occasions were detected pregnant at one of the one-week post transfer pregnancy ex...
Takagi M, Nishimura K, Oguri N, Ohnuma K, Ito K, Takahashi J, Yasuda Y, Miyazawa K, Sato K.The viability of embryos before flushing from donor mares (n = 5) and after transfer to recipient mares (n = 7) was monitored in mare serum by detecting early pregnancy factor (EPF) using the rosette inhibition test (RIT). The EPF activity was measured in donor mares before and after natural mating at natural estrus; after ovulation on Days 2, 5 and 8; and after embryo flushing (Day 8) on Days 8, 9, 10 and 13 after ovulation. The collected embryos were transferred immediately after flushing. The EPF activity in recipient mares were measured on the day of transfer and after embryo transfer on D...
Seidel GE.Principles for selecting future research projects include interests of investigators, fundability, potential applications, ethical considerations, being able to formulate testable hypotheses and choosing the best models, including selection of the most appropriate species. The following 10 areas of assisted reproduction seem especially appropriate for further research: efficacious capacitation of bovine spermatozoa in vitro; improved in vitro bovine oocyte maturation; decreasing variability and increasing efficacy of bovine superovulation; improved fertility of sexed semen; improving equine IV...
Wegner W.In 1893 a major component of veterinary practice derived from 4 million horses and from farm animals, whereas thousands of dogs were even slaughtered every year, especially in Saxony and Silesia. Nowadays companion animals figure as an element of ever growing importance for the practitioner. Teaching and research adapted themselves to this altered situation, research funding and professional politics not yet so completely. In former days veterinarians even presided the licensing committees for breeder animals--today the animal breeding act merely mentions them in connection with institutions o...
Bromfield JJ.Artificial insemination has been a landmark procedure in improving animal agriculture over the past 150 years. The utility of artificial insemination has facilitated a rapid improvement in animal genetics across agricultural species, leading to improvements of growth, health and productivity in poultry, swine, equine and cattle species. The utility of artificial insemination, as with all assisted reproductive technologies side-steps thousands of years of evolution that has led to the development of physiological systems to ensure the transmission of genetics from generation to generation. The ...
Azuma T, Choi YH, Hochi S, Oguri N.The development of in-vitro matured and microfertilized horse oocytes was examined in vitro. Fertilized oocytes were produced by 20-h insemination of in-vitro matured and partially zona-removed oocytes with frozen spermatozoa that had been treated with caffeine/calcium ionophore A23187 (fertilization rate 34.2%, monospermy rate 76.9%). Embryonic development was assessed by the number of nuclei stained with Giemsa solution. In Experiment 1, a continuous 8-day culture of the microfertilized oocytes in TCM199 or modified synthetic oviduct fluid (m-SOF) supplemented with 10% fetal bovine serum or ...
Hinrichs K, Riera FL, Klunder LR.Embryo transfer was performed in three mares with gonadal dysgenesis. Karyotypes of the mares were as follows: Mare 1, 63,XX, 64,XX, 65,XX; Mare 2, 63,X; and Mare 3, 65,XXX. The mares were administered progesterone in oil, 300 mg intramuscularly daily, starting 1 or 2 days after donor mare ovulation. Embryos were transferred on day 7 after donor ovulation. Mare 1 became pregnant after the first embryo transfer and had a normally developing fetus on necropsy on day 45 of gestation. Mare 3 became pregnant after the third embryo transfer, but the embryo was lost between day 14 and day 18 of gesta...
Woods GL, Hillman RB, Schlafer DH.To evaluate embryo transfer as a possible method to circumvent infertility in mares, embryos from 14 normal and 14 infertile mares were collected three times and examined. Fewer flushes (p less than 0.05) from normal than infertile mares (1/42 vs 9/42) contained only abnormal embryos whereas more flushes (p less than 0.05) from normal than infertile mares contained one or more normal embryos (28/42 vs 8/42). More flushes (p less than 0.05) from normal than infertile mares contained embryos (29/42 vs 17/42). The embryo diameters (mm) at either day-7 or day-8 post ovulation were greater (p less ...
Wang X, Asgenbaatar N, Shen Y, Yi M, Zhao B, Ren H, Davshilt T, Ulaangerel T, Wang M, Burenbaatar A, Tian S, Li B, Dugarjav M, Bou G.Proliferation of embryonic fibroblasts under the same cell culture conditions, hinny embryonic fibroblasts (HiEFs) was slower than horse embryonic fibroblast (HEFs), donkey embryonic fibroblasts (DEFs) and mule embryonic fibroblasts (MuEFs). The imprinted genes IGF2 and IGF2R are important for cell proliferation. Therefore, we investigated whether the slower proliferation of HiEFs is related to an aberrant gene expression of IGF2 or its receptors or genes influencing the expression of the IGF2 system. Results: Real-time polymerase chain reaction, immunofluorescence and cell starving experiment...
Budik S, Tschulenk W, Kummer S, Walter I, Aurich C.Live cell RNA imaging has become an important tool for studying RNA localisation, dynamics and regulation in cultured cells. Limited information is available using these methods in more complex biological systems, such as conceptuses at different developmental stages. So far most of the approaches rely on microinjection of synthetic constructs into oocytes during or before fertilisation. Recently, a new generation of RNA-specific probes has been developed, the so named SmartFlare probes (Merck Millipore). These consist of a central 15-nm gold particle with target-specific DNAs immobilised on i...
Newcombe JR, Wilsher S, Silva ESM, Sala-Ayala L, Cuervo-Arango J.Two experiments were conducted in the Northern (UK) and Southern (Brazil) hemispheres to determine the effect of season (month of conception) on the development of supplementary CL (SCL) and the relationship with pregnancy loss. In experiment 1, 199 pregnancies were followed between Day 14 and term, to determine the number of SCL and pregnancy viability (Northern Hemisphere). From the 199 pregnancies, 178 were obtained from inseminations during the breeding season (March-September), while the rest, 21 pregnancies resulted from conceptions in the non-breeding season (October to February). Pregn...
Cortez JV, Hardwicke K, Grupen CG, Herrid M, Machaty Z, Vajta G.A 6-year-old mare, a valuable polo horse, died of complications following postcolic surgery. To preserve its genetics, ear skin samples were collected immediately after death and stored in an equine embryo transfer medium at 4°C for 5 days. After trypsin digestion, monolayer fibroblast cultures were established, but signs of massive bacterial infection were found in all of them. As an ultimate attempt for rescue, rigorously and repeatedly washed cells were individually cultured in all wells of four 96-well dishes. New monolayers were established from the few wells without contamination and us...
Sielhorst J, Koether K, Volkmann N, Blanco M, Vicioso R, Baade S, Kemper N, de Mestre AM, Sieme H.Little is known about the incidence and outcome of high-risk pregnancies in equine practice and clinical studies on spontaneous occurring placentitis cases and treatments are missing. Therefore, the aims of this retrospective field study were to (1) describe the incidence and severity of ultrasonographic assessed placental abnormalities (UPA) in 4,192 pregnancies on a large commercial warmblood stud farm in 2017 - 2019 and (2) characterize these UPA cases and their pregnancy outcome. UPA severity (Placental abnormality score (PSc) 1-3; low to high), nine treatment regimens (TM1-9) used in UPA ...
