Sperm Motility Parameters Research Articles

Effect of sodium molybdate on cadmium-related testicular damage in adult male Wistar rats

BackgroundMolybdenum, as a trace element, has various pharmacological effects, including antioxidant, antiviral, anti-allergic, anti-osteoporosis, anti-tumor, anti-inflammatory, anti-diabetic, anti-obesity, and free radical-scavenging activities. This study aimed at investigating the sodium molybdate impacts on cadmium chloride (CdCl2)-induced testicular toxicity in adult Wistar rats. MethodsThe impacts of oral administration of sodium molybdate (0.05, 0.1, 0.2, and 0.4 mg/kg) was evaluated in healthy and infertile animals. Animals were randomly assigned to nine groups, including healthy control, sodium molybdate alone, infertile control (3 mg/kg of CdCl2), and sodium molybdate plus CdCl2. Following 30 days of administration, animals were sacrificed for biochemical and histopathological assays. ResultsThe results indicated that administration of sodium molybdate to infertile rats significantly mitigated the cadmium impacts on sperm appearance, concentration, and motility parameters. Also, sodium molybdate reduced the production of malondialdehyde (MDA) and enhanced antioxidant enzymes activities in the testicular homogenates in rats; these findings were supported by histopathological examinations. Treatment with sodium molybdate significantly increased aquaporin-9 (AQP9) expression in the testicular tissues of infertile rats. ConclusionsThe current findings suggested that sodium molybdate performs as a strong protective agent from CdCl2-related testicular toxicity in rats.

Vanadium adversely affects sperm motility and capacitation status via protein kinase A activity and tyrosine phosphorylation

Vanadium is a chemical element that enters the atmosphere via anthropogenic pollution. Exposure to vanadium affects cancer development and can result in toxic effects. Multiple studies have focused on vanadium’s detrimental effect on male reproduction using conventional sperm analysis techniques. This study focused on vanadium’s effect on spermatozoa following capacitation at the molecular level, in order to provide a more detailed assessment of vanadium’s reproductive toxicity. We observed a decrease in germ cell density and a structural collapse of the testicular organ in seminiferous tubules during vanadium treatment. In addition, various sperm motion parameters were significantly decreased regardless of capacitation status, including sperm motility, rapid sperm motility, and progressive sperm motility. Curvilinear velocity, straight-line velocity, average path velocity, beat cross frequency, and mean amplitude of head lateral displacement were also decreased after capacitation. Capacitation status was altered after capacitation. Vanadium dramatically enhanced protein kinase A (PKA) activity and tyrosine phosphorylation. Taken together, our results suggest that vanadium is detrimental to male fertility by negatively influencing sperm motility, motion kinematics, and capacitation status via abnormal PKA activity and tyrosine phosphorylation before and after capacitation.

Assessment of preclinical effect of (+)-catechin hydrate on sexual function: An in silico and in vivo study.

Considering dopamine-enhancing effect of (+)-catechin, the present study was designed to evaluate dopamine-2 (D2) receptor agonistic and phosphodiesterase-5 (PDE5) enzyme inhibitory effects in in silico and effect on male sexual function of Sprague Dawley rats in vivo. (+)-Catechin and standard (sildenafil and bromocriptine) were docked using Autodock Vina 1.1.2 and visualised by UCSF Chimera 1.14. Significant interactions in terms of binding energies were observed for catechin with both proteins. In in vivo study, the rats were dosed orally for 54days with (+)-catechin hydrate (50mg/kg), sildenafil citrate (standard, 4mg/kg) and carboxymethylcellulose (vehicle, 0.25%w/v). The aphrodisiac effects were evaluated on the day 14, 28, 42 and 54 using the behavioural parameters of mounting and intromission. After the study, animals were sacrificed and testes and spermatozoa were assessed for safety profile. Results showed a significant increase in mount and intromission frequencies and a significant reduction in mount and intromission latencies in the catechin group on all tested days when compared to vehicle control. (+)-Catechin was found to be safe on histology of testes, sperm count, sperm motility and sperm morphology parameters. In conclusion, catechin demonstrated an enhancement in sexual behaviour without eliciting toxicity on the male reproductive system in rats.

