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.
Sotgia S, Taras A, Zinellu A, Cherchi R, Mangoni AA, Carru C, Bogliolo L.The dependence of a stallion's spermatozoa on oxidative phosphorylation for energy requirements results in an unconventional relationship between reactive oxygen species (ROS) production and fertility. In such a scenario, antioxidant activity must be finely controlled and not affect the essential functions of ROS. Some in vivo evidence suggests that the naturally occurring antioxidant ergothioneine (ERT) interferes with the critical roles of ROS/reactive nitrogen species (RNS) in pro-oxidant states but not in healthy tissues. The measurement of ERT in seminal plasma collected from 14 stallions...
Bubenickova F, Postlerova P, Simonik O, Sirohi J, Sichtar J.Seminal plasma (SP) is the natural environment for spermatozoa and contains a number of components, especially proteins important for successful sperm maturation and fertilization. Nevertheless, in standard frozen stallion insemination doses production, SP is completely removed and is replaced by a semen extender. In the present study, we analyzed the effects of the selected seminal plasma protein groups that might play an important role in reducing the detrimental effects on spermatozoa during the cryopreservation process. SP proteins were separated according to their ability to bind to hepar...
Ebel F, Vallejos A, Gajardo G, Ulloa O, Clavel E, Rodríguez-Gil JE, Ramírez-Reveco A.The aim of this study was to evaluate seasonal changes in basic parameters of sperm quality and freezability behaviour of ejaculates from 10 fertile heavy draft stallions. A total of 140 ejaculates were collected, processed and evaluated during both the breeding (September-November) and non-breeding seasons (April-June). Fresh semen was evaluated for volume, concentration, total spermatozoa per ejaculate, plasma membrane integrity and total sperm motility. Cryopreserved samples were evaluated for plasma membrane integrity and sperm motility by the CASA system, and for the freezability index (F...
Catalán J, Papas M, Gacem S, Mateo-Otero Y, Rodríguez-Gil JE, Miró J, Yeste M.Previous studies in other mammalian species have shown that stimulation of semen with red-light increases sperm motility, mitochondrial activity, and fertilizing capacity. This study sought to determine whether red-light stimulation using a light emitting diode (LED) at 620-630 nm affects sperm motility and structure of motile subpopulations, sperm viability, mitochondrial activity, intracellular ATP levels, rate of O consumption and DNA integrity of horse spermatozoa. For this purpose, nine ejaculates were collected from nine different adult stallions. Upon collection, semen was diluted in Ke...
Kumar P, Mehta JS, Ravi SK, Dedar RK, Purohit GN, Legha RA, Tripathi BN, Talluri TR.The present study was conducted with the hypothesis that addition of cholesterol to the extender would stabilize the sperm membranes by increasing the cholesterol-to-phospholipid (C:P) ratio and would result in an improved post-thaw semen quality and reduce oxidative stress in the jack (Martina franca) semen. Forty-eight ejaculates from six jacks were collected and analyzed for the present study. The freshly collected semen sample of each jack stallion was divided into five equal fractions after addition of the primary extender without cholesterol-loaded cyclodextrin (CLC) (C) and with 1, 1.5,...
Bahrami A, Divar MR, Azari M, Kafi M.In the present study, we aimed to evaluate the possible protective effects of the nicotinic acid (NA) at three concentrations (10, 20, and 40 mM) on the equine cooled and frozen-thawed spermatozoa quality markers including viability, plasma membrane or acrosome integrity, DNA fragmentation, lipid peroxidation, and total oxidant levels. We also evaluated the effects of NA on preservation of the post-thaw sperm quality after 6 hours of cold storage before freezing. Five stallions were used for semen collections. The current experiment was repeated six times using pooled semen samples from two ...
Ferrer MS, Canisso IF, Ellerbrock RE, Podico G, Lister BN, Hurley DJ, Kline K, Palomares RA.Freezing cooled-transported semen allows veterinarians and breeders to collect and process the semen of stallions on farm, and then ship the semen to a semen freezing center. There, however, is a lack of standardization of shipping and freezing protocols. The objectives were to optimize and simplify protocols to freeze cooled-shipped semen. In Experiment 1, cooled-transported semen was centrifuged at room temperature or 5 °C before freezing. Sperm variables (motility, membrane integrity, acrosome integrity, membrane fluidity) were evaluated before and after freezing. Centrifugation temperat...
