Equid semen refers to the reproductive fluid produced by male horses, which contains sperm cells necessary for fertilization. The quality and viability of equid semen are important for successful breeding programs and can be influenced by various factors, including genetics, age, diet, and environmental conditions. Key parameters used to assess semen quality include volume, concentration, motility, and morphology of sperm cells. Techniques for collecting, evaluating, and preserving equid semen, such as cryopreservation, are critical for artificial insemination practices. This page compiles peer-reviewed research studies and scholarly articles that explore the physiology, assessment, and management of equid semen in the context of equine reproduction.
Greiser T, Sieme H, Martinsson G, Distl O.Objectives of the present study were to estimate genetic parameters for frozen-thawed semen traits of 271 fertile German Warmblood stallions and genetic correlations with raw semen quality traits. Semen samples were collected from stallions utilized for semen collection and artificial insemination (AI) on the Lower Saxon National Stud Celle and the North Rhine-Westphalian National Stud Warendorf. Semen quality variables were analyzed in 63,972 raw (gel-free volume, concentration, progressive motility, number of sperm) and 3681 frozen-thawed samples (motility, DNA fragmentation index (DFI), non...
Whitesell K, Stefanovski D, McDonnell S, Turner R.The stallion Breeding Soundness Examination (BSE), as proposed by the Society for Theriogenology, recommends that a stallion produce a minimum of one billion progressively motile, morphologically normal sperm (PMMNS) in the second of two ejaculates collected 1 h apart to be classified as a Satisfactory Prospective Breeder. With this in mind, the first objective of this study was to determine if the classification outcome of the traditional BSE differs depending on the methods used to evaluate sperm motility, morphology and concentration. We hypothesized that application of Computer Assisted ...
Del Prete C, Stout T, Montagnaro S, Pagnini U, Uccello M, Florio P, Ciani F, Tafuri S, Palumbo V, Pasolini MP, Cocchia N, Henning H.During cold storage stallion spermatozoa experience undergo oxidative stress, which can impair sperm function and fertilizing capacity. Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) are the main endogenous enzymatic antioxidants in stallion seminal plasma, and counteract reactive oxygen species. Semen dilution reduces the endogenous antioxidant concentrations. The aim of this study was to investigate whether addition of 15 IU/mL each of SOD, CAT, and GPX to diluted stallion semen would ameliorate a reactive oxygen-mediated decrease in semen quality during 72 h o...
Poirier M, Smith OE, Therrien J, Rigoglio NN, Miglino MA, Silva LA, Meirelles FV, Smith LC.Cell reprogramming by somatic cell nuclear transfer and in induced pluripotent stem cells is associated with epigenetic modifications that are often incompatible with embryonic development and differentiation. For instance, aberrant DNA methylation patterns of the differentially methylated region and biallelic expression of H19-/IGF2-imprinted gene locus have been associated with abnormal growth of fetuses and placenta in several mammalian species. However, cloned horses are born with normal sizes and with no apparent placental anomalies, suggesting that H19/IGF2 imprinting may be epigenetical...
Kumar P, Kumar R, Mehta JS, Chaudhary AK, Ravi SK, Chandra Mehta S, Ansari MM, Legha RA, Tripathi BN, Talluri TR.The present study was designed to study the adverse effects of cryopreservation and evaluation of the cryoprotective effect of reduced glutathione (GSH) and ascorbic acid (AA) supplementation on exotic jack semen in combination or alone. For this, 24 semen samples from four adult and fertile jacks were collected via artificial vagina using an estrus jenny as dummy. After semen collection, the semen was evaluated for various qualitative and quantitative parameters in fresh, cooled, and frozen-thawed semen. The semen pellet was extended with the freezing extender containing either AA (0.9 g/L),...
Papas M, Arroyo L, Bassols A, Catalán J, Bonilla-Correal S, Gacem S, Yeste M, Miró J.This study compared the activities of four antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; glutathione peroxidase, GPX; and glutathione reductase, GSR) in the seminal plasma of stallions and jackasses. Eighteen stallion ejaculates and 24 jack ejaculates were collected through an artificial vagina. Seminal plasma was obtained by several centrifugations at 3000×g and 4 °C for 10 min, and activities of SOD, CAT, GPX and GSR were subsequently determined. We also evaluated whether the collecting season had any influence on the activities of these four enzymes in both stallions ...
