Collection and quality of rhesus monkey semen

Electroejaculation is an accepted method of semen collection from nonhuman primates. Although both penile and rectal probe stimulation techniques have been used, there has been a general lack of consistency and detail regarding their application. This report describes the collection, processing, and evaluation of rhesus monkey semen contrasting two methods of penile electroejaculation: 1) a constant-voltage method where stimulus current is a variable and 2) a constant-current method where stimulus current is operator-controlled. The constant-current method was the more efficient procedure, requiring a lower stimulus current for successful electroejaculation. The influence on semen quality of potentially toxic agents used in the procedure, surgical glove powder and electrolyte cream, was tested; both were detrimental as measured by motility loss. No correlation was found between coagula volume and sperm numbers. The intra- and interanimal variability in semen samples from six monkeys was also evaluated. Penile electroejaculation, combined with control of stimulus current, provides a consistent, successful, and humane method for the collection of semen in the rhesus monkey.     

Methods of semen collection in an ambulatory greater one-horned rhinoceros (rhinoceros unicornis)

Illegal poaching and reduced habitats have led to the endangered status of rhinoceroses. Collection of semen for preservation and future artificial insemination would therefore increase the reproductive potential of the rhinoceros. Although various collection methods have been attemped with individual rhinos, comparison between methods on a single animal have not been reported. This report details the application of six semen collection techniques on an unsedated and unrestrained greater one-horned rhino. The methods included different types of penile and/or rectal stimulation. Artificial vaginas and an inflatible probe for electroejaculation were specially constructed for semen collection. Of the various methods employed, penile massage consistently resulted in sperm-poor seminal fluid, but when preceded by either rectal massage or electroejaculation, seminal fluid with high sperm concentration was obtained.     

Comparison of electroejaculation and transrectal massage for semen collection in range and yearling feedlot beef bulls

Two experiments were conducted to compare electroejaculation (EE) and transrectal massage (RM) of the ampullary region for semen collection from beef bulls, and to determine the effect of semen collection method on semen traits. In experiment 1, semen was collected either by EE or RM randomly assigned on an alternate basis in 137 range beef bulls unaccustomed to being handled. The maximum time allowed for RM was 4 min and if no semen was obtained, EE was used. In experiment 2, semen was collected from 39 yearling feedlot beef bulls that were accustomed to being handled, by RM followed immediately by EE. The maximum time allowed for semen collection by both methods was 4 min. In both experiments, sperm concentration, percent of progressively motile sperm, percent of sperm staining alive, and sperm morphology were determined. In experiment 1, RM resulted in fewer (P<0.001) successful semen collections and fewer bulls with penile protrusion than EE (80.9% versus 100% and 54.4% versus 91.5%, respectively). The success of RM was not influenced by bull age or breed, or by the veterinarian performing the massage. Transrectal massage required more time (30s, P<0.001) for obtaining a semen sample and resulted in samples with lower sperm concentration (P<0.001), percent motile sperm (P<0.05) and percent live sperm (P<0.001) when compared to EE. In experiment 2, EE and RM were equally effective for obtaining a semen sample (97.4 and 94.9%, respectively), but the proportion of bulls exhibiting penile protrusion during semen collection was lower (P<0.0001) with RM compared to EE. Percent of sperm staining alive was also lower (P<0.01) in samples collected by RM. Sperm morphology (normal sperm, head defects, midpiece defects, proximal cytoplasmic droplets, and detached sperm heads) did not differ between samples collected by EE and RM. In conclusion, semen could be collected by transrectal massage from approximately 80% of range beef bulls and from 95% of yearling beef bulls accustomed to handling. Sperm morphology was not affected by the method of semen collection, but percent of motile sperm and live sperm were lower in samples collected by RM. A reduced ability to stimulate penile protrusion with RM precluded examination of the penis in a large proportion of bulls.     

L’AMP chez les blessés médullaires: quelles indications pour quels résultats?

