Semen Evaluation Research Articles

Production and Evaluation of Frozen Semen of Sahiwal Bull Using Two Different Diluters and Protocols

The experiment was designed to evaluate the effects of diluters and freezing protocols on frozen semen production of Sahiwal breeding bulls. Imported 100% Sahiwal breeding bulls were then reared in the American dairy limited (ADL). Semen samples are processed and frozen once a week in the ADL. Spermatozoa with various motility patterns were collected; these included post-thawing, progressive, fast, slow, circular, and local. Computer Assisted Semen Analyzer (CASA) was used to measure the straight-line velocity (VSL), curvilinear velocity (VCL) and linearity (LIN) parameters. Two diluters (tris egg-yolk and tris egg-yolk aqua) and freezing protocols (slow freezing and fast freezing) were used for frozen semen production of these bulls. The significantly highest (P<0.05) number of post-thawing total, progressive and progressive fast motility (78.42±1.88, 73.50±1.90 and 59.68±2.10%), VCL (115.31±3.82µm/s) and lowest slow, circular and local motility (13.44±1.40, 0.57±0.08 and 4.92±0.41%), LIN (0.39±0.01%) was found in diluter 1 and freezing protocol 2. On the other hand, highest number of circular motility (0.77±0.18%) and VSL (44.83±1.17µm/s) was found in diluter 1 and freezing protocol 1 whereas highest number of slow and local motility (18.84±1.23 and 7.46±0.57%) and LIN (0.61±0.20%) and lowest progressive fast motility (48.73±2.28%), VCL (98.90±3.42µm/s), VSL (39.76±1.33µm/s) was found in diluter 2 freezing protocol 2 and also lowest post-thawing and progressive motility (73.44±1.94 and 66.51±2.02%) was observed in diluter 2 freezing protocol 1. The study reveals that diluter 1(one step egg yolk based) and freezing protocol 1 (5 steps) and 2 (2 steps) seems to be an efficient method for frozen semen production.

Evaluation of specific fractions of Large White Yorkshire boar semen

The current study was conducted to evaluate the quality of specific fractions of three Large White Yorkshire (LWY) boar semen samples. Fifteen semen samples were collected by gloved hand method as an initial 10 mL of sperm rich fraction (SRF- F1), remaining SRF (F2) and whole semen. Samples were evaluated for colour, volume, pH, sperm concentration and progressive motility. The first 10 mL of SRF of boar semen contained 25 per cent of the total ejaculate spermatozoa and was able to withstand cooling, freezing and thawing better than the spermatozoa of the bulk ejaculate. A significant difference between the fractions was observed in the volume of semen ejaculated, pH and sperm concentration across different fractions of the same ejaculate. The differences in semen fractions were attributed to the differences in the composition of seminal plasma of the initial fraction of SRF Keywords: Boar semen, sperm rich fraction, semen evaluation

Effect of a single dose of letrozole on ejaculation time, semen quality, and testicular hemodynamics in goat bucks subjected to heat stress.

Letrozole (LTZ) is an aromatase inhibitor that limits estrogen (E2) production and increases testosterone (T) levels. This research aimed to examine the impact of a single dose of LTZ on testicular hemodynamics, ejaculation time, and semen quality in goats under heat stress (HS). Therefore, Doppler examination and semen evaluation were performed on twelve mature bucks for two weeks (W-1, W-2) as pre-heat stress control during winter. Then during summer HS bucks were subjected to Doppler examination, semen evaluation, and hormonal analysis (T, E2, and LH) at 0h. Afterward, bucks were assigned into two groups and subcutaneously injected with physiological saline (n = 6; CON) or LTZ (0.25mg/kg BW; n = 6; LTZ). Both groups were subjected to Doppler scanning and hormonal analysis at 2, 4, 24, 48, 72, 96,144, and 168h. Semen evaluation was performed at 48 and 168h. The LTZ group showed increasing (P < 0.05) in semen volume, sperm motility, and viability and decreasing (P < 0.05) in ejaculation time and sperm abnormalities compared to CON group at 48h. Additionally, T concentrations increased (P < 0.01) at 2, 24, and 48h, E2 decreased (P < 0.01) from 2 to 48h, and LH raised (P < 0.01) at 2 and 72h in LTZ group compared to CON one. Doppler indices reduced (P < 0.05) at 96h in LTZ group. semen pH and scrotal circumference were not affected by LTZ. In conclusion, LTZ administration shortened ejaculation time and enhanced semen quality in bucks during HS.