Use of Glass-Forming Water–Salt Systems for Preparing Human Sperm to Hypothermic Semen Preservation

A protectant for hypothermic preservation of human sperm based on glass-forming water–salt systems of vital metals (magnesium, zinc, and calcium) was developed for the first time. The method of sperm preservation without preliminary washing of seminal plasma is proposed, which retains the sperm viability and sperm motility parameters comparable to the values of native sperm. The proposed protectant and method of preparing human sperm for hypothermic storage at the first stage help to slow down the metabolism as evidenced by the complete loss of sperm motility and maintenance of the nearly 100% viability of the studied sperm. Once the protectant was removed, sperm motility is restored up to 90–100% compared to the indices of native sperm.

The Effect of Resorcinol on Bovine Spermatozoa Parameters in Vitro.

The goal of this study was to observe the effect of resorcinol on motility, viability and morphology of bovine spermatozoa. The semen was used from six randomly chosen breeding bulls. Ejaculate was diluted by different solutions of resorcinol in 1:40 ratio. Samples were divided into 7 groups with different concentrations of resorcinol (Control, RES1 - 4 mg/ml, RES2 - 2 mg/ml, RES3 - 1 mg/ml, RES4 - 0.5 mg/ml, RES5 - 0.25 mg/ml and RES6 - 0.125 mg/ml). Motility of spermatozoa was detected using CASA method at temperature of 37 °C in time periods 0, 1, 2, 3, 4 hours from the start of the experiment. Significant motility differences between all groups except control and RES6 with difference of 5.58 %, as well as between RES1 and RES2 groups with difference of 2.17 % were found. Progressive motility had the same significant differences. Spermatozoa viability (MTT test) decreased compared to control in all experimental groups during the entire duration of experiment. Observing morphologically changed spermatozoa, no significant changes were observed and a higher percentage of spermatozoa with separated flagellum in all experimental resorcinol groups compared to control were detected. Also, increased number of spermatozoa with broken flagellum, acrosomal changes and other morphological forms in the group with the highest concentration of resorcinol (RES1) were found. Results of our study clearly show negative effects on motility parameters of spermatozoa which depend on concentration, cultivation temperature and time period.

Effect of relaxin on cryopreserved beef bull semen characteristics

This study aimed to improve the quality of cryopreserved beef bull (Piedmontese) semen by incorporation of relaxin in diluted semen before cryopreservation procedures. Semen samples were collected from 4 proven fertile bulls, using artificial vagina, once per week for 8 consecutive weeks and pooled together then diluted with Bullxcell® extender, and supplemented with different concentrations of relaxin (0 (control), 25, 50 and 100 ng/ml) before cooling, equilibration and freezing procedures. Frozen semen was thawed and assessed for motility by Computer-Assisted Sperm Analysis and vitality parameters such as acrosome, plasma membrane and DNA integrities, apoptosis, mitochondrial membrane potential, mucus penetration and SOD activity. The developmental potential of bovine embryos produced in vitro by using relaxin-treated was also investigated. In the present study, 50 and 100 ng/ml relaxin incorporation in extended bull semen before cryopreservation induced a reduction of sperm motility immediately after thawing (0h), whereas, during long incubation periods (1–2 h), relaxin showed a significant positive effect on sperm quality by improving the sperm motility and velocity parameters. Interestingly, sperm vitality was improved by 25 and 100 ng/ml relaxin and the blastocyst developmental rate was significantly increased in the 25 ng/ml relaxin group compared with controls (52/118, 44.0% vs. 32/116, 27.6%, respectively). These findings suggest a potential use of relaxin at the doses tested in the present study as an additive in the cryopreservation media of bull semen to improve sperm quality.