The fertility of men with spinal cord injury (SCI) is severely impaired because of ejaculatory dysfunction and poor semen quality. Only a few patients are able to ejaculate during either sexual intercourse or masturbation. Fortunately, ejaculation can usually be obtained either by penile vibratory stimulation as first treatment option or electroejaculation as the second option. When assisted ejaculation techniques fail because of lack of response or complications such as autonomic dysreflexia, spermatozoa can be retrieved from the vas deferens or epididymis or directly from the testes. Motivated couples with adequate semen quality can be offered penile vibratory stimulation combined with self-insemination at home before resorting to assisted reproductive technology. However, most couples require an assisted reproduction technique. When semen quality is consistently good, up to three or four intrauterine inseminations can be initially recommended. However, this technique achieves only modest pregnancy results andin vitro fertilization techniques are often required. We perform standardin vitro fertilization (IVF) when semen quality is considered to be sufficient, otherwise we perform intracytoplasmic sperm injection (ICSI). With the new techniques now available, the majority of spinal cord injured men stand a fair chance of fathering a child. Availability of ICSI is important to maximize the probability of success for men with very poor semen quality. There are also a number of concerns about the safety of ICSI and the potential risks for the offspring. This new technique must therefore be used very cautiously and requires further surveillance.     

Variations in seminal parameters over a 12-month period in captive bonnet monkeys

Semen samples were collected from adult fertile bonnet monkeys twice a month by penile electroejaculation for twelve consecutive months. Various parameters like semen volume, weight of ejaculate and coagulum, sperm count, sperm motility, sperm morphology, and functional parameters e.g. plasma membrane integrity,in vitro nuclear chromatin decondensation and acrosomal status were evaluated to assess within and between animal variations. Effects of seasonality, if any, on quantity and quality of semen were also studied. Considerable intra- and inter-individual variations in the geometric mean values were observed for semen volume, weights of ejaculate and coagulum, and sperm counts during the study period. On the other hand, sperm motility, morphology, and functional parameters showed less within and between animal variations. Results on motility, morphology, and functional parameters indicated that good semen quality was maintained throughout the year. Various routine and functional parameters did not show any annual variations. The diurnal rhythmicity in circulatory testosterone levels was observed throughout the year. The study shows lack of seasonality in exocrine and endocrine testicular functions and further suggests that motility, morphology, and functional parameters are better indicators of semen quality in captive bonnet monkeys.     

Quality and quantity of smelt (Osmerus eperlanus L.) sperm in relation to time after hormonal stimulation

The effect of Ovaprim (salmon GnRH analogue and a dopamine antagonist) treatment on the quantity and quality of smelt (Osmerus eperlanus L.) sperm was studied in relation to time after hormonal stimulation. Sperm was obtained at 0, 24, 48 and 72 h after treatment (n=13/each time point). Computer Assisted Sperm Analysis (CASA) was used to evaluate sperm motility parameters and histological analysis was used to examine the testis morphology. Only a small volume of semen (1-5 μl) was collected at the beginning of the experiment (time 0) but it dramatically increased 24 h after hormonal treatment. A further increase in semen volume was recorded 48 h after hormonal stimulation. CASA parameters, such as percentage of motile cells, progressive motility, curvilinear velocity, straightlinear velocity, straightness and amplitude of lateral head displacement of stripped sperm increased 48 h after hormonal treatment, which indicates high quality of sperm. No further increase in sperm quality was recorded at 72 h. Compared to stripped semen, testicular semen was characterized by a slightly lower quality. In addition, histological analysis indicated that 24 h after hormonal treatment, a high number of spermatozoa was released from the testis. Ovaprim-stimulated smelt became clearly darker than the control fish. In conclusion, our results suggest that smelt semen should be collected 48 h after hormonal stimulation to ensure high quality of semen. This time may vary depending on maturation status of testis.     

