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Topic:Embryo Transfer
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.
Efficiency of transvaginal oocyte aspiration (TVA), post-mortem ovary harvest (OH), and Intracytoplasmic Sperm Injection (ICSI) in a Quarter Horse mare population in North America. In a clinical setting, Intracytoplasmic Sperm Injection (ICSI)-derived equine blastocysts can be obtained either by transvaginal oocyte aspiration (TVA) or post-mortem ovary harvest (OH). Most peer-reviewed retrospective reports have described the suitability of these techniques for obtaining blastocysts, pregnancies, and live foals, particularly in Warmblood horse populations in Europe. In this study, retrospective data from Quarter Horse client-owned mares subjected to TVA-ICSI (n = 105; 151 cycles) or OH-ICSI (n = 42) at an academic institution in North America between 2023 and 2025...
Achieving Success in Conventional Equine Embryo Transfer: A Practical Review. Equine embryo transfer is a well-established procedure in horse breeding programs and it's used to increase the number of offspring from valuable mares, obtain embryos from mares in competition and produce foals from mares unable to carry a pregnancy to term. The success of embryo recovery depends mainly on the day of flushing, management of insemination, and the fertility of donor mare and stallion. Other relevant factors include donor age and reproductive status, number of ovulations, operator experience, and flushing technique. Pregnancy rates after transfer are influenced by donor-recipien...
Cracks in the ice: biological challenges and incremental progress in equine oocyte vitrification. Equine oocyte vitrification could enhance the flexibility of in vitro embryo production, yet developmental competence remains reduced. Only vitrification of oocytes matured in vivo yields foaling rates comparable to fresh oocytes. Recent advances in vitrification at the germinal vesicle (GV) stage, supported by systematic comparisons of cryoprotective agents and exposure times, has enabled consistent blastocyst production and birth of several foals. Comparative studies of GV and metaphase II (MII) stages suggest biological advantages of vitrification after in vitro maturation (IVM), warranting...
Application of Anti-Müllerian Hormone in Regulation of Ovarian Function and Its Practical Relevance for Fertility and Embryo Production in Cows and Mares. In the female organism, Anti-Müllerian hormone (AMH) is produced exclusively by granulosa cells, particularly in secondary and early tertiary follicles. The amount of AMH released into the blood is proportional to the total number of follicles present in the ovaries and is therefore used to estimate the ovarian functional reserve. In both cattle and mares, a strong and highly repeatable association has also been demonstrated between AMH concentrations and antral follicle count. Therefore, AMH may serve as a biomarker with broad diagnostic utility in reproductive medicine in these species, as ...
Vitrification of in vivo produced equine embryos: Size really does matter! Vitrification rather than slow-freezing has become the method of choice for the cryopreservation of in-vivo produced embryos. It is a simple technique that does not require a programmable freezing machine and relies chiefly on placing the embryo on a surface, rather than within a straw, in a small volume of vitrification solution to achieve extremely rapid cooling rates. However, in the field of equine embryo vitrification size truly does matter. Methodologies for both vitrification and warming prior to transfer are largely dictated by the size of the embryo. Since embryo size is directly rela...
Embryo developmental kinetics and vitrification in relation to pregnancy outcome, foaling percentage and foal sex. While fast in vitro embryo development has been associated with improved outcomes, the relationships of slow in vitro embryo development and vitrified versus fresh in vitro produced (IVP) embryo transfer to foaling percentage and foal sex are less explored. Objective: To determine the relationships of (1) day of blastocyst formation (D7-11 after intracytoplasmic sperm injection (ICSI)) and (2) vitrification and warming method (one- and three-step) versus fresh transfer with pregnancy, early pregnancy loss (EPL), foaling percentage and foal sex. Methods: Retrospective clinical study. Methods: B...
Conventional IVF in the horse past, present, and future: physiological and technical insights. For decades, conventional in vitro fertilization (IVF) in the horse was ineffective, leading to reliance on intracytoplasmic sperm injection (ICSI) for both research and clinical in vitro embryo production. However, the ability to achieve IVF is essential to the study of key reproductive processes, including sperm capacitation, fertilization, and early embryonic development. Recently we have established a repeatable and efficient IVF system for horses, using long-term (9-22 h) sperm pre-incubation. For clinical use, IVF has many features that may support higher embryo quality than does ICSI, a...
Molecular detection of pathogenic Leptospira spp. in mares with reproductive disorders enrolled in embryo transfer programs. Equine genital leptospirosis (EGL) is associated with reproductive disorders in mares. However, its occurrence in embryo transfer (ET) programs remains poorly characterized. Objective: Given the scarcity of data on the circulation of Leptospira spp. in mares enrolled in ET programs, this study aimed to investigate the serological and molecular occurrence of Leptospira spp. in mares with a history of reproductive disorders from 13 stud farms dedicated to embryo transfer in Brazil. Methods: Serum and endometrial cytobrush samples (ECS) were collected from 70 unvaccinated mares. Serological analy...