In-vitro Antimicrobial Effect of Silybum marianum Extract on Bacterial Isolates from the Semen of Infertile Men in Iran

Background: Genital tract infection is one of the main causes of male infertility. The increasing frequency of drug resistance and side effects of antibiotics have urged researchers to seek alternative sources of antimicrobial agents, such as medicinal herbs, for the treatment of antibiotic-resistant infections. Objectives: The present study aimed to determine the frequency of potentially pathogenic microorganisms in the semen of infertile men and investigate the antimicrobial activity of Silybum marianum extract against the isolates. Methods: Semen samples were collected from 96 infertile men who referred to the Pasteur Laboratory in Gorgan, Iran. Semen samples were first analyzed according to the World Health Organization guidelines and then underwent microbiological tests to identify pathogens. Antimicrobial susceptibility to S. marianum extract was evaluated using the disk diffusion (Kirby-Bauer) method. Moreover, the active components of the extract were identified by the gas chromatography-mass spectrometry technique. Results: In the semen analysis, 64% of the samples had problems in the parameters of sperm count, motility, and morphology. In addition, the bacterial contamination was observed in 36% of semen samples. The most and the least common isolates were Staphylococcus aureus (45%) and Klebsiella pneumoniae (11%) and Staphylococcus epidermidis (11%), respectively. In Gram-positive isolates, S. aureus was mostly resistant to azithromycin (57%). In Gram-negative isolates, Escherichia coli was mostly resistant to gentamicin (37%). The inhibitory activity of S. marianum flower extract was significantly higher than that of S. marianum leaf extract (P < 0.01). Silybin (2.64%) and silychristin (3.07%) were the most abundant constituents of S. marianum flower extract. Conclusions: Bacterial infections play an important role in male infertility and S. marianum extract after purification can be potentially used for the treatment of antibiotic-resistant genital tract infections.

Gamete management for artificial reproduction of northern pike Esox lucius (Linnaeus, 1758)

Eyeing and hatching rates with regard to sperm: egg ratio, the length of time eggs remain viable post-activation in water, and duration of storage time in testicular and stripped sperm were determined in northern pike Esox lucius. In a trial of seven sperm: egg ratios, we found 4 × 105 to be the minimum tested number of spermatozoa per egg to produce an acceptable hatching rate (66 ± 4%). To determine the period of time that eggs remain viable after activation, eggs were placed in hatchery water and fertilized at 0, 15, 30, 45, 60, 90, 120, 240, and 480 s post-activation. With elapsed time, the eyeing and hatching rates decreased significantly (P < .05), dropping from 83 ± 8% and 73 ± 1% at time 0 to 69 ± 7% and 62 ± 1%, respectively, after 30 s. At 480 s post-activation, the eyeing and hatching values were 23 ± 8% and 15 ± 4%. Produced eyeing and hatching rates and sperm motility parameters of testicular and stripped sperm were compared over the course of 48 h storage. Testicular sperm remained viable for 24 h compared with 3 h for stripped sperm. The obtained data clarified important biological characteristics and interaction of gametes for use in developing optimal techniques for artificial reproduction of northern pike under controlled conditions.

Beneficial influence of fetal bovine serum on in vitro cryosurvival of chicken spermatozoa

Chicken spermatozoa are highly susceptible to cryopreservation often requiring extenders containing additives to enhance their post-thaw quality. Although protective properties of fetal bovine serum (FBS) during freezing of tissue cultured cells are widely known, its potential as a cryoprotectant for sperm cells has not been largely explored. Thus, the aims of our study were to (i) investigate the protective effect of FBS at different concentrations (0, 5, 10, 15 and 20%) against cryodamages in chicken spermatozoa, and (ii) test the FBS concentration that yielded the best preservation versus 1 mg/mL of cholesterol-loaded cyclodextrins (CLCs). Samples were assessed before and after freezing for sperm motility parameters, plasma membrane and acrosomal integrities, mitochondrial membrane potential, oxidative stress and plasma membrane peroxidation. Our findings showed that, despite their beneficial effects on fresh spermatozoa, higher FBS concentrations (15 and 20%) obtained the worst results for most motility and functional parameters after thawing. In contrast, lower FBS concentrations (5 and 10%) improved all post-thaw variables when compared to control. Afterwards, based on regression analysis, the concentration of 7% FBS was chosen to be assessed against CLCs in an experiment composed by four groups: control, FBS, CLCs, and FBS + CLCs. FBS and FBS + CLCs groups exhibited higher progressive motility in fresh samples, whereas only FBS maintained higher post-thaw progressive motility. Additionally, the incorporation FBS into extenders increased the percentage of rapid cells and reduced free radicals production and plasma membrane peroxidation. Together, these outcomes indicated that FBS minimize some harmful effects of cryopreservation, providing an alternative for chicken semen extenders that in many aspects appears to be superior to CLCs at 1 mg/mL.