Dominguez EM, Moreno-Irusta A, Rodriguez MB, Salamone DF, de Arruda RP, Losinno L, Giojalas LC.The successful use of assisted reproduction techniques (ART) depends in part on the sperm physiological status. Several sperm selection procedures have been applied to improve quality of sperm population when using the ART. There has previously been development of a Sperm Selection Assay (SSA) for humans which is based on the attraction of capacitated sperm by chemotaxis towards progesterone (P), resulting in an enriched sperm population with an optimal physiological status similar to capacitated spermatozoa, with these cells having very little DNA fragmentation and optimal concentrations of r...
Oldenhof H, Wolkers WF, Sieme H.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...
Nery IHAV, Araújo Silva RAJ, Souza HM, Arruda LCP, Monteiro MM, Seal DCM, Silva GR, Silva TMS, Carneiro GF, Batista AM, Câmara DR, Guerra MMP.l-Carnitine (LC) plays a key role in sperm metabolism, easily providing energy through β-oxidation, which positively affects motility. The objective of this study was to investigate the association between blood plasma and seminal plasma LC levels, as well as the effect of LC as an additive in a skimmed milk-based extender during sperm storage at 5°C. In the first experiment, semen and blood samples from 14 Quarter Horse stallions were used. The LC content in blood plasma and seminal plasma was determined by spectrophotometry and their relationships with seminal parameters were evaluated. In...
Andrade Junior LRP, Segabinazzi LGTM, Oliveira SN, Dell'Aqua JA, Papa FO.A high amount of blood and not the mere presence of blood in equine semen impacts fertility. The aim of this study was to develop an approach to rescue the fertility of stallions with high hemospermia levels. Semen from 15 stallions was divided into four experimental groups: (a) Control-pure raw semen, (b) WB50-50% (v/v) whole blood added into semen, (c) E1-WB50 extended in a 1:1 (v/v) ratio with milk-based extender and (d) E2-WB50 extended in a 2:1 ratio with milk-based extender. Sperm kinetics, plasma membrane integrity (PMI), lipid peroxidation (PER) and intracellular superoxide (O ) produc...
Cavalero TMS, Segabinazzi LGTM, Scheeren VFC, Canuto LEF, Gobato MLM, Papa FF.A 7-year-old Quarter Horse stallion was admitted at the hospital with a history of ejaculatory failure for 12 months. The stallion revealed no physical or psychological abnormalities, as well as, normal libido and erection. In addition, there were no abnormalities in accessory sex glands or the aorta artery detected by transrectal ultrasonography. Based on clinical findings, the stallion was diagnosed with an idiopathic ejaculatory dysfunction; therefore, alternative attempts of semen collection were performed. Thermal compress on the basis of the stallion's penis, semen collection on the gro...
Campos GA, Garcia VFC, Freitas-Dell'Aqua CP, Segabinazzi LGTM, Maciel LFS, Alvarenga MA, Papa FO, Dell'Aqua JA.This study aimed to assess the effects of sodium caseinate and cholesterol to extenders used for stallion semen cooling. Two ejaculates from 19 stallions were extended to 50 million/mL in four different extenders and cooled-stored for 24 hours at 5°C. The extender 1 (E1) consisted of a commercially available skim milk-based extender. The extender 2 (E2) consisted of E1 basic formula with the milk component being replaced by sodium caseinate (20 g/L). The extender 3 (E3) consisted of E1 basic formula added to cholesterol (1.5 mg/120 million sperm). The extender 4 (E4) consisted of a combinat...
Treulen F, Aguila L, Arias ME, Jofré I, Felmer R.In vitro manipulation of spermatozoa leads to deleterious changes of structure and function that occur mainly due to oxidative stress, therefore, prevention or treatment is a strategy to improve the functions of processed sperm. In the present study, the aim was to evaluate the effects of MnTBAP supplementation, a compound with antioxidant activity, on in vitro capacitation conditions of thawed equine sperm. For this purpose, stallion spermatozoa (2 × 10 cells/mL) were incubated in the sperm-TLP base medium for 4 h in which there were three different conditions: non-capacitating, capacitating...