Age, embryo donor status, and insemination with chilled semen all predispose to persistent breeding-induced endometritis in warmblood mares. Persistent breeding-induced endometritis (PBIE) is a common reason for mares failing to establish pregnancy. Objective: To examine the hypothesis that advancing mare age and embryo donor status predispose to PBIE and compromise fertility. Methods: Retrospective analysis of the impact of mare age, breeding system (embryo donor versus broodmare), and semen type (chilled versus frozen-thawed) on the likelihood of PBIE and subsequent pregnancy/embryo recovery. Methods: Warmblood mares (n = 769) inseminated during 1745 oestrous cycles were divided into three age categories (≤6, 7-13 and ≥14...
Equine Oocyte and Embryo Metabolism in the ART Setting: Current Knowledge and Clinical Significance. Equine in vitro embryo production has become very efficient and is used extensively worldwide for the clinical production of foals. What we know so far about offspring health and performance is promising; however, elements of the in vitro process remain unphysiological when compared to in vivo conditions. Studying the metabolism of oocytes and embryos can help inform the optimisation of culture systems and identify aberrations if they occur. Given the growing body of knowledge on the downstream effects of an inadequate peri-conception environment, both in vivo and in vitro, it is imperative th...
Comparative analysis of the transcriptome and methylome in distinct trophoblast populations at the time of embryo implantation in mares. BACKGROUND: During equine implantation, two distinct trophoblast subpopulations are identified: the invasive chorionic girdle (CG) and the non-invasive allantochorion. The invasive CG trophoblast is believed to modulate the maternal immune response and facilitates embryo attachment to the endometrium, while the allantochorion contributes to placental development without breaching the endometrium. However, the mechanisms underlying differential adhesion and immune modulation by these two lineages remain poorly understood. This study aimed to investigate the regulation of biological processes un...
Impact of donor mare and stallion identity on the likelihood of pregnancy after transfer of in vitro produced embryos. The success of in vitro embryo production (IVEP) is influenced by donor mare and stallion. Objective: To determine whether donor mare and stallion influence the pregnancy rate after transfer of in vitro produced (IVP) blastocysts and to identify factors influencing the likelihood of obtaining one or more pregnancies from a single IVEP session. Methods: Retrospective clinical study. Methods: The influence of the following variables on pregnancy outcome was assessed in 2292 transfers of IVP blastocysts: speed of in vitro embryo development, embryo grade after thawing, age of the donor mare, blas...
Sex determination and genetic screening of equine embryos with whole genome amplification and real-time PCR. Reliable pre-implantation sex determination and genetic screening enables informed embryo transfer decisions in equine breeding while avoiding later interventions. We developed a streamlined workflow that couples rapid whole genome amplification (WGA) with a multiplex real-time PCR targeting ETSTY5 as a Y-specific marker and UBC as an autosomal control. On purified equine DNA, sex was correctly assigned down to 10 pg gDNA and to a single fibroblast cell. Direct testing of embryo biopsies without WGA yielded inconsistent results, whereas introducing a short WGA step produced tight allelic-discr...
Clinical pregnancy outcomes after transfer of fresh and vitrified-warmed conventional IVF equine embryos. In vitro fertilization (IVF) in horses has emerged as a valuable assisted reproductive technology, yet the impact of embryo cryopreservation on pregnancy outcomes when utilizing this technique remains unclear. This study reports pregnancy outcomes following transfer of conventional equine IVF embryos, with both fresh and vitrified-warmed embryos. Conventional IVF (cIVF) was used to produce 40 embryos, which were transferred to recipient mares, pregnancy diagnosis was performed at 6 days post-transfer followed by embryonic heartbeat detection between day 25 and 30 of gestation. Observed pregnan...
Embryonic vesicle diameter at first pregnancy diagnosis in in vivo- and in vitro-derived pregnancies and its relationship with pregnancy loss in the mare. This retrospective study compared the relationship between embryonic vesicle diameter at first pregnancy diagnosis and early embryonic loss (EEL) across three pregnancy types: artificially inseminated broodmares (AI; n = 263), recipients carrying in vivo-derived flushed embryos (IVD-F; n = 312), and recipients carrying in vitro-produced embryos by intracytoplasmic sperm injection (IVP-ICSI; n = 161). Vesicle diameter was recorded at Day 12 post-ovulation in AI, Day 4 post-embryo transfer (ET) for IVD-F, and Day 7 post-ET for IVP-ICSI pregnancies. Mean (±SD) vesicle diameters and E...