Trehalose enhanced the freezability of Poodle dog sperm collected by an artificial vagina (AV)

In an attempt to develop a suitable freezing method for Poodle dog sperm, an experiment was conducted to investigate semen collection methods of digital stimulation and an artificial vagina (AV), using Tris and trehalose-egg yolk extender, on the characteristics and cryopreservation of sperm. Two dogs (dogs A and B) were subjected to semen collection by digital stimulation and AV. The volume, sperm concentration, sperm motility index (SMI) and acrosome status of ejaculates were determined immediately after collection. The remainder was frozen as pellets in Tris and trehalose-egg yolk extender. Sperm motility index was evaluated after thawing and during a thermal resistance test, and acrosome integrity was also assessed. No significant differences regarding sperm concentrations, SMI and acrosome integrity were observed between semen collected by AV and digital stimulation. However, when dog sperm were collected by an AV and frozen in trehalose-egg yolk extender, the motility index of frozen-thawed sperm was significantly improved compared to sperm frozen in Tris-egg yolk extender which were collected by digital stimulation. In conclusion, semen collected by an AV and frozen in trehalose-egg yolk extender was effective in enhancing the freezability of Poodle dog sperm.     

Stratégies biologiques à l’heure de l’Assistance Médicale à la Procréation chez le blessé médullaire

Most men with spinal cord injury (SCI) men have fertility problems caused by anejaculation and decreased fertility of the ejaculate. There are two main causes for the impaired reproductive potential in SCI men: ejaculatory dysfunction and poor quality semen. However, current treatment techniques allow a large number of SCI males to achieve ejaculation (rectal electro-stimulation, penile vibrator stimulation). Firstly, masturbation and/or penile vibrator are used at home allowing the couple to perform insemination themselves. The semen of men with spinal cord injuries has commonly been characterised by small volume, abnormal count (low or high), decreased sperm mobility, increased formation of reactive oxygen species, sperm autoimmunity, necrospermia. This impairment is thought to be due to insufficient drainage, genitourinary infections and raised scrotal temperature. Testicular biopsy reveals varying degrees of tubule degeneration and decreased spermatogenetic activity. Semen could be used for various assisted reproductive technologies such as intrauterine insemination,in vitro fertilisation (IVF) and microinsemination (ICSI). The literature reports pregnancy rates by intrauterine insemination of about 15 to 20% per couple. Clinical pregnancy rates after IVF and ICSI techniques are 30% per cycle and these results are comparable to the clinical pregnancy rates when these techniques are performed for female infertility. However, semen must be frozen as soon as possible after the injury and the patient must be informed about the various available assisted reproductive technologies.     

Comparative Examination of Capercaillie (Tetrao urogallus L.) Behaviour Responses and Semen Quality to Two Methods of Semen Collection

Artificial insemination (AI) is very helpful in solving the reproductive and biodiversity problems observed in small, closed avian populations. The successful production of fertilized eggs using AI is dependent on the collection of good quality semen. Two methods of male sexual stimulation and semen collection from captive kept capercaillie (Tetrao urogallus L.), one of the most seriously endangered grouse species in Europe, are compared in this study. Ejaculates were obtained either with the use of a dummy female or by the dorso-abdominal massage method. Differences in the individual responses of the males to the two methods of semen collection as well as in their semen quality were noted. Only sperm concentration (432.4 x 10⁶ mL^-1 with dummy female and 614.5 x 10⁶ mL^-1 for massage method) was significantly affected by capercaillie stimulation method. Sperm motility and morphology were not affected (P≥0.05). Thus, for semen collection from captive kept capercaillie both methods can be used successfully. The dummy female can be an alternative to dorso-abdominal massage method, commonly used for semen collection from domesticated bird species.     

Rhesus monkey sperm penetration into zona-free hamster ova: Comparison of preparation and culture conditions

A major problem in development of nonhuman primate in vitro fertilization is the selection of donor males and repeated collection of consistent sperm samples. In practice, collection of a viable semen sample is highly dependent on operator technique and the type of animal restraint. We report an updated method for semen collection from the rhesus monkey (Macaca mulatta), use of TES-Tris (TEST) Yolk Buffer (TYB) for prolonged sperm storage and improved results of hamster ovum penetration assay. Semen was obtained from adult males restrained with 2.0 mg/kg IM ketamine hydrochloride prior to direct penile stimulation (Grass SD-9, frequency 150, delay 9, duration 7, volts 12–18, repeat mode, twin pulse). Liquified semen was washed and centrifuged twice at 100 × g for 5 min in BWW, Ham's F-10 and TALP and allowed to swim-up 60 min at 37° in 5% CO₂ and air. Alternatively, semen was mixed 1:1 with TYB, refrigerated 20 h at 4°C, centrifuged at 100 × g for 5 min, and the pellet resuspended in 1.0 ml of TALP or BWW prior to use. Hamster ova penetration was achieved with capacitated macaque sperm. Penetration was significantly improved (P < .001) with preincubation in TYB followed by resuspension in TALP (79%).     