Al-Kass Z, Guo Y, Vinnere Pettersson O, Niazi A, Morrell JM.Bacteria colonize stallion semen during collection and processing which may cause disease in inseminated females or negatively affect sperm quality during storage prior to insemination. Antibiotics are added to semen extenders to control the growth of these bacteria but may induce antimicrobial resistance. Research into alternatives to antibiotics for this purpose requires knowledge of which bacteria are present in semen. Not all bacteria in semen, however, can be identified by conventional microbiological techniques. The objectives of the study were to: i) determine which bacteria are present...
Catalán J, Llavanera M, Bonilla-Correal S, Papas M, Gacem S, Rodríguez-Gil JE, Yeste M, Miró J.The aim of this study was to evaluate whether red-light stimulation increases the longevity and resilience of cryopreserved stallion sperm to withstand post-thaw incubation for 120 min. Sixteen frozen straws of 0.5 mL from eight stallions were used. Samples were cryopreserved, thawed through incubation at 38 °C for 30 s and divided into the control and samples exposed to red-light using a triple LED photo-activation system (wavelength: 620-630 nm). Three irradiation protocols consisting of different light-dark-light intervals (1-1-1, 2-2-2 and 3-3-3 min) were tested. Sperm quality param...
Oliveira SN, Andrade LRP, Silva LFMC, Araujo EAB, Rayashi RM, Segabinazzi LGTM, Alvarenga MA, Dell'Aqua CP, Dell'Aqua JA, Papa FO.Treatments for seminal vesiculitis have poor outcomes in stallions; thus, the development of alternative strategies is warranted. This study aimed to evaluate fractionated semen collection as a method to restore the fertility of stallions diagnosed with seminal vesiculitis. Eighteen ejaculates from six stallions (three ejaculates/stallion) diagnosed with seminal vesiculitis were harvested in fractions, as follows: Fraction A (FA), the first two jets; Fraction B (FB), the third and fourth jets; and Fraction C (FC), the fifth and remaining jets of the ejaculate. All fractions were subject to sta...
Dini P, Bartels T, Revah I, Claes AN, Stout TAE, Daels P.A high degree of inbreeding has been reported to negatively impact semen quality in Friesian horses and Shetland ponies. Both breeds are characterized by a closed studbook, small population size, and high incidence of inbreeding. The Dutch Warmblood studbook (KWPN: Koninklijk Warmblood Paardenstamboek Nederland) is a much larger studbook with two distinct populations: the KWPN-Riding horses, managed as an 'open' studbook, and the KWPN-Harness horses, representing a much smaller subpopulation within the KWPN breed and managed as an 'almost closed' studbook. It was recently reported that the deg...
Alamaary MS, Haron AW, Hiew MWH, Ali M.Present study aimed to investigate the effect of adding antioxidants, cysteine and ascorbic acid on the levels of glutamic oxaloacetic transaminase (GOT), glutamic-pyruvate (GPT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and γ-glutamyl transpeptidase (GGT) enzymes of post-thawed stallion sperm. Ten ejaculates were collected each from four healthy stallions and cryopreserved using HF-20 freezing extender containing either 0 mg/ml cysteine or ascorbic acid, 0.5 mg/ml cysteine and 0.5 mg/ml ascorbic acid. All samples in freezing extender containing cysteine or ascorbic acid or ...
Data in briefJune 20, 2020
Volume 31 105887 doi: 10.1016/j.dib.2020.105887
Martín-Cano FE, Gaitskell-Phillips G, Ortiz-Rodríguez JM, Silva A, Gil C, Ortega-Ferrusola C, Peña FJ.This paper provides the dataset of proteins of stallion ejaculates before and after cryopreservation. The data report the analysis and identification of stallion sperm proteins obtained from the same ejaculates and split in two subsamples. The first aliquot consisted on fresh spermatozoa and the second aliquot was frozen and thawed spermatozoa. Samples were analyzed using a UHPLC/MS/MS system consisting of an Agilent 1290 infinity series UHPLC coupled to an Agilent 6550 Q-TOF mass spectrometer (Agilent Technologies, Santa Clara, CA, USA). We provide a data set of 2226 different proteins, with ...