Deficiency in homozygous haplotypes reveals recessive lethal variants affecting fertility and viability in the Friesian horse. BACKGROUND: Recessive lethal alleles causing pre- or postnatal death in homozygous mutant animals, could lead to reduced fertility success. The Friesian horse breed has signs of reduced fertility and has faced high inbreeding rates in the past (∆F > 1%). Consequently, by genetic drift lethal alleles may have reached moderate to high frequencies in the population. Our aim was to identify lethal recessive alleles that — when homozygous — may cause pre- or postnatal death in the Friesian horse. RESULTS: We analyzed genotypes (70 K SNP) of over 8,000 Friesian horses, looking for hapl...
Does the Reproductive Technique Affect Neonatal Health Parameters in Foals? This retrospective study evaluated whether different reproductive biotechnologies influence neonatal behaviour parameters in foals. Data from 102 foalings in two commercial breeding farms were analysed, including foals conceived by artificial insemination (AI), conventional embryo transfer (ET) and intracytoplasmic sperm injection (ICSI). Neonatal parameters recorded were birth weight, time to achieve sternal recumbency, time to stand, time to nurse and time to meconium elimination. Mixed statistical models were applied to assess the influence of reproductive technique, breed, sex and farm. Si...
Long-term micronutrient and amino acid supplementation increases embryo recovery per flush and ovarian responsiveness in donor mares. Background: Nutritional strategies may modulate ovarian function and embryo recovery in donor mares, but field evidence for long-term multinutrient supplementation remains limited.Aims/objectivesTo determine whether long-term micronutrient and amino acid supplementation increases embryo recovery per flush and ovarian responsiveness in donor mares.Methods: A total of 115 uterine flushes were performed under a crossover design across two consecutive breeding seasons in 12 multiparous, non-lactating crossbred donor mares (4-12 years). Mares received a basal diet or the same diet plus 40 g/day of ...
Temporal characterization of conditions that promote functional capacitation of stallion sperm. Equine in vitro fertilization (IVF) has historically been challenged due to lack of conditions that promote functional capacitation of stallion sperm. Capacitation success is often attributed to prolonged sperm incubation requirements. Advances to equine IVF are hindered by incomplete characterization of conditions that promote the functional capacitation of stallion sperm over time. Herein, we describe conditions that promote the functional capacitation of fresh, cooled, and frozen-thawed stallion sperm isolated by Percoll, Swim-Up, or Microfluidics (VetMotl). Fresh (diluted sperm) and previo...
Metabolomics reveals early predictors of blastocyst formation in equine ICSI-derived embryos. Equine in vitro embryo production (IVP) via ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) has gained prominence in sport-horse breeding due to high success rates, efficient semen utilization, and year-round embryo production, including from mares with fertility challenges or postmortem. However, assessing viability of early-stage ICSI-produced equine embryos remains challenging due to subjective morphological evaluations and limited visibility of developmental features. Non-invasive metabolomic analysis of culture media offers a promising alternative to enhance embryo selectio...
Treatment of luteal phase mares with the aromatase inhibitor letrozole does not change characteristics of the estrous cycle, ovarian function, and embryo characteristics. Letrozole is a potent and highly selective third-generation aromatase inhibitor. In the present study, we aimed to investigate the effects of letrozole treatment of non-pregnant mares on estrous cycle characteristics. Cyclic, non-lactating mares (n = 9) were included. After estrus synchronization, mares were monitored to detect ovulation (d0:) of the treatment cycle. Each three mares were randomly assigned to one of three treatments: control, 0.5, and 1 mg/kg body weight letrozole in three consecutive cycles (d7 to d13 of the treatment cycle). Mares were inseminated in the estrus after trea...
Incidence of spontaneous ultrasonographic regression of the corpus luteum in pregnant embryo recipients and broodmares: a field study. Early embryonic loss (EEL), particularly between Days 14 and 42 of pregnancy, remains a key concern in equine reproduction. This study investigates the incidence of spontaneous luteolysis-used here as a marker of maternal recognition of pregnancy (MRP) failure-in both pregnant Thoroughbred (TB) broodmares and recipient mares carrying either in vivo-derived (IVD) or in vitro-produced (IVP) embryos. Retrospective reproductive data from 2078 pregnancies in TB and 532 pregnancies in recipients were analysed to determine the incidence and characteristics of clinical luteolysis during pregnancy. Ove...
The efficacy of embryo recovery on consecutive days from Day 6.5 to obtain small embryos for vitrification. Vitrified embryos ≤300 μm give better pregnancy rates following warming and transfer than larger ones. Embryo recovery undertaken close to when the embryo enters the uterus (Day 6-6.5) helps in the recovery of embryos ≤300 μm. However, flushing early can mean missing an embryo not yet in the uterus, whereas later can result in embryos >300 μm. Objective: To evaluate if repeated embryo flushing on consecutive days from Day 6.5 would increase the number of embryos ≤300μm recovered. Methods: Retrospective case series. Methods: Four hundred and ninety-six inseminations with cool...
