Topic:Artificial Insemination
Artificial insemination (AI) in horses is a reproductive technique involving the collection of semen from a stallion and its subsequent introduction into the reproductive tract of a mare. This method is utilized to facilitate breeding, enhance genetic diversity, and improve the management of equine reproduction. AI can employ fresh, cooled, or frozen semen, each with specific handling and storage requirements to maintain sperm viability. The process requires careful timing to coincide with the mare's estrous cycle, maximizing the likelihood of successful fertilization. This page compiles peer-reviewed research studies and scholarly articles that explore the methodologies, applications, and outcomes of artificial insemination in equine reproduction.
Stored Stallion Sperm Quality Depends on Sperm Preparation Method in INRA82 or INRA96. Removal of seminal plasma facilitates stallion sperm survival during storage, but washing may damage sperm chromatin. Therefore, sperm quality was compared in samples following single-layer centrifugation (SLC) or sperm washing and controls (extension only) in two extenders, INRA82 and INRA96. Ejaculates from six stallions were split among six treatments: SLC, sperm washing, and controls, in INRA82 and INRA96. Sperm motility and acrosome status were evaluated at 0, 24, 48, 72 and 96 hours; morphology at 0, 24, 48, 72 hours and chromatin integrity at 0 and 96 hours, with storage at 6°C. Sperm ...
Addition of caffeine to equine thawed sperm increases motility and decreases nitrite concentration. The aim of this study was to improve the quality of frozen-thawed equine sperm by the addition of caffeine to it. Semen from nine stallions was frozen and different concentrations of caffeine (3, 5 and 7.5 mM) were added to frozen-thawed semen. The sperm kinetic parameters, membrane functionality and integrity, and acrosome integrity and spontaneous acrosome reacted sperm were evaluated with a computer-assisted sperm analysis, a hypoosmotic swelling test and epifluorescent microscopy, respectively. Nitrite and hydroperoxide concentrations of frozen-thawed semen were measured using spectrophot...
Microbiota of semen from stallions in Sweden identified by MALDI-TOF. Stallion semen is known to contain environmental bacteria and normal commensals, and in some cases may contain opportunistic pathogens. These bacteria may negatively influence sperm quality during storage before artificial insemination. The bacteria isolated depend on the culture conditions and method of identification; therefore, the aim of this study was to identify as many of the bacteria present in stallion semen as possible by culturing aliquots of semen under a variety of conditions. Eleven semen samples were available: five extended semen samples from one stud together with a sample of ...
Sperm transport and endometrial inflammatory response in mares after artificial insemination with cryopreserved spermatozoa. This study aimed to determine whether the insemination site and dose with cryopreserved sperm of reproductively normal mares affect the sperm population in uterine tubes and the intensity of endometrial inflammatory response. Experimental subjects were estrous mares inseminated, in the mid-uterine body (Body) or the tip of the uterine horn (Tip), ipsilateral to the dominant follicle, with one 0.5 mL straw with 50 × 106 sperm (50) or with eight straws with 50 × 106 sperm/straw (400). Mares were slaughtered 2 h, 4 h and 12 h after artificial insemination (AI) and randomly assigned to f...
Seminal plasma AnnexinA2 protein is a relevant biomarker for stallions which require removal of seminal plasma for sperm survival upon refrigeration†. Some stallions yield ejaculates that do not tolerate conservation by refrigeration prior to artificial insemination (AI), showing improvement after removal of most of the seminal plasma (SP) by centrifugation. In this study, the SP-proteome of 10 different stallions was defined through high-performance liquid chromatography with tandem mass spectrometry and bioinformatic analysis in relation to the ability of the ejaculates to maintain semen quality when cooled and stored at 5°C. Stallions were classified into three groups, depending on this ability: those maintaining good quality after direc...