Castro R, Morales P, Parraguez VH.In this work, we investigated the influence of age and seasonality on sperm motility and DNA fragmentation in post-thawing semen from Chilean Purebred Stallions (CPS), a horse breed presenting the oldest genealogy record in South America with an interesting reproductive industry. Despite that semen from aged CPS is frozen all year round, there is a lack of studies characterizing the breed semen freezability in accordance with age and seasonality. Twenty fertile CPS were grouped into the young group, the middle group, and the aged group. Ten ejaculates from each stallion were obtained by using ...
Loureiro KC, Lima-Verde IB, Johannisson A, Ntallaris T, Jager A, Štěpánek P, da Costa Mendonça M, Severino P, Morrell JM.Cryopreservation of stallion spermatozoa tends to cause plasma membrane damage due to the low ratio of cholesterol to phospholipids. Gums have been suggested as an alternative cryoprotectant to glycerol for stallion spermatozoa. Therefore, the present experiment was designed to verify whether the effect of addition of cashew gum (CG), or nanoparticles (NP) containing CG, to the extender before cooling on sperm quality in stallion semen. Ejaculates from 6 stallions were extended and split between six treatment groups (control, a-tocopherol [TOC], CG1, CG0.5, NP1 and NP0.5), stored in cryotubes ...
Guasti PN, Souza FF, Scott C, Papa PM, Camargo LS, Schmith RA, Monteiro GA, Hartwig FP, Papa FO.During ejaculation, a large amount of seminal plasma proteins interact with the sperm membrane, leading to a series of biochemical and structural changes implicated in sperm function and gamete interaction. However, the roles of the majority of these proteins remain unknown. This study aimed to investigate the proteome and functionality of the major equine proteins of seminal plasma and the sperm membrane. Seminal plasma and enriched-membrane proteins (150 μg) were separated by two-dimensional gel electrophoresis, and the respective maps were analyzed. Protein identification was performed by...
Aurich J, Kuhl J, Tichy A, Aurich C.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...
Rubessa M, Feugang JM, Kandel ME, Schreiber S, Hessee J, Salerno F, Meyers S, Chu I, Popescu G, Wheeler MB.The capacity for microscopic evaluation of sperm is useful for assisted reproductive technologies (ART), because this can allow for specific selection of sperm cells for in vitro fertilization (IVF). The objective of this study was to analyze the same sperm samples using two high-resolution methods: spatial light interference microscopy (SLIM) and atomic force microscopy (AFM) to determine if with one method there was more timely and different information obtained than the other. To address this objective, there was evaluation of sperm populations from boars and stallions. To the best of our k...
Quiñones-Pérez C, Martínez A, Crespo F, Vega-Pla JL.Contagious equine metritis is receiving renewed attention due to the continuous detection of carriers in apparent agent-free farms. Interactions of Taylorella with the seminal microflora may be the plausible cause behind these spontaneous changes of the carrier state. Accordingly, the aim of this study was to compare the differences in the seminal microbiome composition of one stallion in the contagious equine metritis carrier state and non-carrier state. Samples were cryopreserved after their extraction. Cell disruption was performed by high-speed homogenization in grinding media. Bacterial f...
Miró J, Marín H, Catalán J, Papas M, Gacem S, Yeste M.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...
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...
Novello G, Podico G, Segabinazzi LGTM, Lima FS, Canisso IF.The objective of this study was to compare semen parameters and embryo recovery rates of cooled stallion semen extended with INRA 96 or BotuSemen Gold. In experiment 1, 45 ejaculates from nine mature stallions were collected, assessed, and equally split between both extenders and then extended to 50 million sperm/mL. Then, the extended semen was stored in three passive cooling containers (Equitainer, Equine Express II, and BotuFlex) for 48 hours. In experiment 2, the same ejaculates extended in experiment 1 were cushion-centrifuged, the supernatant was discarded, and the pellets were resuspend...
Gáspárdy A, Renkó E, Somoskői B, Bába A, Cseh S.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...
Squires EL, Keith SL, Graham JK.Although use of cryopreserved stallion spermatozoa is currently accepted by many breed registries, utilization of this technique remains limited due to poor fertility for some stallions. One reason for these results is osmotic stress that spermatozoa experiences when the cryoprotectant (glycerol) is added to the cells prior to freezing and removal from the cells after thawing. In an effort to minimize osmotic damage, alternative cryoprotectants, having lower molecular weights and greater membrane permeability than glycerol, were evaluated to determine their effectiveness for cryopreserving sta...