Tricyclic antidepressives, such as imipramine, indirectly induce ejaculation by increasing the noradrenaline concentration, which triggers an α-adrenergic response, whereas α-adrenergic agonists, such as xylazine and detomidine, directly trigger ejaculation by activating the α-1 adrenergic receptors. Furthermore, serum oxytocin concentrations in stallions increase drastically before ejaculation, but decline immediately thereafter, implicating the role of this hormone in emission. The objectives of the present study were to: 1) compare the efficiency of various protocols for inducing ex copu...
May CE, Guthrie AJ, Schulman ML.We report herein the use of crude extracts obtained from samples of Taylorella equigenitalis-infected horses for the purpose of multi-locus sequence typing (MLST). Samples (n = 36) were collected from horses in South Africa from 1996 to 2017: 34 from genital swabs (stored at -20°C for 2-3 y) and 2 from cryopreserved raw semen aliquots (stored at -70°C for 18 y) prior to assay. The MLST assay showed a single sequence type (ST), designated ST4, that supported a point introduction and thus a common source for the South African outbreak of contagious equine metritis.
Moraes CR, Runcan EE, Blawut B, Coutinho da Silva MA.The iSperm is a newly released semen analysis tool from Aidmics Biotechnology Co. LTD, which allows an iPad Mini to be transformed into a handheld microscope with objective semen analysis software for equine available through the Apple Store (version 4.5.2). The aim of this study was to compare iSperm values for sperm motility and sperm concentration to current acceptable methods for semen analysis and to determine the agreement with these methods using statistical methods. Two ejaculates from each of five Standardbred stallions were used to compare sperm motility (computer-assisted semen anal...
Wilson M, Williams J, Montrose VT, Williams J.Most stallions within breeding programmes are expected to breed and compete concurrently. The exercising of stallions with regards to training regimes during the breeding season is a controversial subject. Daily exercise at low intensities is important for the mental and reproductive well-being of the stallion, however higher intensities of exercise, as seen in competing stallions, may have detrimental effects on seminal quality. To calculate if competition does affect semen quality, this study investigated the effect that equestrian discipline and timing of competition had on a range of stall...
The aim of this study was to compare the effect of three sugars and Equex paste in a freezing extender for donkey sperm cryopreservation. Ejaculates (n = 18) were collected from six Andalusian donkeys of proven fertility were pooled (two ejaculates per pool) and cryopreserved using a freezing extender containing three different sugars (glucose, fructose and sorbitol), with or without the addition of Equex paste. Sperm quality was assessed before and after freezing-thawing for motility, morphology, plasma membrane integrity, acrosome integrity and DNA integrity. The use of sorbitol in the f...
Kowalczyk A, Czerniawska-Piątkowska E, Kuczaj M.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...
Tsunoda RH, Serrano-Recalde EC, Arruda RP, Oliveira BMM, Pinto SCC, Celeghini ECC.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...
van Dorland A, Janett F, Bruckmaier R, Wach-Gygax L, Jeannerat E, Bollwein H, Sieme H, Burger D.Effects of a plasmolysed yeast product enriched with herbs, malt, honey and orange syrup on semen characteristics and oxidative status in stallions were evaluated. Twenty stallions (mean age ± standard deviation = 9.5 ± 4.5 years) were randomly divided into a treatment group (n = 10) receiving 0.06 mL/kg bodyweight of plasmolysed herbal yeast, and a control group (n = 10) receiving the same amount of placebo daily in the feed for 10 weeks. Ejaculates were collected weekly from all stallions starting at Week 0. Volume, sperm concentration, motility, and velocity were eva...
Sales FA, Ferreira-Silva JC, Vieira JI, Chaves MS, Caldas EL, Filho JP, Freitas VJ, Oliveira MA.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...
Ortiz-Rodriguez JM, Balao da Silva C, Masot J, Redondo E, Gazquez A, Tapia JA, Gil C, Ortega-Ferrusola C, Peña FJ.The population of stallion spermatozoa that survive thawing experience compromised mitochondrial functionality and accelerated senescence, among other changes. It is known that stallion spermatozoa show very active oxidative phosphorylation that may accelerate sperm senescence through increased production of reactive oxygen species. Rosiglitazone has been proven to enhance the glycolytic capability of stallion spermatozoa maintained at ambient temperature. Thus, we hypothesized that thawed sperm may also benefit from rosiglitazone supplementation. Thawed sperm were washed and resuspended in Ty...