Cryopreservation of Semen from Domestic Livestock: Bovine, Equine, and Porcine Sperm. In modern livestock breeding, cryopreserved semen is routinely used for artificial insemination. Sperm cryopreservation allows for long-term storage of insemination doses and secures reproduction at a desired time point. In order to cryopreserve semen, it needs to be carefully processed to preserve its vital functions after thawing. In this chapter, we describe the processes involved in cryopreservation of bull, stallion, and boar sperm. These include preparation of diluents, dilution of sperm in primary and freezing extender, slow cooling from room temperature to 5 °C, packaging of insemina...
Successful Eradication of Taylorella asinigenitalis, Pseudomonas aeruginosa, and Klebsiella pneumoniae Venereal Bacterial Pathogens Using Domestic Steam Disinfection: Implications for AI Practice. Steam disinfection has become established as a trusted method of microbial decontamination; however, there have been no reports on the use of this technology to disinfect equipment used in collection of semen in artificial insemination practice. Hence, it was the aim of this study to examine the survival of Taylorella asinigenitalis, Pseudomonas aeruginosa, and Klebsiella pneumoniae venereal bacterial pathogens using domestic steam disinfection. Sixteen bacterial pathogens from three genera Taylorella, Pseudomonas, and Klebsiella each at an inoculum density of approximately 1.5 × 107 colony-...
Efficiency of Semen Cryopreservation in Stallions. Differences in the cryotolerance of spermatozoa exist among stallions, but it remains to be determined to what extent such differences are affected by breed. In this study, post-thaw semen quality in stallions presented for semen cryopreservation was analysed retrospectively (1012 ejaculates from 134 stallions of 5 breeds). The percentage of frozen-thawed ejaculates acceptable for artificial insemination (AI) and the number of insemination doses per cryopreserved ejaculate was calculated. Logistic regression analysis revealed sperm motility in raw semen as the most important explanatory variab...
Seminal Plasma, Sperm Concentration, and Sperm-PMN Interaction in the Donkey: An In Vitro Model to Study Endometrial Inflammation at Post-Insemination. In the donkey, artificial insemination (AI) with frozen-thawed semen is associated with low fertility rates, which could be partially augmented through adding seminal plasma (SP) and increasing sperm concentration. On the other hand, post-AI endometrial inflammation in the jenny is significantly higher than in the mare. While previous studies analyzed this response through recovering Polymorphonuclear Neutrophils (PMN) from uterine washings, successive lavages can detrimentally impact the endometrium, leading to fertility issues. For this reason, the first set of experiments in this work inten...
Practical experience with artificial insemination (AI) using fresh chilled and frozen semen in mares. The objective of this study was to compare the efficiency of artificial insemination (AI) carried out with frozen and fresh, diluted and chilled semen under field conditions. One hundred and twenty-nine mares of different breeds were included in the study. Eighty-one out of the 107 mares inseminated with fresh, chilled semen got pregnant. Seven pregnant mares aborted and 74 foals were born. Out of the 22 mares inseminated with frozen semen, 17 mares got pregnant. Two mares out of the 17 pregnant mares aborted and finally 15 healthy foals were born. No difference was found between the two group...
Identification and quantification of coding and long non-coding RNAs in stallion spermatozoa separated by density. It is not unusual for stallions to have fertility problems. For many, artificial insemination with more dense spermatozoa (isolated by density gradient centrifugation) results in greater pregnancy rates compared with the rates when using unfractionated spermatozoa. RNAs in spermatozoa delivered to the oocyte at conception are required for embryo development. Novel molecular assays of spermatozoa that reflect function are needed to predict the fertility of stallions. To describe and compare the RNA populations in more dense and less dense spermatozoa from stallions. Spermatozoa from five stalli...