Gökkuşağı Alabalığının (Oncorhynchus mykiss) Sperm Kalitesinde Bilgisayarlı Otomatik Sperm Analiz Sisteminde (CASA) Leja ve Makler Lamlarının Performansının Karşılaştırılması

In this study, the effect of the chamber used for the automated analysis of sperm motility and sperm kinematics parameters by CASA was evaluated of rainbow trout (Oncorhynchus mykiss) sperms. The assessment of motility parameters was carried out using CEROS II (Hamilton-Thorne, Beverly, MA, USA) connected to CX41 microscope (Olympus, Japan) at room temperature. Sperm samples were collected from five adult males by abdominal massage during the reproduction season and analyzed with two different chambers as follows specialty: Leja 2 cell chambered with 20 µl deep (Leja Products, Netherlands) and Makler chamber, round shape with 10 µl deep (Sefi-Medical Instrument, Haifa, Israel). Total sperm motility (Mot, %), and Velocity of Curvilinear (µm/s) were measured. For fertilization test, eggs from one female (550 g approximately 7500 eggs) were separated in equal five parts. Each part of eggs (approximately 1.500 eggs) was fertilized with each analyzed sperm (5ml). Fertilization, incubation procedure and calculation of fertilization rates have been kept as used routinely for rainbow trout culture procedure. The fertilization rates were found ˃80% for all used males. The motility percentage of samples analyzed by Leja has been found higher 90% while by makler changed between 26-45%. There is significantly effect on different chambers used in this study to determining the motility percentage. The high stability results and matched the fertilization success were detected in Leja 2-chamber. Statistical study with motility percentage showed a significant difference between Leja and Makler chambers (p<0,05).

DNA damage, oxidative stress, decreased viability and motility in common carp (Cyprinuscarpio L.) spermatozoa induced by tryptophan, phenylalanine and cysteine amino acids during short-term storage

Free amino acids are found in spermatozoa and seminal plasma. Their qualitative and quantitative presence have been affected by many factors. Under in vitro storage conditions, their effects on spermatozoa in animal species and also human have been concerned over the last decades. In this study, the effects of tryptophan, phenylalanine and cysteine on DNA integrity, lipid peroxidation, viability and motility parameters in spermatozoa of the common carp (Cyprinuscarpio) were assessed during short-term storage. Spermatozoa were incubated at 1, 5, 25, and 50 mM of these amino acids in vitro at 4 °C in a time?dependent manner (6, 24, and 48 h). The supplementation of cysteine, phenylalanine and >5 mM of tryptophan to the incubation medium attenuated the spermatozoa motility parameters and viability. Particularly, >1 mM of cysteine completely inhibited progressive motility and decreased viability to

Altered testicular cell type composition in males of two outbred mouse lines selected for high fertility.

Recently we described two outbred mouse lines which have been selected for high fertility. These mouse models doubled the number of offspring per litter. Although selected for a primarily female-trait of high fertility (increased litter size), we were interested whether also males of the fertility lines show differences within their reproductive organs. We investigated males from two outbred mouse lines which have been selected for the phenotype "high fertility" for more than 170 generations. In the present study, we analysed the testicular cell type composition by flow cytometry. We further investigated the weights of reproductive organs, histomorphometry of testis as well as studied sperm motility parameters using a thermal stress assay as well as a sperm hyperactivation assay. Here, we describe that males of the fertility line (FL) 1 show an increased percentage of diploid cells within the testis. Flow cytometric analysis identified this enlarged cell population as Leydig cells. Testis weights were unaffected whereas the weights of seminal vesicles of FL1 and FL2 were increased compared to Ctrl bucks. FL2 males show decreased diameter of tubulus seminiferi and an enhanced spermatid/Sertoli cell index. Sperm motility parameters of FL1 and Ctrl males are initially indistinguishable but FL1 spermatozoa show a better performance in a thermal stress experiment over a 5hours observation period. These data indicate that although selected for a primarily female-trait of high fertility also males from the fertility lines are effected by defined alterations in their reproductive organs. Some of these alterations are FL1-specific others are FL2-associated, indicating that different molecular strategies warrant the high-fertility phenotype on the female as well as on the male side.