Hernández-Avilés C, Love CC, Serafini R, Teague SR, Varner DD.Commonly marketed semen extenders contain various antibiotic types and concentrations to control bacterial growth from stallion's external genitalia. An experiment was conducted to test the effects of supplemental amikacin disulfate (1,000 μg/mL) + potassium penicillin G (1,000 IU/ML: INRA-AP), or ticarcillin-clavulanate (1,000 μg/mL: INRA-TIM) in the INRA 96 extender, on sperm function and antimicrobial activity, compared with extender without supplemental antibiotics (INRA-C). In freshly extended semen (Time 30m), no differences were observed among the three treatment groups for sperm m...
Dini P, Troch L, Lemahieu I, Deblende P, Daels P.The objective of this study was to evaluate the accuracy of a novel, portable device (iSperm Equine for assessing concentration and motility of stallion semen). In the first experiment, semen concentration was determined by the iSperm Equine (Aidmics Biotechnology), Androvision (Minitube) and NucleoCounter SP-100™ (ChemoMetec). The total motility and progressive motility were determined by the iSperm Equine and the Androvision using the manufacturer's guidelines. Frozen/thawed semen samples (n = 33) at various dilutions were analysed for concentration and motility with the above-mentioned ...
Gonzalez-Castro RA, Trentin JM, Carnevale EM, Graham JK.We examined the effects of different freezing extenders, cryoprotectant agents (CPA) and initial thawing temperatures for preparing doses of refrozen stallion sperm for intracytoplasmic sperm injection (ICSI). Single ejaculates, from twelve stallions, were frozen in lactose-EDTA-egg yolk extender (LE) with 5% glycerol. In experiment 1, sperm were initially thawed to 5 °C or 37 °C, before being diluted in LE or skim milk-egg yolk extender (SMEY) containing either 5% glycerol (GLY), 5% methylformamide (MF) or 5% of a combination of both (GMF). In experiment 2, frozen sperm were initially t...
Al-Kass Z, Eriksson E, Bagge E, Wallgren M, Morrell JM.Although artificial insemination (AI) was developed as a means of controlling disease transmission, pathogens can still be transmitted to females in semen used for AI. In addition, bacteria can cause deterioration in sperm quality during storage. Semen becomes contaminated by the male's normal bacterial flora as it passes out of the reproductive tract but potential pathogens may also contaminate the semen. Therefore, semen samples from stallions to be used for AI are tested before the breeding season to minimize transmission of pathogens to inseminated mares. In Sweden, semen samples are teste...
Consuegra C, Crespo F, Dorado J, Diaz-Jimenez M, Pereira B, Ortiz I, Hidalgo M.Sperm vitrification is a rapid freezing method in which carbohydrates are used as cryoprotectants. The aim of this study was to determine the optimal volume, concentration and type of carbohydrates for stallion sperm vitrification using 0.25 ml straws in comparison to conventional freezing. Ejaculates (n = 54) were collected from six stallions. For vitrification, straws were filled with different volumes (30, 70, 100 μl), sperm concentrations (50, 100, 200 × 10 sperm/ml) and extenders containing sucrose (20, 100, 200 mM), trehalose (50, 100, 200 mM) and raffinose (50, 100, 20...
Neuhauser S, Bollwein H, Siuda M, Handler J.Cryopreservation of epididymal sperm allows the saving of genetic material in case of unexpected death or emergency castration. The aim of the present study was the comparison of five different combinations of extenders commercially available for equine frozen semen processing for cryopreservation of epididymal sperm. Epididymal sperm were harvested from gonads of 10 healthy stallions after routine castration by retrograde flush technique. Then, samples were split and diluted with (1) INRA96 + INRA Freeze, (2) BotuSemen + BotuCRIO, (3) EquiPlus + Gent Freeze, (4) EquiPlus + EquiPlus Freeze...