Preservation of Epididymal Stallion Sperm in Liquid and Frozen States: Effects of Seminal Plasma on Sperm Function and Fertility. Three separate experiments were conducted to improve preservation of stallion epididymal sperm. In the first one, two different cooling extenders (Kenney and Gent) were compared. Sperm viability and motility patterns were assessed in 10 different epididymal sperm samples after 0 hours, 24 hours, 48 hours, 72 hours, and 96 hours of preservation at 4°C. No significant differences were observed in any of the evaluated parameters either between extenders or throughout the storage period. The second set of experiments was designed to determine whether supplementing thawing medium (INRA Freeze...
Birth of first foals through embryo transfer after artificial insemination using frozen semen in Japan. Until now, there have been no reports of foals born through embryo transfer after artificial insemination using frozen semen in Japan. The aims of this study were to develop a riding crossbred horse and evaluate the prospects of embryo transfer technology in multiplying horse population. In both donor and recipient mares, luteolysis was induced by the administration of 0.1 mg Cloprostenol to synchronize the onset of estrus, and ovulation was induced by administering 2000 IU human chorionic gonadotropin (hCG) or 0.75 mg Deslorelin. Frozen semen from an Irish Connemara pony stallion was used to ...
Effect of Timing of Postovulatory Insemination Relative to Human Chorionic Gonadotropin/Buserelin Treatment With 1 Straw of Frozen-Thawed Semen on Mare Fertility. The reproductive management of mares for frozen semen artificial insemination (AI) can be costly and labor intensive. Predicting the exact time of ovulation can be challenging even when ovulation-inducing drugs are used. The main objective of this retrospective study was to determine whether there was an effect of interval between examinations to detect ovulation on likelihood of pregnancy and early embryonic loss in mares after postovulatory breeding with a single straw of frozen/thawed semen. The second objective was to determine the efficacy of two different drugs (human chorionic gonadotro...
Stallion Sperm Integrity After Centrifugation to Reduce Seminal Plasma Concentration and Cool Storage for 4 days. The objective of the study was to investigate if reducing the seminal plasma of stallion extended semen by centrifugation once will suffice to maintain acceptable semen quality for insemination after 4 days of cool storage. Collected semen was extended to 25 × 106 sperm/mL and subjected to one of the following treatments: noncentrifuged (control), centrifuged for 10 minutes at 900 × g and 1800 × g. The supernatant was partially removed, and the sperm pellet, reconstituted and re-extended. It was then placed in a passive cooling device overnight and then transferred to a refrigerator fo...
Relationships between antral follicle count, blood serum concentration of anti-Müllerian hormone and fertility in mares. The anti-Müllerian hormone (AMH) plays an inhibitory role during folliculogenesis by regulating the number of follicles entering the growing pool. Antral follicle counts (AFC) are highly correlated with serum AMH concentrations and both appear to be related to the ovarian reserve in several species. Few data on AMH and AFC in mares exist, especially with regard to fertility. Therefore, the objective of the current study was to investigate the interrelationship between antral follicle count, serum AMH concentrations and fertility outcome in mares. One hundred and twenty-seven mares were enroll...
Breed and stallion effects on frozen-thawed semen in warmblood, light and quarter horses. The objectives of the present study were to analyze systematic effects on semen quality traits from fresh and frozen-thawed semen collected in and outside season. A total of 4,681 reports on semen traits of 121 stallions representing Arabian, Thoroughbred, Quarter Horse and four warmblood breeds used for artificial insemination at the Lower Saxon National stud Celle were edited for analysis of gel-free volume, sperm concentration, total number of sperm and total number of motile sperm in fresh semen and in frozen-thawed semen progressive motility, DNA fragmentation index and non-viable sperm. ...
Factors Influencing the Popularity of Artificial Insemination of Mares in Europe. The purpose of this review was to analyze factors affecting the popularity of artificial insemination of mares in Europe in the context of sperm quality. Taking into account the prices of stallion semen on the world market, efficiency is important for the profitability of its use in artificial insemination programs in Europe. To increase the efficiency of a semen insemination facility, it is necessary to correctly and objectively assess the quality of semen. The available range of tools allows an effective evaluation of the potential fertility of a stallion. For several years, artificial insem...