Semen quality of postpubertal boars during increasing and decreasing natural photoperiods

The present study analyses the effects of increasing and decreasing photoperiods on the semen quality of 20 selected postpubertal Landrace boars. The boars were exposed, throughout 75 days, to increasing and decreasing photoperiods of natural light, a constant temperature of 21 +/- 1 degrees C and 60-70% of humidity, fed with a nutritious diet and, submitted to a rhythm of semen collection of twice a week. During the last 2 weeks of each treatment, semen samples were analysed and the parameters measured were: ejaculate volume and pH, sperm concentration, sperm production and the number of semen doses per ejaculate, sperm vitality, sperm motility, osmotic resistance of spermatozoa and sperm morphology. The comparative analysis between increasing and decreasing photoperiods indicated that the semen quality of boars exposed to a decreasing photoperiod was reduced as a consequence of decreases in sperm concentration, sperm production and the number of semen doses. There was no difference between increasing and decreasing photoperiods in terms of sperm vitality and sperm motility, nor in the osmotic resistance of spermatozoa to isotonic and hypotonic media. The analysis of sperm morphology showed significantly lower frequencies of mature and immature spermatozoa with a distal cytoplasmic droplet, and significantly higher frequencies of immature spermatozoa with a proximal droplet in boars exposed to the decreasing photoperiod. These results indicate that the sperm quality of the selected boars decreased during decreasing photoperiods, in comparison with increasing photoperiods, mainly due to impaired testicular function.     

Influence of Griseofulvin treatment on semen quality in the dog

Griseofulvin is used to treat dermatomycosis in many species and causes oligospermia in supra-pharmacological doses. The aim of the present study was to evaluate the effect of Griseofulvin administered at therapeutic doses upon semen quality in dogs. Four dogs were treated with Griseofulvin (25 mg/kg per day) for 30 days. Semen collections and analyses were performed before, during and for 100 days after treatment for the Griseofulvin group and 10 untreated control dogs. Semen analyses included mean percentage of forward progressively motile sperm, total sperm output, normal live sperm and normal dead sperm. There was no significant difference between control and treated dogs for each of the semen quality parameters. Therapeutic dosage of Griseofulvin had no deleterious effect upon semen quality in dogs, although this does not preclude potential embryotoxic and teratogenic effects.     

Edible bird’s nest supplementation in chilled and cryopreserved Arabian stallion semen

Diluents and various biological products have been used in different animal species, with promising outcomes in post-thaw sperm quality. Nevertheless, only a few reports are available for the semen of Arabian horses. Edible bird’s nest (EBN) – a product of the salivary secretions of swiftlet species is widely known to have both antioxidant and anti-inflammatory properties. Presently, there is no data available on the role of EBN supplemented in different extenders and its effect on semen quality in stallion semen. Two in vitro experiments were conducted to examine the effects of edible bird’s nest (EBN) on the quality of chilled and post-thawed cryopreserved Arabian stallion spermatozoa. In experiment one, 10 ejaculates were collected, divided into two equal parts, diluted using EquiPlus® and INRA 96® and supplemented with 0 % (control), 0.12 %, 0.24 % EBN concentrations. The semen samples were stored at 5 ℃ and observed at 0, 24, and 48 h. Sperm kinetics variables (% total motility [TM] and progressive motility [PM], curvilinear velocity; VCL, straightness; VSL, average path velocity; VAP) were analyzed using computerized assisted sperm analysis. For chilled semen, there was no significant difference in any of the sperm quality parameters between control (0 %), 0.12 %, and 0.24 % EBN supplementation either in INRA96® or EquiPlus®. In experiment two, nine ejaculates were diluted and cryopreserved using EquiPlus Freeze® and INRA Freeze® containing 0 %, 2.4 %, and 4.8 % EBN, and evaluated after thawing. Sperm kinetics, DNA integrity and antioxidant capacity - Biological Anti-oxidant Potential (BAP) and Reactive Oxygen Metabolites (d-ROMs) test were evaluated. In chilled semen, there was no significant difference in any of the sperm quality parameters between control (0 %), 0.12 %, and 0.24 % EBN supplementation either in INRA96® or EquiPlus®. For frozen semen supplemented with 2.4 % and 4.8 % EBN had higher sperm motility parameters compared to control in INRA Freeze® and EquiPlus Freeze®, but the values were not statistically significant (P > 0.05). Also, EBN supplementation had no significant effects on the DNA integrity, biological antioxidant potential, and reactive oxygen metabolites. EBN supplementation had no significant effects on sperm quality and antioxidant status in chilled and frozen Arabian Stallion semen. Future studies might consider different methods of EBN preparation and concentrations to elucidate the potential biological impact of EBN in Arabian stallion semen.     