Boye JK, Katzman SA, Kass PH, Dujovne GA.In equids, it is common to inject lidocaine into the testicles at the time of routine castration to provide analgesia. The effects of lidocaine on equine sperm have not been evaluated in vitro or on epididymal sperm collected following castration. The aims of this study were to determine effects of clinically relevant doses of lidocaine on equine spermatozoa in vitro using freshly collected semen and to compare the characteristics of epididymal spermatozoa after routine castration with or without intra-testicular lidocaine administration. We hypothesized that increasing concentrations of lid...
de la Torre J, Crespo F, Arroyo F, Zabal-Aguirre M, Abdoon AS, Gosálvez J.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...
Ghallab ARM, Abou-Ahmed MM, Fad AM, El-Badry DA, Shahat AM, Moawad AR.Cryopreservation of Arabian stallion semen is important in order to improve the function and fertility of frozen/thawed semen in this breed. Objective: To investigate the effects of centrifugation, type of semen extenders, and type of cryoprotectants on the quality of frozen/thawed Arabian stallion spermatozoa. Methods: Semen samples collected from four adult Arabian stallions (one ejaculate per week for 10 consecutive weeks) were either processed directly without centrifugation (no centrifugation; NC) or subjected to centrifugation on the gel-free fraction. Centrifugation protocols were divid...
Sales F, Ferreira-Silva J, Vieira J, Basto S, Filho P, Rocha J, Moura MT, Oliveira M.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...
Martins HS, Martins-Filho OA, Araujo MS, Martins NR, Lagares MA.Frozen equine semen has lower fertility compared to cooled semen. Due to the difficulty to obtain equine oocytes, a heterologous zona pellucida binding assay (ZBA) is an alternative method to predict the fertilizing capability of equine frozen sperm. The rate of capacitated and hyperactivated sperm according to their motility characteristics were analyzed with a Computer Assisted Sperm Analyzer. We believe this report describes for the first time the in vitro hyperactivation induction and the heterologous ZBA to predict the fertilizing ability of frozen equine sperm. Objective: This work aimed...
Delerue M, Breuil MF, Duquesne F, Bayon-Auboyer MH, Amenna-Bernard N, Petry S.Taylorella equigenitalis can be transmitted during artificial insemination. This report describes clinical T. equigenitalis transmission by cryopreserved stallion semen. T. equigenitalis isolates from a mare's vaginal discharge and semen from the same batch of the cryopreserved semen used for the insemination gave identical API ZYM, antibiotic susceptibility, and multilocus sequence typing results (ST-46); furthermore, the multilocus sequence typing lineage ST-46 is known to circulate in the country of semen collection. These results support the need for strict contagious equine metritis scre...
Yasine A, Ashenafi H, Geldhof P, Bekana M, Tola A, Van Brantegem L, Van Soom A, Duchateau L, Goddeeris BM, Govaere J.Trypanosoma equiperdum (T. equiperdum) causes dourine, a venereally transmitted infection in horses. Purification of semen by single layer centrifugation (SLC) has been proven to be successful in reducing venereally transmitted diseases when dealing with other pathogens. The objective of this study was to evaluate the purification of T. equiperdum spiked semen by SLC. Semen was spiked using cryopreserved T. equiperdum stabilates (Dodola strain isolate 943). In total, 6 concentrations, varying from 10 to >5 × 10 trypanosomes, were added to semen samples. Subsequently, SLC was performed ...
Yasine A, Daba M, Ashenafi H, Geldhof P, Van Brantegem L, Vercauteren G, Demissie T, Bekana M, Tola A, Van Soom A, Duchateau L, Goddeeris B, Govaere J.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...
Pasing S, von Lewinski M, Wulf M, Erber R, Aurich C.The aim of this study was to evaluate the stress response of stallions (n = 16) aged 3-13 years with previous sexual experience to semen collection by determination of heart rate, heart rate variability, and cortisol in saliva. Recordings were done on two consecutive days. The time intervals from leaving the box until arrival in the collection barn and from first exposure to the teaser mare until ejaculation as well as the number of mounts until ejaculation were neither affected by collection day nor by age, sexual experience (i.e., the number of breeding seasons the stallion experienced), or ...