Predicting Boar Sperm Survival during Liquid Storage Using Vibrational Spectroscopic Techniques.

Artificial insemination (AI) plays a critical role in livestock reproduction, with semen quality being essential. In swine, AI primarily uses cool-stored semen adhering to industry standards assessed through routine analysis, yet fertility inconsistencies highlight the need for enhanced semen evaluation. Over 10-day storage at 17 °C, boar semen samples were analyzed for motility, morphology, sperm membrane integrity, apoptosis, and oxidative stress indicators. Additionally, machine learning tools were employed to explore the potential of Raman and near-infrared (NIR) spectroscopy in enhancing semen sample evaluation. Sperm motility and morphology gradually decreased during storage, with distinct groups categorized as "Good" or "Poor" survival semen according to motility on Day 7 of storage. Initially similar on Day 0 of semen collection, "Poor" samples revealed significantly lower total motility (21.69 ± 4.64% vs. 80.19 ± 1.42%), progressive motility (4.74 ± 1.71% vs. 39.73 ± 2.57%), and normal morphology (66.43 ± 2.60% vs. 87.91 ± 1.92%) than their "Good" counterparts by Day 7, using a computer-assisted sperm analyzer. Furthermore, "Poor" samples had higher levels of apoptotic cells, membrane damage, and intracellular reactive oxygen species on Day 0. Conversely, "Good" samples maintained higher total antioxidant capacity. Raman spectroscopy outperformed NIR, providing distinctive spectral profiles aligned with semen biochemical changes and enabling the prediction of semen survival during storage. Overall, the spectral profiles coupled with machine learning tools might assist in enhancing semen evaluation and prognosis.

PSXIII-16 Kinematics and viability of spermatozoa from Criollo and Angus bulls in a low-input system

Abstract The Chihuahuan Criollo cattle is a genetic resource with special adaptive characteristics to rough, arid and semi-arid ecosystems. There is a great interest in incorporating such adaptations into beef production systems as one of the strategies for sustainable production to face the climate change challenge. Reproductive success is determinant to achieve sustainability in animal production systems. Most research efforts to improve reproductive efficiency have focused on the study of females. However, it has been shown that much of the economic losses caused by embryonic deaths are due to the bull. The aim of this study was to evaluate the quality of thawed semen from Angus and Criollo bulls kept in a low-input system. Thirteen healthy bulls (n = 7 Angus and 6 = Chihuahuan Criollo), with an average age of 4.3 ± 0.48 yr, grazing in the same rangeland and subjected to natural pasture conditions, were studied. Semen obtained by electroejaculation, were frozen and subsequently thawed for analyses. Characteristics of motility, vitality (V), morphology (M) and plasma membrane integrity (PMI) were evaluated in fresh and thawed semen, whereas movement kinetics (MK) and acrosome reaction (AR) were evaluated in thawed semen. Statistical analysis was performed using the PROC GLM of SAS Studio 2022 including the fixed effects of breed, semen condition and their interaction for variables of progressive motility (PM), V, M, PMI and the effect of breed for variables of MK and AR. As expected, PM and V were less (P &amp;lt; 0.05) in thawed (57.93 ± 4.1 and 53.69 ± 2.7) compared with fresh (72.64 ± 3.9 and 74.98 ± 2.6) semen, whereas no differences (P &amp;gt; 0.05) were found in M. However, the interaction between semen condition and breed for PMI tended to the significance (P = 0.08) with a PMI of 68.95 ± 4.6 and 58.98 ± 4.3 for Criollo and Angus, respectively, in fresh semen, whereas in thawed semen this difference was inverted (52.08 ± 4.6 and 58.40 ± 4.6). It is important to note that the semen retained quality traits with values above the minimum accepted for use in artificial insemination, despite the scarcity of resources of the system in which the animals were kept. There were no differences by breed in the main MK variables; however, AR reported as live unreacted sperm was less (P &amp;lt; 0.05) in Criollo (40.49 ± 4.49) compared with Angus bulls (58.48 ± 4.49). In conclusion, it is important to incorporate the assessment of membrane integrity and acrosome reaction in thawed semen evaluation to obtain a more accurate prediction of the potential fertility of samples, even if motility parameters appear to be optimal.