Carossino M, Dini P, Kalbfleisch TS, Loynachan AT, Canisso IF, Shuck KM, Timoney PJ, Cook RF, Balasuriya UBR.Equine arteritis virus (EAV) can establish long-term persistent infection in the reproductive tract of stallions and is shed in the semen. Previous studies showed that long-term persistence is associated with a specific allele of the CXCL16 gene (CXCL16S) and that persistent infection is maintained despite the presence of a local inflammatory and humoral and mucosal antibody responses. In this study, we demonstrated that equine seminal exosomes (SEs) are enriched in a small subset of microRNAs (miRNAs). Most importantly, we demonstrated that long-term EAV persistence is associated with the dow...
Zemanová J, Lukác N, Massányi P, Trandzík J, Burócziová M, Nad P, Capcarová M, Stawarz R, Skalická M, Toman R, Koréneková B, Jakabová D.In this study, the concentration of nickel in stallion, bull, ram, boar and fox semen, and its relation with spermatozoa quality was analyzed. The concentration of nickel in semen was 0.20 mg kg(-1) in stallion, 0.12 mg kg(-1) in bull, 0.31 mg kg(-1) in ram, 0.06 mg kg(-1) in boar and 0.36 mg kg(-1) in fox. Seminal nickel concentration was significantly higher (P < 0.05) in foxes than that in bulls and significantly higher (P < 0.01) in rams and foxes in comparison with boars. Evaluation of total pathological spermatozoa revealed the highest number in stallions followed by rams, bulls, boars a...
Macías García B, González Fernández L, Morrell JM, Ortega Ferrusola C, Tapia JA, Rodriguez Martínez H, Peña FJ.The present study attempted to select the subpopulation of stallion spermatozoa that best survived a conventional freezing and thawing procedure, using centrifugation of post-thawed semen samples through a single layer of a glycidoxypropyltrimethoxysilane-coated silica colloid with a species-specific formulation (Androcoll-E). After freezing and thawing, four sperm subpopulations were identified, listed as FT1 to FT4. While subpopulations FT1 and FT2 were characterized by low sperm velocity, high velocities characterized the ones called FT3 and FT4. The single-layer centrifugation (SLC)-handle...
Balao da Silva CM, Ortega Ferrusola C, Morillo Rodriguez A, Gallardo Bolaños JM, Plaza Dávila M, Morrell JM, Rodriguez Martínez H, Tapia JA....At present, the only repeatable means of selecting the sex of offspring is the Beltsville semen sorting technology using flow cytometry (FC). This technology has reached commercial status in the bovine industry and substantial advances have occurred recently in swine and ovine species. In the equine species, however, the technology is not as well developed. To better understand the changes induced in stallion spermatozoa during the sorting procedure, pooled sperm samples were sorted: sperm motility and kinematics were assessed using computer assisted sperm analysis, sperm membrane integrity wa...
Gaitskell-Phillips G, Martín-Cano FE, Ortiz-Rodríguez JM, Silva-Rodríguez A, Gil MC, Ortega-Ferrusola C, Peña FJ.Although cryopreservation is widely used in animal breeding, the technique is still suboptimal. The population of spermatozoa surviving the procedure experiences changes attributed to alteration in their redox regulation. In order to expand our knowledge regarding this particular aspect, the proteome in fresh and frozen thawed aliquots of equine spermatozoa was studied to identify the proteins most severely affected by the procedure. If alteration of redox regulation is a major factor explaining cryodamage, proteins participating in redox regulation should be principally affected. Using a spli...
Minervini F, Lacalandra GM, Filannino A, Garbetta A, Nicassio M, Dell'aquila ME, Visconti A.Fumonisin B1 (FB1) is a mycotoxin produced by Fusarium species that exerts its toxic effect through interference with the sphingolipid pathway by inhibiting ceramide synthase. A FB1-dependent sperm toxicity was reported in boars. No information on FB1-related reproductive toxicity in stallions, the most sensitive animal species, has been reported. The aim of the present study was to assess the in vitro toxicity of FB1 on fresh and frozen-thawed equine spermatozoa by analyzing sperm viability, chromatin stability (SCSA) and reactive oxygen species (ROS) production by flow cytometry and sperm mo...