Single bovine sperm sex typing by amelogenin nested PCR

Sex-sorted bovine semen has become a valuable tool in animal production for sex preselection. Development of novel sperm sexing technologies, or evaluation of the quality of existing methods, often requires a single-sperm, sex-typing method that is reliable and easy to perform. In the present study, we report the development, validation, and application of a simple, reliable, and cost-effective method for single-sperm sex typing using nested polymerase chain reaction (PCR), based on the amelogenin gene. Several hundred single sperm were isolated using a simple manual technique, or a high-speed flow-sorter, and were successfully sex-typed using the amelogenin nested PCR. Based on the pooled results of individual sperm, there was no significant difference in the semen sex ratio of unsorted (44.6% X-sperm and 55.4% Y-sperm) or X/Y-sorted semen (91.4% X-sperm and 94.0% Y-sperm), as compared to the expected ratio in unsorted semen or the post-sorting reanalysis data, respectively. The amelogenin single-sperm sexing method was an adaptable, accurate, and reliable tool for single-sperm sex typing.     

Influence of thawing method on motility, plasma membrane integrity and morphology of frozen-thawed stallion spermatozoa

Different thawing methods are used for stallion semen, however, it is unclear which method is the optimal one. To determine if the thawing temperature has an effect on semen quality, we compared 2 thawing temperatures, 75 degrees C and 37 degrees C. The following parameters were used to measure sperm quality: sperm motility, sperm viability, plasma membrane integrity and sperm morphology. Twenty-three ejaculates from 10 Dutch Warmblood stallions were thawed either at 37 degrees C for 30 sec or at 75 degrees C for 7 sec. Sperm motility was evaluated by a Hamilton Thorn Motility Analyser. Plasma membrane integrity and sperm viability were evaluated by using a live/dead fluorescein stain containing a calcein AM probe and ethidium homodimer-1 probe. The eosinaniline blue staining method was used to evaluate the percentage of live and dead cells, as well as sperm morphology. There was no significant difference (P = 0.84) between sperm motility after thawing at 37 degrees C and 75 degrees C. There was also no significant difference (P = 0.053) between the percentage of live spermatozoa using the calcein AM/ethidium homodimer stain after thawing at 37 degrees C and 75 degrees C. There was, however, a significant difference (P = 0.032) between the percentage of live spermatozoa using the eosin-aniline blue stain after thawing at 37 degrees C compared with that at 75 degrees C. In conclusion, our laboratory results indicated that stud farms using frozen semen should thaw the straws at 37 degrees C instead of 75 degrees C. The lower temperature is easier to work with, as thawing at the higher temperature requires special equipment and has to be timed very carefully to avoid damage to the spermatozoa.     