Flow cytometry evaluation of Dalagh ram post-thaw semen incorporated with curcumin-loaded niosomal nanocarriers

Optimization of cryopreservation media to attenuate oxidative damage on post-thaw spermatozoa is crucial. This study aimed to assess the antioxidant functionality of curcumin-loaded niosomal nanocarriers (CurLNN) on the quality of cryopreserved ram spermatozoa. Semen samples (n=16) of four Dalagh rams were extended with different concentrations (10 or 20 μL) of curcumin (Cur) or CurLNN and then cryopreserved using a standard protocol. The findings indicated that sperm cells exposed to curcumin, specifically CurLNN at a concentration of 20 μL, displayed increased levels of total and progressive motility, greater lateral head displacement and linearity values, improved plasma membrane function, and reduced abnormality (P < 0.05). Viability and plasma membrane functionality increased by supplementing cryopreservation medium with Cur20 and CurLNN (P < 0.05). A significant decline (P < 0.05) in malondialdehyde (MDA) and reactive oxygen species (ROS) concentrations was observed by supplementing cryopreservation medium with both curcumin and its nano-sized. There was a significant (P < 0.05) increase in the percentage of spermatozoa with mitochondrial activity when the cryopreservation medium was incorporated with CurLNN. Spermatozoa treated with CurLNN demonstrated higher viability and lower late apoptosis (P < 0.05) compared with other groups. There were no differences among groups in terms of the percentage of necrotic and early apoptotic spermatozoa. In conclusion, the results of this study demonstrated that nano-sized curcumin, particularly at a 20 µL concentration, had a higher antioxidant potential, leading to improved sperm functional quality by mitigating oxidative damage during cryopreservation.

Comparison of computer-assisted sperm analysis and smartphone-applied sperm analysis for evaluation of frozen-thawed bull semen.

This study aims to compare the efficacy of computer-assisted sperm analysis (CASA) and smartphone-applied sperm analysis (SASA) in assessing the quality of frozen-thawed bull semen. A total of 75 straws (n = 75) semen samples were used from different production batches of five Holstein bulls. The semen analyses were conducted in three groups: Group I (CASA-37°C), semen samples were evaluated using the CASA system at 37°C (n = 25); Group II (SASA-25°C), semen samples were assessed using the SASA system at a temperature of room heat (25°C) (n = 25); and Group III (SASA-37°C), semen samples were evaluated using the SASA system at 37°C (n = 25). The frozen-thawed bull semen samples were analysed in terms of total motility (TM), progressive motility (PM), immotile, velocity average path (VAP), velocity curve linear (VCL), velocity straight line (VSL) and sperm concentration. There was no significant difference between the groups in terms of spermatozoa concentration (p > .05). However, significant differences among the groups were observed for total motile spermatozoa values (p < .001). Values of progressive motile spermatozoa were lower in Group I and Group II compared to Group III (p < .001). The immotile spermatozoa values were significant between the groups (p < .001) and were found to be proportional to total motile spermatozoa values. Additionally, the VAP, VCL and VSL values were comparable between Group II and Group III, but lower when compared to Group I. In conclusion, the results of the study demonstrate that the Sperm Cell™ system can accurately analyse the concentration of frozen-thawed bull semen. The analyses performed at room temperature indicate a parallelism between the PM value and CASA results. However, it is thought that SASA devices require a series of standardization studies in different semen extenders, different sample concentrations and different animal species, analogous to the standardization evolution process of CASA devices in semen analysis.

N-Acetylcysteine Supplementation Improves Testicular Haemodynamics, Testosterone Levels, Seminal Antioxidant Capacity and Semen Quality in Heat-Stressed Goat Bucks.