Burger D, Dolivo G, Wedekind C.Sperm competition theory predicts semen characteristics to be affected by the social environment. We used the polygamous horse (Equus caballus) to experimentally study within-subject plasticity in response to different social environments. Stallions were sequentially exposed, over a period of 8 weeks each, to other stallions and then singly to mares, or vice versa (in adjacent boxes separated by grills). Ejaculates were collected to determine semen characteristics. Highest sperm numbers were found in stallions that were first exposed to other stallions and then to mares, while lowest sperm num...
Khlifaoui M, Battut I, Bruyas JF, Chatagnon G, Trimeche A, Tainturier D.The cryoprotective effect of l-glutamine and an approach of its mechanism of action, in preserving motility of stallion spermatozoa during the freezing-thawing process, were studied. In Experiment 1, thirty-six ejaculates were collected from six stallions (two good, two middle, and two of poor sperm freezability) and were diluted with 10 different freezing media derived from INRA 82 medium supplemented with 20 mM HEPES and 2% (v/v) centrifuged egg yolk (BM). After thawing, sperm motility was evaluated by a computer-assisted semen motility analyser. The effects of glutamine and glycerol at diff...
Schmidt K, Deichsel K, de Oliveira RA, Aurich J, Ille N, Aurich C.We hypothesized that housing of stallions in a thermoneutral temperature zone during autumn and winter does not only influence metabolism and hair shedding but also improves the characteristics of raw and processed semen. Fertile Shetland pony stallions were followed from October to June. This time coincided with the seasons autumn, winter and spring. Ponies were kept in outside paddocks (group CON, n = 8) or in indoor stables (group ST, n = 8) from October to March when ST stallions returned to outdoor paddocks, but ponies remained in the same groups. The rectal temperature was measured o...
Suliman Y, Becker F, Tuchscherer A, Wimmers K.Horses are seasonal breeders with a natural breeding season beginning in spring and extending through midsummer. In this study, quantitative and qualitative parameters of chilled stallion semen were compared between fertile and subfertile stallions and between the breeding and the non-breeding season. Semen quality parameters compared included ejaculate volume, sperm concentration, total sperm number, sperm morphology, and computer-assisted semen analysis (CASA)-derived sperm movement characteristics obtained from two groups of warmblood stallions ( ; four fertile stallions and four subfert...
Brum AM, Sabeur K, Ball BA.The objective was to evaluate apoptotic markers in ejaculated equine spermatozoa after separation by density-gradient centrifugation and after cryopreservation. Subpopulations of percoll-separated equine spermatozoa differed (P<0.05) in the percentage of live, caspase-activated spermatozoa (2.9+/-0.7% vs 14.2+/-6.4%; mean+/-S.E.M.), low mitochondrial membrane potential (MMP; 6.8+/-1.1 vs 23.8+/-3.7), altered plasma membrane permeability (1.3+/-0.2 vs 3.0+/-0.5), DNA fragmentation (2.0+/-1.3 vs 14.3+/-3.6), total motility (81.8+/-3.3 vs 35.1+/-5.4), and progressive motility (66.3+/-4.3 vs 24...
Morrell JM, Mari G, Kútvölgyi G, Meurling S, Mislei B, Iacono E, Rodriguez-Martinez H.Some stallions produce ejaculates of low quality and/or low fertility when used for artificial insemination (AI). The purpose of these five case studies was to use Single Layer Centrifugation (SLC) to select the best spermatozoa from 'problem' ejaculates for subsequent use in AI. Sperm quality, in terms of motility, morphology and chromatin integrity, was improved in the SLC-selected samples compared to the corresponding uncentrifuged samples, with the exception of one stallion thought to have ampullary stasis. In this stallion, neither the incidence of spermatozoa with detached heads nor the ...
Choi YH, Love CC, Chung YG, Varner DD, Westhusin ME, Burghardt RC, Hinrichs K.We investigated the use of direct nuclear injection using the Piezo drill and activation by injection of stallion sperm cytosolic extract for production of cloned equine embryos. When metaphase II horse oocytes were injected with either of two dosages of sperm extract and cultured 20 h, similar activation rates (88% vs. 90%) and cleavage rates (49% vs. 46%) were obtained. The successful reconstruction rate of horse oocytes with horse somatic cell donor nuclei after direct injection using the Piezo drill was 82%. Four dosages of sperm extract (containing 59, 176, 293, or 1375 microg/ml protein)...