Plasma Estradiol and Endometrial Edema Profile in Acyclic Mares After Single Administration of 17-β Estradiol, Estradiol Benzoate and Estradiol Cypionate. A longer endometrial exposure to estradiol before progesterone has been shown to be beneficial in cyclic and acyclic recipient mares. Therefore, the selection of an estradiol ester that promotes longer endometrial exposure to estradiol using a single administration would be advantageous when preparing acyclic mares as embryo recipients. This study investigated plasma estradiol profiles in acyclic mares after a single administration of 17-β estradiol (17-β), estradiol benzoate (EB) and estradiol cypionate (EC), and the correlation between plasma concentrations and endometrial edema. Fifteen n...
Abnormal cleavage patterns in equine in vitro-produced embryos lead to higher early pregnancy loss. Despite significant advances, in vitro production (IVP) of equine embryos continues to lack standardised embryo classification criteria and is associated with increased rates of early pregnancy loss compared with in vivo-derived blastocysts. Objective: To evaluate morphokinetic characteristics of the first mitotic division and early embryonic development in IVP blastocysts and their association with embryo development, as well as pregnancy rate and early pregnancy loss following embryo transfer. Methods: Retrospective analysis of archived material and clinical records. Methods: We retrospectiv...
A comparison of the efficacy of three commercial human embryo vitrification kits for cryopreservation of in vivo produced equine embryos. Different cryoprotectants can influence the ability of embryos to successfully survive vitrification and subsequent warming before transfer. Objective: To compare pregnancy rates for embryos ≤500 μm vitrified, without puncture or aspiration of the blastocoele cavity, with one of three commercial human embryo vitrification kits containing the same penetrating cryoprotectants (DMSO and EG) but varying in their non-penetrating cryoprotectants (NPCPAs; sucrose, trehalose, dextran serum supplement [DSS], and hydroxypropyl cellulose [HPC]). Methods: In vivo experiments. Methods: Embryos (n =...
Follicular fluid secretome enhances the meiotic competence of in vitro matured equine oocytes, but not the developmental competence of zygotes after intracytoplasmic sperm injection (ICSI). In vitro embryo production is widely used in equine clinical reproduction; however, its efficiency remains suboptimal due to the undermined developmental competence of in vitro-matured oocytes. In this work, we evaluated the effect of mare preovulatory follicular fluid (PFF) secretome supplementation during in vitro maturation (IVM) of equine oocytes recovered post-mortem. Secretome was isolated from PFF and was added to the IVM medium at concentrations of 20 µg/ml (S20) or 40 µg/ml (S40). The percentage of mature oocytes, as well as embryo cleavage, blastocyst development, and gene expr...
Biphasic CAPA-IVM Improves Equine Oocyte Quality and Subsequent Embryo Development Without Inducing Genetic Aberrations. In vitro maturation (IVM) of oocytes retrieved from ovum pick-up (OPU) or ovarian tissue (OT) is a standard approach for patients with specific conditions where prior hormonal stimulation is contraindicated. However, the developmental competence of oocytes matured in vitro is still inferior to that of oocytes matured in vivo. Capacitation IVM (CAPA-IVM) includes an extra step of pre-maturation culture (PMC) with c-type natriuretic peptide (CNP) as a meiotic arrestor to better synchronize cytoplasmic and nuclear maturity in oocytes by allowing the cytoplasm additional time to acquire essential ...
In vitro embryo production via ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) in pure and crossbred Japanese Hokkaido native ponies. This study evaluated the viability of in vitro embryo production using ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) as breeding techniques for pure and crossbred Hokkaido native ponies (n = 9). Oocytes were collected using transvaginal ultrasound-guided follicle aspiration. ICSI was performed on in vitro matured oocytes using frozen semen. Embryonic cultures were monitored using time-lapse cinematography. Blastocysts were cryopreserved and, after thawing, were transferred non-surgically into recipient mares. Over nine OPU sessions, the mean number of aspirated follicles was 2...
The Influence of Aspiration Pressure, Follicle Flushing Method and Needle Rotation During Single-Operator OPU Technique on Oocyte Recovery and Embryo Production in the Mare. The objective of this study was to determine the effect of two aspiration pressures (75 vs. 150 mmHg), the follicle flushing method (injection pump controlled by a foot pedal vs. a plastic syringe) and the twisting of the OPU needle on oocyte recovery and in vitro embryo production. OPU data from a total of 104 warmblood sport mares belonging to a commercial OPU-ICSI program were collected as part of a prospective study split into three experiments. Each mare was used only once for OPU. In Experiment 1, the mares' follicles were aspirated using either a high aspiration pressure (flow rate of 1...