Effects of pentoxifylline supplementation to semen extender on post-breeding inflammation response assessed by endometrial cytology and vascular perfusion in mares. The aim was to evaluate effects of addition of pentoxifylline to skimmed milk semen extender on uterine inflammatory response. Thirty-six estrous cycles of 15 mares were randomly divided into five groups for artificial insemination (AI): Control: mimicking the AI procedure (n = 7); Extender: deposition of skimmed milk based extender (n = 7); Extender + PTX: skimmed milk based extender plus pentoxifylline (7.18 mM; n = 8); Semen: semen diluted with extender without pentoxifylline (n = 7), and Semen + PTX: semen diluted with extender containing pentoxifylline (n = 7). M...
In Vitro and In Vivo Efficiency of Extenders Added to Thawed Stallion Semen. Addition of extenders to thawed semen could improve fertility. Objective: To determine the efficiency of extenders to increase viability of thawed semen, measured by sperm parameters in vitro and pregnancy rates after artificial insemination (AI). Methods: Sperm motility and acrosin activity were measured during a thermoresistance test (TRT). Results: Progressive motility decreased (P<0.05) after 30 min in thawing semen treated with saline solution (SS) and only after 60 min with Tyrode's solution (TS) or freezing diluent (FD). The total motility decreased (P<0.05) after 60 min in thawed semen...
Periovulatory administration of firocoxib did not alter ovulation rates and mitigated post-breeding inflammatory response in mares. Non-steroidal anti-inflammatory drugs (NSAIDs) are a therapeutic option for the treatment of inflammation. However, negative effects of non-selective NSAIDs for treatment of mares with endometritis have been described, including delayed uterine clearance and impairment of ovulations. Firocoxib is a specific cyclooxygenase-2 (COX-2) inhibitor and has the ability to act in the uterus of mares. We investigated the effects of firocoxib on ovulation rate, numbers of polymorphonuclear neutrophils (PMNs), and COX-2 protein levels in the endometrial tissue of susceptible mares after insemination. Two ...
Effect of sperm dosage transportation in stallions: Effect on sperm DNA fragmentation. Artificial insemination programs for horses usually involve ex vivo handling and transporting of sperm. The present experiment was designed to: (i) assess the effect of transportation on sperm DNA integrity at different time post semen collection, and (ii) evaluate if sperm DNA quality deteriorates rapidly beyond 24 h of cooled storage. After collection, the ejaculates were extended using INRA 96 and semen was prepared for prompt analysis (A0) or 24 h/48 h cooled-shipping (B24 and C48 respectively). Each sample was assessed for sperm DNA fragmentation index (SDFI) at time 0 and after inc...
Freezing of Stallion Semen: in vitro and in vivo Evaluation of Sperm Motility and Acrosin Activity in Frozen-thawed Semen with Addition of Post-diluent Extenders. Post-diluents could potentially increase semen cryotolerance, but remain poorly explored in horses. The aim was to evaluate the efficiency of post-diluents on frozen-thawed semen viability of two stallions (S1-S2). The cryopreserved semen was thawed at 50°C for 40 seconds. Semen motility and acrosin activity (AA) were determined during the thermo-resistance test (TRT). Progressive motility of S2 semen decreased after 60 and 90 minutes of TRT (TRT60 and TRT90) on the control compared to both post-diluents. The total motility of both S1 and S2 decreased on TRT60 and TRT90 semen control versus b...
Tissue (re)distribution of Trypanosoma equiperdum in venereal infected and blood transfused horses. Dourine, caused by Trypanosoma equiperdum, is a life-threatening venereal disease in equidae. So far, there is no clear evidence on how and when stallions become infectious, nor which tissues are affected by the parasite in diseased animals. Post-infection, after a transient, temporary phase of parasitaemia, the parasite disperses to different tissues in an unknown distribution pattern. This study describes the distribution of the parasite after infection by artificial insemination (AI) or blood transfusion. Mares (N = 4) were artificially inseminated with T. equiperdum spiked semen wherea...