Pickett BW, Sullivan JJ, Byers WW, Pace MM, Remmenga EE.The effect of centrifugation of diluted and undiluted semen on equine and bovine spermatozoan motility and fertility was examined, as was the effect of seminal plasma and dilution on stallion spermatozoa during incubation before and after freezing. Centrifugation at 370 g or 829 g was not detrimental (P greater than 0.05) to prefreeze or postfreeze motility if a final concentration of 10% seminal plasma was present. A reduction of seminal plasma from 10% to 2% significantly (P smaller than 0.05) reduced motility. A centrifugal force of 956 g significantly reduced prefreeze but not postfreeze m...
Macías García B, González Fernández L, Ortega Ferrusola C, Morillo Rodríguez A, Gallardo Bolaños JM, Rodríguez Martinez H, Tapia JA....Fatty acids and plasmalogens were extracted from the phospholipids of the plasma membrane of stallion spermatozoa, to determine their relation with sperm quality after freezing and thawing. Sperm quality was rated using a quality index that combined the results of the analysis of sperm motility and velocity (CASA analysis), membrane status and mitochondrial membrane potential (flow cytometry) post thaw. Receiving operating system (ROC) curves were used to evaluate the value of specific lipid components of the sperm membrane herein studied as forecast of potential freezeability. From all parame...
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 ...
Catalán J, Yánez-Ortiz I, Tvarijonaviciute A, González-Arostegui LG, Rubio CP, Yeste M, Miró J, Barranco I.This study investigated whether the activities of the antioxidant components of donkey seminal plasma (SP)-both enzymatic (superoxide dismutase (SOD), catalase-like (CAT), glutathione peroxidase-like (GPX), and paraoxonase type 1 (PON1)) and non-enzymatic (measured in terms of total thiol, copper-reducing antioxidant capacity (CUPRAC), ferric-reducing ability of plasma (FRAP), and Trolox equivalent antioxidant capacity (TEAC))-and oxidative stress index (OSI) are related to sperm cryotolerance. For this purpose, 15 ejaculates from jackasses (one per individual) were collected and split into tw...
Al-Essawe EM, Johannisson A, Wulf M, Aurich C, Morrell JM.Addition of seminal plasma (SP) prior to cryopreservation may influence stallion sperm cryosurvival. The objective of this study was to investigate the addition of pooled SP from "good" or "bad" freezer stallions to spermatozoa selected by single layer centrifugation (SLC) prior to cryopreservation on post-thaw sperm quality. Semen from 12 stallions was collected; 5 mL was frozen as control (C) and the remainder was processed by SLC to remove SP and was divided into three aliquots: i) SLC sample without SP (SLC); ii) SLC plus pooled SP from "good freezer" stallions (SLC-GF); iii) SLC plus po...
Delgado-Bermúdez A, Noto F, Bonilla-Correal S, Garcia-Bonavila E, Catalán J, Papas M, Bonet S, Miró J, Yeste M.Aquaporins (AQPs), a family of ubiquitous water channels divided into orthodox AQPs, aquaglyceroporins (GLPs), and superAQPs, are present in stallion spermatozoa. The aim of this study was to elucidate the functional relevance of each group of AQPs during stallion sperm cryopreservation through the use of three different inhibitors: acetazolamide (AC), phloretin (PHL) and propanediol (PDO). Sperm quality and function parameters were evaluated in the presence or absence of each inhibitor in fresh and frozen-thawed samples. In the presence of AC, different parameters were altered ( < 0.05), b...
Martins HS, da Silva GC, Cortes SF, Paes FO, Martins Filho OA, Araujo M, Stahlberg R, Lagares MA.During cryopreservation, sperm was submitted to an increase in reactive oxygen species generation. This work aimed to improve the quality of frozen equine sperm after the addition of antioxidants lactoferrin (Lf) and catalase (Cat) to a freezing extender. Semen from six stallions was frozen with the extenders: F1) control, INRA 82 freezing extender, F2) F1 + 500 μg/ml Lf and F3) F1 + 200 IU/ml Cat. After thawing, sperm motility parameters, membrane functionality and integrity, and acrosome integrity and spontaneous acrosome-reacted sperm were evaluated with a computer-assisted sperm anal...