Heat stress (HS) disrupts testicular homeostasis because of oxidative stress. N-acetylcysteine (NAC) is a thiol compound with antioxidants, anti-inflammatory and anti-apoptotic properties. As a sequel, this research aimed to assess the ameliorative effects of NAC supplementation on the reproductive performance of goat bucks kept under environmental HS. Primarily, Doppler examination as well as semen collection and evaluation were conducted on 12 mature bucks for 2 weeks (W) as pre-heat stress control (W1 and W2) during winter (February 2023). The temperature-humidity index (THI) was 63.4-64.3 (winter season). Then during summer HS conditions (from the beginning of July till the end of August 2023) bucks were assessed before NAC supplementation (W0), afterwards they were arbitrarily assigned into two groups. The control group (CON; n = 6) received the basal diet while the NAC group (n = 6) received the basal diet in addition to oral NAC daily for 7 weeks (W1-W7). The THI was 78.1-81.6 (summer season). Testicular blood flow parameters, serum concentration of nitric oxide (NO) and testosterone were measured. Additionally, total antioxidant capacity (TAC) and malondialdehyde (MDA) content in seminal plasma and semen quality parameters were evaluated. There were marked reductions (p < 0.05) in the resistive index (RI; W1, W4 and W5), pulsatility index (PI; W2 and W4-W7), and systolic/diastolic ratio (S/D; W4-W7) in the NAC group compared to the CON group. Furthermore, testosterone and NO levels were higher (p < 0.01 and p < 0.05, respectively) in the NAC group (W2, W3, W5 and W3-W5, respectively). Seminal plasma TAC increased (p < 0.05) and MDA decreased (p < 0.05) in the NAC group (W2, W4 and W5) compared to the CON group. Moreover, there were marked improvements (p < 0.05) in semen quality parameters (mass motility, total motility, viability and normal morphology) in the NAC group. In conclusion, oral NAC supplementation could be used to enhance the reproductive performance of goat bucks during HS conditions which is supported by remarkable enhancement in testicular haemodynamics, NO, testosterone levels and semen quality parameters.

P-102 Incorporation of an automated sperm counting method: a matter of time

Abstract Study question Is a fully-automated system a faster and reliable sperm counting method? Summary answer A fully-automated sperm counting system is a reliable tool that significantly reduces time spent for semen evaluation. What is known already The emergence of automated tools in our everyday life is a reality. Fully-automated systems for the evaluation of seminal samples are available for the andrology laboratories. The expected advantages of such analysers would be a higher reliability and reproducibility of results, with lower inter and intra-individual variability. Although there are few published validation studies, available data have evidenced that results obtained with these methods are, in general, consistent with those obtained with manual/conventional analysis. Moreover, automatic sperm assessment could be useful in reducing the time spent for analysis. However, no data have been published in this regard. Study design, size, duration Observational retrospective study of 220 semen samples analyzed in 2022. Every sample was analyzed using three different methodologies: manual analysis, semi-automated and fully-automated. Manual assessment was performed by senior embryologists undergoing periodical quality control evaluations and included counting with improved Neubauer chamber, motility assessment with slide with phase-contrast optics, and morphology analysis by Diff-Quick stain. The semi-automated analysis was performed by SCATM plus manual morphology assessment. The fully-automated analysis was exclusively performed by SQA-VisionTM. Participants/materials, setting, methods Male patients consulting for infertility and undergoing a spermiogram in a single centre. The sperm parameters evaluated for correlation study among methodologies were: total sperm, motile sperm and progressive sperm concentrations (106 per ml), and morphologically normal forms (%). Duration of analysis (sperm count + motility + morphology) using each method was recorded and compared. Intraclass correlation coefficient (ICC[95%CI]) was used to compare agreement between methodologies and Wilcoxon’s test was used for time (median (p25-p75)). Main results and the role of chance The observed correlation between the three methodologies was moderate (ICC=0.5-0.79) to high (ICC &amp;gt;0.8) for all parameters analysed. Concerning total sperm concentration, both fully and semi-automated systems showed a high correlation with manual assessment (ICC=0.93 (0.90-0.95) fully-automated vs manual; ICC=0.83 (0.73-0.89) semi-automated vs manual, respectively). Concerning motile sperm concentration and progressive motile sperm concentration, both fully and semi-automated systems showed a high or moderate correlation with manual assessment (ICC =0.86 (0.77-0.91) and ICC=0.87 (0.76-0.92) fully-automated vs manual; ICC=0.54 (0.27-0.70) and ICC=0.62 (0.39-0.76) semi-automated vs manual, respectively). In terms of normal forms, fully-automated showed a moderate correlation with manual assessment (ICC=0.69 (0.57-0.78)). Regarding time spent for analysis, both fully and semi-automated analysers showed to be faster than manual method (p &amp;lt; 0.001) (1343 (1199-1472) seconds faster fully-automated vs manual and 608 (429-769) seconds faster semi-automated vs manual). Comparing time spent for analysis between the fully and the semi-automated systems, fully-automated was 707 (610-784) seconds faster (p &amp;lt; 0.001) than the semi-automated. Limitations, reasons for caution Retrospective study. SCATM is a semi-automated operator-dependent system based on image analysis and its accuracy relies on the number of sperm analysed and the operators’ ability in reviewing images. SQA-VisionTM is fully-automated, based on electro-optical signal analysing millions of sperm. Therefore, conclusions may not apply to other software operators. Wider implications of the findings Fully-automated systems for sperm assessment can be reliably introduced in andrology laboratories offering a dramatic reduction in the time spent for analysis. However, such analysers may only be cost-efficient in centres with a very high activity. Trial registration number not applicable