Effects of reduced glutathione on ram sperm parameters, antioxidant status, mitochondrial activity and the abundance of hexose transporters during liquid storage at 5 ℃

The objective of this study was to evaluate the effects of reduced glutathione (GSH) supplementation on sperm motility, kinematics, mitochondrial activity and the abundance of sperm hexose transporters during liquid storage of ram semen at 5 ℃. Semen samples from eight rams were pooled and diluted with Tris-egg yolk extender supplemented with 0 (control), 50, 100 and 200 mM GSH, and they were assessed at different processing periods (fresh diluted, stored for 24, 48 and 72 h at 5 ℃). Sperm motility and kinematic parameters were analyzed by computer-assisted sperm analysis (CASA). The hypo-osmotic swelling test (HOST) was used to evaluate the functional integrity of sperm membrane. Eosin-nigrosin and JC-1/PI staining methods were used to assess sperm viability and mitochondrial activity, respectively. The antioxidant status of sperm was examined using biological assays. The changes of sperm hexose transporters (GLUT 3, 5 and 8) were determined using western blot analysis. As compared to the control group, supplementation with 100 and 200 mM GSH improved (P ＜ 0.05) sperm total motility, membrane integrity, motion parameters and mitochondrial activity at 72 h of storage, as well as resulted in the higher sperm antioxidant status after storage from 24 to 72 h. The cooling induced a reduction of GLUT 3 and 8, and GSH treatment at 200 mM retarded the gradual decline of these proteins during the prolonged storage time. Our data suggest that GSH can reduce the decline of semen quality by influencing the antioxidant status and energy metabolism of liquid stored ram spermatozoa. The concentration of 200 mM could be appropriate for ram semen during liquid storage at 5 ℃.

Identification of circular RNAs in porcine sperm and evaluation of their relation to sperm motility

Circular RNAs (circRNAs) are emerging as a novel class of noncoding RNAs which potential role as gene regulators is quickly gaining interest. circRNAs have been studied in different tissues and cell types across several animal species. However, a thorough characterization of the circRNAome in ejaculated sperm remains unexplored. In this study, we profiled the sperm circRNA catalogue using 40 porcine ejaculates. A complex population of 1,598 circRNAs was shared in at least 30 of the 40 samples. Generally speaking, the predicted circRNAs presented low abundances and were tissue-specific. Around 80% of the circRNAs identified in the boar sperm were reported as novel. Results from abundance correlation between circRNAs and miRNAs together with the prediction of microRNA (miRNA) target sites in circRNAs suggested that circRNAs may act as miRNA sponges. Moreover, we found significant correlations between the abundance of 148 exonic circRNAs and sperm motility parameters. Two of these correlations, involving ssc_circ_1458 and ssc_circ_1321, were confirmed by RT-qPCR using 36 additional samples with extreme and opposite sperm motility values. Our study provides a thorough characterization of circRNAs in sperm and suggests that circRNAs hold potential as noninvasive biomarkers for sperm quality and male fertility.

Stallion Semen Cooling Using Native Phosphocaseinate-based Extender and Sodium Caseinate Cholesterol-loaded Cyclodextrin-based Extender

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 resuspended at 100 million sperm/mL with their respective extender. Semen was then cooled and stored as in experiment 1. In both experiments, sperm motility parameters, plasma membrane integrity, and high mitochondrial membrane potential were assessed at 0, 24, and 48 hours post cooling. For experiment 3, 12 mares (n = 24 cycles) were bred with 48 hour–cooled semen from one stallion. Semen was processed as described in experiment 1. Mares had embryo flushing performed by 8-day post-ovulation. In experiment 1, BotuSemen Gold displayed superior total and progressive motility relative to INRA 96 (P < .05). There were no significant differences between the types of containers in any experiment. In experiment 2, INRA 96 and BotuSemen Gold extenders had similar total and progressive motility, but BotuSemen Gold had superior sperm velocity parameters at all timepoints. Embryo recovery was identical for both extenders (50%). Finally, the results obtained herein suggest that BotuSemen Gold is a suitable alternative to be included in semen cooling tests against INRA 96 in clinical practice.