Troedsson MH, Liu IK, Crabo BG.Following the deposition of semen in the mares uterus, spermatozoa must be transported to the site of fertilization, be maintained in the female tract until ovulation occurs, and be prepared to fertilize the released ovum. Sperm motility, myometrial contractions, and a spontaneous post-mating uterine inflammation are important factors for the transport and survival of spermatozoa in the mares reproductive tract. Fertilizable sperm are present in the oviduct within 4 hours after insemination. At this time, the uterus is the site of a hostile inflammatory environment. Our data suggest that sperm...
Morrell JM, Rodriguez-Martinez H, Johannisson A.An improvement in sperm quality after single layer centrifugation (SLC) has been seen in previous studies using small sample sizes (for example, n = 10 stallions). There is a need to investigate whether this improvement is repeatable over several breeding seasons with a larger number of stallions (n ≥ 30 stallions). Objective: To make a retrospective analysis of the results of SLC performed on more than 250 sperm samples (176 ejaculates) from 31 stallions in 3 consecutive breeding seasons. Methods: Sperm quality (motility, proportion of morphologically normal spermatozoa and the proportion o...
Ghosh S, Serafini R, Love CC, Teague SR, Hernández-Avilés C, LaCaze KA, Varner DD.Effects of different media and promoters on lipid peroxidation (LPO) in viable stallion sperm have not been reported. Aims of this study were to determine effects of three media (INRA-96™, Equipro CoolGuard™, and Biggers, Whitten and Whittingham [BWW]), and promoters (iron sulfate-Fe; ultraviolet light-UV; or control-no exposure to promoters) on viable sperm LPO using four different flow cytometric assays (i.e., BODIPY, Liperfluo, 4-hydroxylnonenal [4HNE], malonaldehyde [MDA]). Significant media x promoter interactions were detected using the Liperfluo, 4HNE, and MDA assays (P < 0.05)...
Veeramachaneni DN, Moeller CL, Sawyer HR.Conventional light microscopic evaluation of a seminal ejaculate does not fully avail potential indicators of functional impairment in spermatozoal organelles. The technique of critical quantitative evaluation of morphologic features of individual structural components of spermatozoa at a light microscopic level in conjunction with critical qualitative evaluation of spermatozoal organelles at an ultrastructural level, as described in this article, is a valuable clinical tool. Compared with a battery of sperm function assays used in human andrology clinics, this relatively less expensive and si...
Love CC.Sperm quality assays have increased in number in the last 10 years. Most of these assays are flow cytometry based in application and are modified from assays that have been developed to measure somatic cell function. The goal of any sperm quality assay should be to advance the clinicians/researchers understanding of sperm cell function and the relationship to fertility. While these assays appear to measure somatic cell-like functions in sperm there tends to be little understanding how the results of these assays relate to fertility.
Turner RM, McDonnell SM, Hawkins JF.Ejaculation was pharmacologically induced in a 13-year-old Quarter Horse stallion with a spiral fracture of the radius. The owners desired to have semen from the stallion frozen prior to euthanatizing the horse, but because of the debilitating injury, standard methods of semen collection could not be used. With the stallion standing quietly in a stall, a plastic collection bag was positioned over the stallion's penis, and clomipramine hydrochloride (2.2 mg/kg of body weight, IV) was administered. Fifty-five minutes later, xylazine hydrochloride (0.5 mg/kg, IV) was administered. The stallion ej...
Rota A, Sgorbini M, Panzani D, Bonelli F, Baragli P, Ille N, Gatta D, Sighieri C, Casini L, Maggiorelli MM, Aurich C, Camillo F.Reproductive management of male donkeys employed for artificial breeding has been poorly studied. The aim of this study was to evaluate the effect of housing system, with the animals grouped together in a paddock or kept in individual boxes, on sexual behaviour, cortisol and testosterone concentration and seminal characteristics of adult male donkeys. The study included four Amiata donkey jacks (stallions) from which ejaculates, saliva and blood were collected during two distinct 3 weeks periods, one in the group and one in the box housing system. Time needed for semen collection was shorter ...