Analisis Semen Babi Yorkshire dalam Modifikasi Extender Alami Buah Lontar dalam Filtrat Jambu Air Hijau (Syzygiumaqueum)

This study aims to see the effectiveness of Yorkshire boar semen natural modification of the natural diluent of green guava filtrate (Syzygiumaqueum). Semen is stored per 72 hours from a 3-year-old sow. Macro and microscopic evaluation of semen includes Volume, Colour, Consistency, pH, Odor and motility, concentration, viability, abnormality and normality of spermatozoa. The semen was diluted with palm fruit juice and added green guava filtrate (Syzygiumaqueum) in tiers: 100μL, 200μL, 300μL, 400 μL and 500 μL. The results showed that the modification of palm fruit juice diluent with the addition of 200 μL of green guava filtrate (Syzygiumaqueum) in Yorkshire boar semen was able to maintain the quality of spermatozoa up to 22 hours of storage.

Association of lifestyle and occupational exposure factors with human semen quality: a cross-sectional study of 1060 participants

The incidence of male infertility (MI) is rising annually. However, the lifestyle and occupational exposure factors contributing to MI remain incompletely understood. This study explored the effects of self-reported lifestyle and occupational exposure factors on semen quality. Among 1060 subjects invited to participate, 826 were eligible. The participants’ general characteristics, lifestyle, and occupational exposure factors were collected immediately before or after semen evaluation through an online questionnaire. Initially, univariate analysis was used to investigate the relationship between the abovementioned factors and semen quality. The results indicated significant associations between low semen quality and various factors, including age, BMI, infertility type and duration, abstinence time, semen and sperm parameters, smoking, alcohol consumption, irregular sleep habits, and frequent exposure to high temperatures and chemicals at work (p < 0.05). Then, multivariate analysis was conducted to identify factors independently associated with low semen quality. Adjustment for relevant confounders was achieved by including factors with a p-value < 0.25 from univariate analyses as covariates in the binomial and ordered logistic regression models. The results suggested that alcohol consumption was a positive factor for sperm concentration (odds ratio [OR] = 0.60; 95% confidence interval [CI] = 0.36–0.99; p = 0.045). The groups with a BMI ≥ 24 and <28 kg/m2 showed a significant decrease in sperm progressive motility when compared to the reference group (BMI < 24 kg/m2) (OR = 0.63; 95% CI = 0.46–0.87, p = 0.005). In addition, the groups that drank green tea <1 time/week (OR = 1.52, 95% CI = 1.05–2.2) and 1–4 times/week (OR = 1.61, 95% CI = 1.02–2.54) exhibited significantly increased sperm DFI values compared with the group that drank green tea 5–7 times/week. In conclusion, these findings underscore the importance of maintaining a normal weight and regularly consuming green tea for men.