Effects of flunixin meglumine on postponement of ovulation in mares. OBJECTIVE To evaluate use of flunixin meglumine as a treatment to postpone ovulation in mares, mare fertility after flunixin meglumine treatment during estrous cycles, and effects of flunixin meglumine on function of the corpus luteum after ovulation. ANIMALS 13 healthy mares. PROCEDURES A single-blinded, placebo-controlled, crossover study was conducted. Flunixin meglumine (1.1 mg/kg, IV, q 24 h) or lactated Ringer solution (placebo treatment) was administered for 2 days to mares with a dominant follicle (≥ 35 mm in diameter) and behavioral signs of estrus. Mares then were bred by artificia...
Increase of pregnancy rate after multiple periovulatory inseminations in mares. There exist differences in the reproductive behavior of stallions and mares under free-running and domestic in-hand breeding conditions. Contrary to artificial insemination programs, a stallion mates a mare multiple times per estrus under natural conditions. The objective of this study was to determine if multiple periovulatory artificial inseminations (MI), four times in two different time intervals instead of two, would result in increased pregnancy outcome or higher incidences of breeding induced endometritis. Methods: Eighty-two estrous mares were allocated randomly to one of three experim...
Altered gene expression in embryos and endometrium collected on day 8 of induced aluteal cycles in mares. Aluteal cycles were induced in the mare to evaluate the effects of progesterone deprivation on the gene expression of embryos and endometrium collected eight days after ovulation. We hypothesized that the transcript expression would be altered during induced aluteal (AL) cycles (low progesterone <1 ng/mL) when compared with control cycles during diestrus (high progesterone; > 4 ng/mL) for 1) the embryonic expression of progesterone-mediated transcripts and those related to normal embryo growth and development and 2) the endometrial expression of progesterone-mediated transcripts and those ...
Analysis of factors affecting the pregnancy rate of mares after inseminations with cooled transported stallion semen. Artificial insemination (AI) with cooled stallion semen has increased markedly during the last decades in all countries, but fertility is often lower than when fresh semen or natural mating is used. The objective of this study was to examine field data (1634 ,cycles 523 Standardbred (SB) mares, 575 Finnhorse (FH) mares, and 90 stallions) using multivariable logistic regression for factors influencing the pregnancy rate (PR) after AI with cooled transported semen from SB and FH stallions. The PR per cycle for the material was 47%: Finnhorses 42% and Standardbreds 53%. When assessed with multiva...
Successful embryo transfer from Hokkaido native pony after artificial insemination with frozen semen. There has been no report of equine embryo transfer in Japan for the last 24 years. Our objective was to establish an effective protocol for embryo transfer in domestic horse. A Hokkaido native pony was bred by deep-horn artificial insemination with frozen semen from a Connemara pony. Embryo collection was performed using a non-surgical method on day 7. Two embryos were obtained from three flushes (67% recovery) and were transferred fresh into crossbred recipient mares. Both recipient mares were diagnosed pregnant (100% pregnancy rate) 5 days after embryo transfer and had normal progesterone le...
Effect of Using Two Cryopreservation Methods on Viability and Fertility of Frozen Stallion Sperm. Studies involving different methods and techniques of cryopreservation and its interactions with the conception rates in artificial insemination (AI) programs are reported in the literature. This study evaluated the sperm kinetics, plasma membrane integrity, and fertility rates of mares inseminated with cryopreserved stallion semen subjected to different freezing methods. For this, four ejaculates from five stallions were collected and frozen in conventional (Styrofoam box) or automated system in Mini-Digitcool ZH 400. Seminal samples were evaluated after thawing for sperm motion parameters by...