Improvement of in vitro fertilization by a tannin rich vegetal extract addition to frozen thawed boar sperm

Boar spermatozoa are very susceptible to cryopreservation injuries and, for this reason, pig remains one of the few species in which fresh semen is still preferred to thawed one for routine artificial insemination (AI). The present work evaluated the effect of supplementing boar sperm thawing medium with Silvafeed SP (SSP), a mixture of Chestnut and Quebracho wood extracts (60/40 w/w) rich in polyphenols (92.4% tannin content) on in vitro fertilization (IVF) and on the following sperm parameters: sperm motility (assessed by CASA), viability, acrosome integrity, mitochondrial function and lipid peroxidation (assessed by flow cytometry) and capacitation status (immunolocalization of tyrosine phosphorylated proteins). Thawed spermatozoa were incubated 1 h at 37°C in BTS without (CTR) or with (5, 10, 20 µg/mL) SSP. After incubation sperm suspension was divided in three aliquots: one was used for IVF trials, one for sperm analysis, and the last one was capacitated for 1 h at 39°C 5% CO2 in IVF medium. Sperm motility parameters, viability, acrosome integrity, mitochondrial functionality, lipid peroxidation and tyrosine phosphorylated protein immunolocalization, used as capacitation parameter, were not influenced by SSP. However, oocytes inseminated with thawed spermatozoa pretreated with all the different SSP concentrations presented a significant (P < 0.01) increase in penetration rate compared to CTR. In addition, 5 µg/mL SSP exerted a positive effect (P<0.05) on the total efficiency of fertilization. These results encourage the use of SSP in the thawing medium since post-thawing fertility is a limit for the large-scale use of boar frozen semen.

Effect of alpha-lipoic acid on oxidative stress, viability and motility in the common carp (Cyprinus carpio) spermatozoa after short-term storage and cryopreservation

As known for different metabolic functions, α-lipoic acid (ALA) has been tested for spermatozoa preservation of animals as well as of human, but not for fish spermatozoa. The present study determined the effects of ALA on short and long-term (cryopreservation) preservation of common carp (Cyprinus carpio) spermatozoa, for the first time. For that, spermatozoa were diluted in extenders containing 0 (control), 0.025, 0.05, 0.1, 0.5, 1, 2, 5, and 10 mM of ALA concentrations in both short-term preservation and cryopreservation. Spermatozoa motility parameters by computer-assisted semen analysis, viability, lipid peroxidation and catalase activity in spermatozoa were conducted in both 2nd and 120th hours of short-term storage and post-thaw samples. Higher percentages of total spermatozoa motility (80 ± 3) and viability (87 ± 3) were observed in 0.5 mM ALA group after 120 h of incubation. In post-thaw samples, higher percentages of these parameters were in 1 mM ALA group (74 ± 3 and 83 ± 2, respectively). Moreover, the results have shown that the addition of ALA until concentrations of 2 mM improved especially spermatozoa curvilinear velocity, maintained viability, and suppressed excessive lipid peroxidation during the preservations. In conclusion, the additions of 0.5 mM ALA for short-term preservation and 1 mM ALA for cryopreservation were the optimal concentrations, and shown the protective effects on common carp spermatozoa, when considering all measured parameters together.

Optimization of Sperm Cryopreservation Protocol for Peregrine Falcon (Falco peregrinus).

Simple SummaryThe process of semen cryopreservation can have multiple advantages in an ex situ conservation program. However, there is a necessity to adapt the protocol to the specificity of each species. With that in mind, we aimed to optimize the sperm freezing/thawing process and study the effect of different cryoprotectants in the peregrine falcon.Sperm cryopreservation is a complex process that needs to be adapted to wild and domestic avian species to ensure proper efficiency. Because of its accessibility, the peregrine falcon may be used as a good model for studying other raptor species. To find the most optimal cryopreservation protocol for peregrine falcon ejaculates, sperm parameters such as motility, viability, DNA fragmentation, acrosome integrity, and mitochondrial activity were analyzed under different conditions by varying the freezing method (slow freezing in straws vs. ultrarapid freezing in pellets), thawing conditions (37 °C for 30 s vs. 5 °C for 1 min), type of cryoprotectant (DMA vs. DMSO), and concentration of DMSO (4% vs. 8%). Results show that slow cryopreservation in straws yielded greater percentages (p < 0.05) of motile spermatozoa (22.5% ± 4.4% vs. 0.0% ± 4.1%), viable spermatozoa with intact acrosomes (84.6% ± 4.3% vs. 77.4% ± 4.3%), and spermatozoa with active mitochondria (41.0% ± 6.7% vs.12.8% ± 6.7%), compared with those obtained by the ultrarapid freezing in pellets. However, no differences were found between different thawing conditions. Moreover, all sperm motility parameters were greater (p < 0.05) when DMSO was used during freezing compared with DMA, although the use of 3% and 8% DMSO produced similar results. In conclusion, these results represent important progress in the study of falcon semen cryopreservation protocol, highlighting the crucial steps of the process and the most suitable conditions.