Kuisma P, Andersson M, Koskinen E, Katila T.The aim of the project was to use current simple and practical laboratory tests and compare results with the foaling rates of mares inseminated with commercially produced frozen semen. In Exp. 1, semen was tested from 27 and in Exp. 2 from 23 stallions; 19 stallions participated in both experiments. The mean number of mares per stallion in both experiments was 37 (min. 7, max. 121). Sperm morphology was assessed and bacterial culture performed once per stallion. In Exp. 1, progressive motility after 0, 1, 2, 3, and 4 h of incubation using light microscopy, motility characteristics measured wit...
Schober D, Aurich C, Nohl H, Gille L.Cryopreservation of spermatozoa is of essential importance for artificial insemination and breeding programs in horses. Besides other factors, spermatozoal motility depends on mitochondrial energy metabolism. Based on changes of single mitochondrial functions it has been suggested that mitochondrial damage during cryopreservation could be a major reason for diminished post thaw semen quality. However, it is still unclear to which extent this influences the whole bioenergetic performance of mitochondria and whether this plays a role during routine cryopreservation procedures. Therefore, it was ...
Restrepo G, Varela E, Duque JE, Gómez JE, Rojas M.Maintaining the integrity of equine sperm subjected to preservation protocols is essential for the successful development of assisted reproduction procedures. The aim of this study was to assess the mitochondrial membrane potential, lipid peroxidation, and DNA integrity of equine sperm subjected to freezing, vitrification, and freeze-drying. Eight ejaculates obtained from four Colombian Creole horses were subjected to programmable freezing, vitrification, and freeze-drying. After thawing or rehydration, sperm motility and kinetics were assessed through a CASA system. The mitochondrial membrane...
Jobim MI, Trein C, Zirkler H, Gregory RM, Sieme H, Mattos RC.The objective was to evaluate protein profiles of equine seminal plasma using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and to determine whether any of these proteins were related to semen freezability. Seminal plasma was collected from 10 stallions, of high and low semen freezability, housed at the State Stud of Lower Saxony, and routinely used in AI programs. Twenty-five protein spots were identified from the two-dimensional gel (12%), seven of which were present in all samples (all proteins were identified by MALDI-MS). Matrix-assisted laser desorption/ionization mass spe...
Peña FJ, Ball BA, Squires EL.Multiparametric assessment of stallion sperm quality using flow cytometry can be a useful adjunct in semen evaluation; however, the availability of flow cytometers in veterinary practice is limited. The ability to preserve and transport sperm samples for later flow cytometric analysis using fixable probes would potentially facilitate this process. In the current study, we validated the combination of live/dead Zombie Green (a fixable dye used to assess live and dead sperm) and MitoTracker Deep Red (used to assess mitochondrial membrane potential). The assay was validated against classic, non-f...
Ortiz-Rodriguez JM, Anel-Lopez L, Martín-Muñoz P, Álvarez M, Gaitskell-Phillips G, Anel L, Rodríguez-Medina P, Peña FJ, Ortega Ferrusola C.Testicular function is particularly susceptible to vascular insult, resulting in a negative impact on sperm production and quality of the ejaculate. A prompt diagnosis of testicular dysfunction enables implementation of appropriate treatment, hence improving fertility forecasts for stallions. The present research aims to: (1) assess if Doppler ultrasonography is a good tool to diagnose stallions with testicular dysfunction; (2) to study the relationship between Doppler parameters of the testicular artery and those of sperm quality assessed by flow cytometry and (3) to establish cut off values ...
Sieme H, Martinsson G, Rauterberg H, Walter K, Aurich C, Petzoldt R, Klug E.The objective of this research was to improve the techniques in processing chilled and frozen-thawed horse semen. In a preliminary experiment (Exp. I), different techniques for sperm selection and preparation [Swim-up, Glass wool (GW) filtration, Glass wool Sephadex (GWS) filtration; Percoll] were tested for their suitability for equine spermatozoa and results were compared with the routine procedure by dilution (Exp. I). In the main experiment (Exp. II), two sperm preparation techniques (GWS, Leucosorb) refering to the results of Exp. I and a previous study of our group (Pferdcheilkunde 1996 ...