Association of heat shock protein 70 with semen quality in boars

This study attempted to clarify the relationship between the levels of 70kDa heat shock protein (HSP70) and semen quality in boars. Semen samples from 29 (13 Duroc, 9 Landrace, and 7 Yorkshire) boars (mean age=25.2+/-2.2 months) were examined. Three to four ejaculates per boar, collected during cool and hot seasons, were evaluated in terms of the sperm concentration, sperm motility, percentage of normal and abnormal sperm, as well as percentage of sperm with proximal and distal plasma droplets. Significant seasonal and breed differences in semen quality were observed. Experimental results indicate that the semen quality of Landrace boars was better than those of Yorkshire and Duroc boars (P<0.05) and semen quality declined significantly during the hot season (P<0.05). One-dimensional SDS-PAGE analysis of spermatozoa proteins indicated that protein profiles did not significantly differ between seasons and among breeds. Both constitutive and stress-inducible form of HSP70 were detected in boar spermatozoa by Western blot analysis. The level of HSP70, which revealed no difference among breeds within a season, was significantly lower during the hot season in all the three breeds (P<0.05). Although there appeared to be low correlation coefficients between the level of HSP70 and semen quality traits, the semen quality tended to decline significantly in samples with a lower level of HSP70. Results in this study suggest that the levels of HSP70 in boar spermatozoa are significantly lower during the hot season and might be associated with semen quality.     

Effectiveness of two systems for transporting equine semen

The storage and transport of cooled, liquid semen is an effective way of facilitating the use of desirable stallions for breeding mares located on distant farms. The Equitainer System is the most widely used transport container and it has been shown that it is possible to ship semen in this container and obtain good conception rates. However, the cost of Equitainers is high, and stud-farms that ship large quantities of semen have tended to rely on cheaper alternatives, even though little documentation exists concerning their reliability, especially under extreme temperature conditions. Two different containers for transporting equine semen (the Equitainer and a styrofoam box) were compared in their effectiveness at maintaining semen quality (i.e. sperm motility and plasma membrane integrity) during 24 h of storage. The transport containers were stored at 2 different environmental temperatures, i.e., room temperature (20 degrees C) and 37 degrees C. Thirty-seven ejaculates from 10 Standardbred stallions (3 to 6 samples per stallion) were examined. Sperm function and plasma membrane integrity were assessed using a Mika Motion Analyzer and a fluorescein stain (Calcein AM/Ethidium homodimer) in fresh diluted semen that had been stored for 24 h at room temperature (20 degrees C). Another 18 ejaculates from 5 stallions were examined using methods described above, but the transport boxes were kept at a high environmental temperature (37 degrees C). After storage at room temperature, there was no significant difference in total sperm motility and frequency of spermatozoa with an intact plasma membrane between the 2 types of transport boxes. A significant difference was seen in linear sperm motility, with the Equitainer being the better container. However, a significant difference was also seen in average path velocity, with the styrofoam box being the better container. After storage at 37 degrees C, the Equitaner maintained semen quality better. A significant difference was seen in total sperm motility, average path velocity, lateral head displacement and frequency of spermatozoa with an intact plasma membrane between the 2 types of transport boxes. Although, both transport containers were satisfactory when used under normal conditions. The Equitainer seemed superior under more extreme temperatures and during longer transport periods (> 24 to 30 h).     

Éjaculation provoquée par le vibromassage ou l’électrostimulation endorectale chez le blessé médullaire: injection intracytoplasmique de spermatozoïdes obtenus par stimulation endorectale chez neuf couples

800 to 1,000 cases of traumatic spinal cord injury (S.C.I.) are observed in France each year. At present time, there are more than 35,000 survivors of S.C.I. in France. Studies show that 80% of injuries occur in men and 82% occur in individuals between the ages of 16 and 45 years. It is well known that more than 80% of men with S.C.I. suffer from ejaculation dysfunction. Since most of these couples are young, many desire a family and seek help to remedy their infertility. Two methods are mainly used to retrieve sperm from S.C.I. men: penile vibratory stimulation and rectal electrostimulation. Penile vibratory stimulation to induce ejaculation is recommended as first-line treatment, and S.C.I. men should only be referred for electroejaculation after failure of this technique. More than 70% of patients respond to penile vibratory stimulation with anterograde ejaculation. Patients who fail to ejaculate by penile vibratory stimulation are treated by electroejaculation. Their ejaculates often have normal sperm counts, but with a higher proportion of immotile sperm than in men without S.C.I. The authors report the pregnancy outcome of a series of 9 couples undergoing combined electroejaculation and in vitro fertilization with intracytoplasmic sperm injection: 25% pregnancies per cycle and 33% pregnancies per couple.     