Correlation between the sperm characteristics evaluated by a computerized system (CASA) and the reproductive performance of sows

Computer-assisted semen analysis (CASA) systems have been one the most used tools to evaluate sperm kinetics. The objective of this research was to estimate the correlation between sperm motility characteristics evaluated by CASA during 72 hours of cooling with the farrowing rate (FR) and total number of piglets born (TNB) after artificial insemination. Multiparous sows (n=464) were inseminated with semen from seven boars (19.6 ± 1.3 ejaculates/male). Sperm motility parameters were determined immediately after dilution and after 24, 48 and 72 hours of cooling at 15°C: total motility (TM-%), progressive motility (PM-%), curvilinear velocity (VCL-μm/s), straight line velocity (VSL-μm/s), average path velocity (VAP-µm/s), amplitude of lateral head displacement (ALH-µm), flagellar beat cross frequency (BCF-Hz), straightness (STR-%) and linearity (LIN-%). Pearson’s correlation coefficient was applied to analyze the data and the comparison of the means of the sperm characteristics between the boars was done by Tukey’s test at 5% probability. TM and PM at time zero (T0) were significant and had a moderate to high correlation with FR and TNB. After 72 hours of refrigeration, the semen quality was reduced and showed a significant and low correlation of the TM and PM with these same parameters. The boar presenting the lowest value of TM and PM after dilution obtained lower FR and TNB. In conclusion, computer-assisted semen analysis soon after dilution can be used to predict fertility of boars.

Does single-layer centrifugation with Bovicoll improve sperm quality of frozen-thawed semen in Fleckvieh bulls?

The aim of the present study was to evaluate the effect of sperm selection by single-layer centrifugation (SLC) performed before freezing on sperm quality after thawing of Fleckvieh bull semen. Ejaculates from 22 bulls were collected by artificial vagina and divided into two aliquots. One aliquot (control sample) was diluted with Steridyl® and frozen over nitrogen vapour in a Digitcool freezer (IMV Technologies). Sperm from the second aliquot (SLC sample) was selected using the SLC technique with Bovicoll colloid and then frozen over nitrogen vapour in a Digitcool freezer. After thawing, both samples (control and SLC) were evaluated by computer-aided sperm analysis (CASA; SCA 6.4 System; Microptic S.L) for sperm motility parameters. Integrity of the plasma membrane (viability), high mitochondrial membrane potential (HMMP) and acrosome integrity were assessed using a Guava® easyCyte flow cytometer (IMV Technologies). Morphological examination of spermatozoa was performed by Differential Interference Contrast microscopy (Leica DMi8). Morphological examination of live, immobilized spermatozoa was analysed under high magnification (≥6,600×). After thawing, the mean sperm viability of the control sample was 51.57%, compared to 40.37% for the SLC sample (p<.01). HMMP was higher (p<.01) in the control sample (40.37% versus 28.96%), and the mean of live spermatozoa with damaged acrosome was significantly higher (p<.03) in the SLC sample (1.63% versus 1.95%). The mean percentage of motile spermatozoa was 80.17% in the control sample, compared to 75.14% in the SLC sample (p<.0195), and rapid subpopulation reduced from 20.08% to 8.99% (p<.0001) after SLC. Percentage of hyperactivated sperm decreased from 12.23% to 4.28% (p<.0001) after SLC. Given the overall results, the sperm quality of thawed Fleckvieh bull semen was not improved when sperm were selected by SLC before freezing.