Hartwig FP, Lisboa FP, Hartwig FP, Monteiro GA, Maziero RR, Freitas-Dell'Aqua CP, Alvarenga MA, Papa FO, Dell'Aqua JA.During the cooling process, sperm may suffer irreversible damage that compromises the fertility rate. Incorporating cholesterol-loaded cyclodextrin (CLC) represents a strategy to increase sperm resistance at low temperatures; however, high levels of cholesterol in the cell membrane can interfere with sperm capacitation. The goals of this study were to determine the CLC concentration and cooling temperature that produce optimal kinetic parameters and viability of sperm from stallions identified as bad coolers (BCs) and good coolers (GCs), as well as the effect of adding CLC on the occurrence of...
Naden J, Amann RP, Squires EL.Puberty was studied using 15 colts of Quarter Horse phenotype. Total scrotal width was measured every 8 weeks from 48 to 96 weeks. Blood samples were taken from 8 colts at 8, 16 and 24 weeks and then every 4 weeks until 100 weeks to measure changes in LH, FSH and testosterone concentrations. Seminal collections were attempted monthly from 48 to 64 weeks and every 2 weeks thereafter until puberty resumed every 3rd day from 96 weeks for 15 ejaculates. For all collections, times to erection, mount and ejaculation and seminal characteristics were recorded. Age at puberty was defined as the first e...
Ortiz I, Dorado J, Ramírez L, Morrell JM, Acha D, Urbano M, Gálvez MJ, Carrasco JJ, Gómez-Arrones V, Calero-Carretero R, Hidalgo M.The aim of this study was to determine the effect of single layer centrifugation (SLC) using Androcoll-E-Large on donkey sperm quality parameters after 24 h of cool-storage. Ejaculates were collected from Andalusian donkeys and then cooled at 5°C. SLC was carried out after 24 h of cool-storage using Androcoll-E-Large. In the first experiment, all sperm parameters assessed (total and progressive sperm motility, viability, sperm morphology and sperm kinematics VCL, VSL, VAP, LIN, STR, WOB, ALH and BCF) were statistically compared between semen samples processed or not with Androcoll-E-Large. Si...
Bertelsmann H, Keppler S, Höltershinken M, Bollwein H, Behne D, Alber D, Bukalis G, Kyriakopoulos A, Sieme H.The essential trace element selenium is indispensable for male fertility in mammals. Until now, little data existed regarding the relationship between selenium and sperm quality in the stallion. Selenium, or selenium-dependent glutathione peroxidase activity, was determined in red blood cells, semen, seminal plasma and spermatozoa, and the percentages of spermatozoa with progressive motility (PMS), intact membranes (PMI), altered (positive) acrosomal status (PAS) and detectable DNA damage, determined by the sperm chromatin structure assay, were evaluated in 41 healthy stallions (three samples ...
Squires EL, Pickett BW, Amann RP.Five ejaculates were collected at hourly intervals from 32 sexually rested stallions. Gel volume, total seminal volume, sperm concentration and spermatozoa per ejaculate declined (P less than 0.01) from the first to the second or third ejaculate. Gel-free seminal volume or percentage of motile spermatozoa did not vary (P less than 0.05) among ejaculates. Ejaculates from 2- to 3-year-old stallions contained less volume and fewer spermatozoa than those from 9- to 16-year-old stallions. Regardless of the stallion's age the first, first 2, first 3 and first 4 ejaculates represented 50, 74, 86 and ...
Bliss SB, Voge JL, Hayden SS, Teague SR, Brinsko SP, Love CC, Blanchard TL, Varner DD.The objective was to determine if decreased cushion-fluid volume and increased sperm number during centrifugation, or if sperm concentration of extended semen following centrifugation, affected stallion sperm quality. Three ejaculates from each of three stallions were subjected to cushioned centrifugation (1,000g for 20 min). Cushion-fluid volume was set at 1 or 3.5 ml, and sperm number per centrifuge tube was set 1 billion or 3 billion. Following centrifugation, sperm pellets were resuspended in semen extender containing 20% seminal plasma (v/v) with sperm concentrations of 25 or 250 million/...