Love CC.The sperm chromatin structure assay (SCSA) was introduced by as a method to determine the susceptibility of sperm DNA to denaturation and how those results related to fertility. This initial study used human sperm and was followed by studies in bulls and boars . This assay was one of the first to introduce the technique of flow cytometry, which has the ability to evaluate specific sperm compartments of large numbers of sperm in a short time, as a methodology to evaluate sperm quality and further define the relationship of sperm quality to fertility. For any assay to be of use clinically, it mu...
Janett F, Thun R, Bettschen S, Burger D, Hassig M.The objective of this study was to investigate seasonal changes of semen quality parameters in Franches-Montagnes stallions and to compare the freezability of ejaculates collected in autumn and winter. Experiments were performed using 15 stallions from the National Stud Farm in Avenches (Switzerland). Ejaculates were collected and evaluated every month during 1 year as well as cryopreserved in autumn and winter (September to February). In fresh semen the gel-free volume, concentration, motility and morphology (normal sperm, major defects, vacuoles and acrosome defects) were evaluated and in fr...
Li N, Yu J, Yang F, Shao Y, Wu S, Liu B, Li M, Wang T, Li J, Zeng S.The aim of this study was to evaluate the effects of L-proline on the extender quality of frozen and post-thawed jackass semen. Jackass (n = 6) semen samples were collected and cryopreserved in gradient concentrations (0-80 mM) of L-proline in extenders; post-thawed semen samples were cultured in L-proline medium for 10 hours at 37°C. For cryopreservation experiment I, the motile parameters, mitochondrial membrane potential (MMP), and plasma membrane, acrosome, and chromatin structure integrities of post-thawed semen were assessed. For culture experiment II, additional ROS contents were ana...
Prasinou P, De Amicis I, Fusaro I, Bucci R, Cavallini D, Parrillo S, Caputo M, Gramenzi A, Carluccio A.Fatty acid-based lipidomic analysis has been widely used to evaluate health status in human medicine as well as in the veterinary field. In equine species, there has been a developing interest in fertility and sperm quality. Fatty acids, being the principal components of the membranes, play an active role in the regulation of the metabolic activities, and their role on spermiogenesis seems to be of great importance for the resulting quality of the sperm and, thus, fertility. With the application of widely used lipidomic techniques, the aim of this study was to evaluate: (a) the fatty acid cont...
Gacem S, Catalán J, Yánez-Ortiz I, Soler C, Miró J.The evaluation of the male fertility potential is based on the analysis of the basic spermatic characteristics of concentration, motility and morphology. Thus, the study of sperm morphology is a fundamental element in the seminal analysis, but its real meaning has been biased by the techniques used for its evaluation. These techniques involve dehydration phases and subsequent staining, which involves the production of artifacts. The aim of the study is to compare two methods for equid semen morphology evaluation, Trumorph using living sperm vs. eosin-nigrosine stain. A total of 49 ejaculates f...
Hoffmann N, Oldenhof H, Morandini C, Rohn K, Sieme H.Cryopreserved stallion sperm displays a high degree of male-to-male variability with respect to cell viability after thawing. Animals that have semen with low viability after cryopreservation are classified as 'poor' freezers, and when post-thaw viability is high they are designated as 'good' freezers. Cryoprotective agents that are used for cryopreserving stallion sperm include glycerol, ethylene glycol, methyl formamide, and dimethylformamide, and are typically used in concentrations ranging from 1% to 4%. The aim of this study was to evaluate the osmotic stresses that stallion sperm is expo...
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...
Catalán J, Papas M, Gacem S, Mateo-Otero Y, Rodríguez-Gil JE, Miró J, Yeste M.Previous studies in other mammalian species have shown that stimulation of semen with red-light increases sperm motility, mitochondrial activity, and fertilizing capacity. This study sought to determine whether red-light stimulation using a light emitting diode (LED) at 620-630 nm affects sperm motility and structure of motile subpopulations, sperm viability, mitochondrial activity, intracellular ATP levels, rate of O consumption and DNA integrity of horse spermatozoa. For this purpose, nine ejaculates were collected from nine different adult stallions. Upon collection, semen was diluted in Ke...