Fertilization rates and in vitro embryo production using sexed or non-sexed semen selected with a silane-coated silica colloid or Percoll

The aim of this study was to evaluate sperm fertilization rates and in vitro embryo development rates for sexed and non-sexed semen selected using a silane-coated silica colloid method (Isolate) or Percoll. Frozen/thawed, sexed and unsexed semen samples from four Holstein bulls were randomly allocated to one of two different density gradient selection methods. Sperm quality (motility, concentration, morphology and membrane integrity) were evaluated and compared before and after sperm selection. Sperm motility and morphology improved (P < 0.005) after the sperm selection process with no differences between the two methods. For non-sexed semen, Percoll gradient increased the mean (± SEM) percentage of sperm recovered (57.3 ± 2.8) compared to Isolate (46.0 ± 1.8; P < 0.01). However, membrane integrity was higher after Isolate than Percoll (sexed semen: 41.0 ± 0.6 vs. 38.8 ± 0.8 and non-sexed semen 60.8 ± 1.6 vs. 58.8 ± 0.5; P < 0.05). The percentage of blastocysts produced was higher when either sexed or non-sexed semen was selected by Isolate (14.0 ± 1.0; 22.0 ± 1.1) than by Percoll (10.5 ± 1.5; 17.0 ± 2.1, respectively; P < 0.05). In summary, Isolate was a more effective method for the recovery of high quality sperm for in vitro fertilization embryo production.     

Edible bird’s nest supplementation in chilled and cryopreserved Arabian stallion semen

Diluents and various biological products have been used in different animal species, with promising outcomes in post-thaw sperm quality. Nevertheless, only a few reports are available for the semen of Arabian horses. Edible bird’s nest (EBN) – a product of the salivary secretions of swiftlet species is widely known to have both antioxidant and anti-inflammatory properties. Presently, there is no data available on the role of EBN supplemented in different extenders and its effect on semen quality in stallion semen. Two in vitro experiments were conducted to examine the effects of edible bird’s nest (EBN) on the quality of chilled and post-thawed cryopreserved Arabian stallion spermatozoa. In experiment one, 10 ejaculates were collected, divided into two equal parts, diluted using EquiPlus® and INRA 96® and supplemented with 0 % (control), 0.12 %, 0.24 % EBN concentrations. The semen samples were stored at 5 ℃ and observed at 0, 24, and 48 h. Sperm kinetics variables (% total motility [TM] and progressive motility [PM], curvilinear velocity; VCL, straightness; VSL, average path velocity; VAP) were analyzed using computerized assisted sperm analysis. For chilled semen, there was no significant difference in any of the sperm quality parameters between control (0 %), 0.12 %, and 0.24 % EBN supplementation either in INRA96® or EquiPlus®. In experiment two, nine ejaculates were diluted and cryopreserved using EquiPlus Freeze® and INRA Freeze® containing 0 %, 2.4 %, and 4.8 % EBN, and evaluated after thawing. Sperm kinetics, DNA integrity and antioxidant capacity - Biological Anti-oxidant Potential (BAP) and Reactive Oxygen Metabolites (d-ROMs) test were evaluated. In chilled semen, there was no significant difference in any of the sperm quality parameters between control (0 %), 0.12 %, and 0.24 % EBN supplementation either in INRA96® or EquiPlus®. For frozen semen supplemented with 2.4 % and 4.8 % EBN had higher sperm motility parameters compared to control in INRA Freeze® and EquiPlus Freeze®, but the values were not statistically significant (P > 0.05). Also, EBN supplementation had no significant effects on the DNA integrity, biological antioxidant potential, and reactive oxygen metabolites. EBN supplementation had no significant effects on sperm quality and antioxidant status in chilled and frozen Arabian Stallion semen. Future studies might consider different methods of EBN preparation and concentrations to elucidate the potential biological impact of EBN in Arabian stallion semen.