Kareskoski AM, Reilas T, Andersson M, Katila T.With the aim of investigating properties of stallion seminal plasma to eventually improve semen-handling techniques, sperm motility and plasma membrane integrity were analysed in different fractions of the ejaculates after storage. Semen was collected using a computer-controlled automated phantom that separates the ejaculates into five successive cups. Samples containing seminal plasma and skim milk extender were compared with samples stored in skim milk extender after the removal of seminal plasma by centrifugation. Fractionated ejaculates were stored cooled for 24 h after dilution with exten...
Massányi P, Trandzik J, Nad P, Toman R, Skalická M, Koréneková B.To determine the seminal concentrations of copper, zinc, iron, cadmium, lead and nickel in bulls, rams, boars, stallions and foxes and study their correlations. Methods: Semen samples were obtained, digested and analyzed by means of the atomic absorption spectrophotometer. Data were analyzed statistically with the Student's t-test and Scheffe's test using PC programs SAS and Excel. Results: The seminal copper concentration was significantly higher in ram [(2.49+/-0.18) mg/kg] and fox [(2.16+/-0.53) mg/kg] than that in bull [(1.64+/-0.21) mg/kg], boar [(1.64+/-0.28) mg/kg] and stallion (0.86 mg...
Ball BA, Gravance CG, Wessel MT, Sabeur K.A testis-specific isoform of angiotensin-converting enzyme (ACE) has been identified in a number of mammalian species. The purpose of this study was to characterize the activity of ACE in equine spermatozoa, seminal plasma, and testis. Activity of ACE was determined in seminal plasma, ejaculated and epididymal spermatozoa from mature stallions as well as from pre- and postpubertal testis. The effect of addition of angiotensin II on equine sperm motility was also evaluated. The activity of ACE in detergent extracted sperm plasma membrane was approximately 13-fold higher than that detected in se...
Love CC, Varner DD, Thompson JA.Dismount semen samples were collected from Thoroughbred stallions during the breeding season to determine the magnitude and significance of the variation in sperm morphology within (intra-) and among (inter-) stallions. The degree of intra-stallion variation in sperm morphology during the breeding season was substantial (coefficient of variation: 20%; range 8-56%), but did not affect the day-to-day fertility of sperm samples taken from the same stallion (intra-stallion). This variability did not affect the pregnancy rate of the mares with which each stallion was mated. The degree of interstall...
Quiñones-Pérez C, Martínez A, Ortiz I, Crespo F, Vega-Pla JL.Despite the advances in reproductive technology, there is still a considerable number of low sperm quality cases in stallions. Recent studies in humans have detected several seminal microflora-spermatozoa associations behind some idiopathic infertility cases. However, no studies are available on horses, and there is limited information on the microflora present in stallion ejaculates. Accordingly, the objective of this study was to examine associations to the presence of bacteria families with five sperm quality parameters: concentration, total number of spermatozoa, total and progressive moti...
Jasko DJ.This article outlines a basic method for conducting a stallion semen evaluation. After the removal of the gel fraction of the ejaculate, semen gel-free volume is determined, and any abnormality in appearance is noted. Concentration of sperm cells in semen can be determined with the use of either a hemacytometer or spectrophotometer after appropriate dilution of raw semen. The percentage of progressively motile sperm is evaluated promptly after collection of semen with the use of a phase-contrast microscope. The total numbers of sperm and progressively motile sperm in the ejaculate are calculat...
Equine germplasm bank management involves not only the conservation and use of semen doses, in addition it can also be a resource to study stallion semen quality and after thawing semen properties for reproductive purposes. A possible criterion to measure quality may be based on differential gene expression of loci involved during spermatogenesis and sperm quality maturation. The rapid degradation of sperm after thawing affects the integrity and availability of RNA. In this study we have analyzed genes expressed in equine cryopreserved sperm, which provided an adequate amplification, specifici...
Saaranen M, Suistomaa U, Vanha-Perttula T.Selenium (Se) and glutathione peroxidase (GSH-Px) were determined from the seminal plasma samples and spermatozoa of human and four different animal species. The human sperm Se concentration was 1.8 +/- 0.8 micrograms/g dry weight, which was about half of that in the bull. Abnormal sperm morphology and motility correlated with low sperm Se content. The volume of sperm mitochondrial sheath in human, bull and stallion was measured using transmission electron microscopy. In these species the sperm Se content was highly correlated with the volume of mitochondria. Among the five species studied, th...