Ultrasonography of Testicular Maturation and Correlation with Body Growth and Semen Evaluation in Beagle Dog Model.

This prospective study investigated the ultrasonographic appearance of the canine testis from birth to adulthood. Eight purpose-bred laboratory Beagle-breed dogs were monitored from 4 to 40 weeks of life. The following parameters were evaluated every two weeks: bodyweight and height, scrotal and testicular volume, ultrasonographically measured testicular volume, echogenicity, heterogeneity, blood-flow score, ratio of the grayscale intensity value of the testis to the capsule, ejaculate volume, motility, viability, and number of spermatozoa. A correlation analysis was carried out between the various measurements obtained. Fertility was achieved in the 36th week of life. The echogenicity of the testicular parenchyma increased with age, and subsequently to the 30th week of life remained constant. The heterogeneity of the testicular parenchyma, as was evaluated by the standard deviation of the values of grayscale intensity of the parenchyma, also increased with age and was >19 at the onset of fertility. The ratio of grayscale intensity of testicular parenchyma had values < 200 at maturity. A colour Doppler evaluation first detected blood flow in the testis in the 22nd week. After the 32nd week, distinct signals were visible. In the 36th week, >80% of the testes imaged had visible vessels. A significant correlation was found between all the evaluation methods. The findings of the study may help clinicians detect the onset of fertility in dogs, especially when semen evaluation is not feasible; however, their applicability in all breeds or individuals might possibly vary due to genetic, physiological, and developmental differences. In summary, the study ultrasonographically explores the testicular maturity in dogs, with the aim to improve clinical assessments and health management in these animals.

La presencia de cuernos determina el rango social de carneros Dorper

The aim was to determine if the presence of horns on rams defines whether they are high-ranking or low-ranking. Dorper rams (n = 20) were evaluated to determine their social rank (SR), either high (HSR) or low (LSR), under intensive management conditions. Upon SR classification, the morphometric variables horns presence (HP) or absence (HA) and distance between them, the height at the withers, body length, thoracic perimeter, and scrotal circumference, male odor, body weight, and body condition score were registered. Afterward, semen evaluations were performed, recording the ejaculated volume, sperm concentration, and mass motility. There were no differences (p &gt; 0.05) for the morphometric variables between the groups. However, HP as well as semen quality were higher (p &lt; 0.05) in HSR males. The presence of horns in the rams determined their social rank and the dominant males showed better semen quality than the subordinate males.

Fatty acid composition and biophysical characteristics of the cell membrane of feline spermatozoa

Sperm membrane composition and biophysical characteristics play a pivotal role in many physiological processes (i.e. sperm motility, capacitation, acrosome reaction and fusion with the oocyte) as well as in semen processing (e.g. cryopreservation). The aim of this study was to characterize the fatty acid content and biophysical characteristics (anisotropy, generalized polarization) of the cell membrane of domestic cat spermatozoa. Semen was collected from 34 adult male cats by urethral catheterization. After a basic semen evaluation, the fatty acid content of some of the samples (n = 11) was evaluated by gas chromatography. Samples from other individuals (n = 23) were subjected to biophysical analysis: membrane anisotropy (which is inversely proportional to membrane fluidity) and generalized polarization (describing lipid order); both measured by fluorimetry at three temperature points: 38 °C, 25 °C and 5 °C. Spermatozoa from some samples (n = 10) were cryopreserved in TRIS egg yolk-glycerol extender and underwent the same biophysical analysis after thawing. Most fatty acids in feline spermatozoa were saturated (69.76 ± 24.45%), whereas the polyunsaturated fatty acid (PUFA) content was relatively low (6.12 ± 5.80%). Lowering the temperature caused a significant decrease in membrane fluidity and an increase in generalized polarization in fresh spermatozoa, and these effects were even more pronounced following cryopreservation. Anisotropy at 38 °C in fresh samples showed strong positive correlations with viability and motility parameters after thawing. In summary, feline spermatozoa are characterized by a very low PUFA content and a low ratio of unsaturated:saturated fatty acids, which may contribute to low oxidative stress. Cryopreservation alters the structure of the sperm membrane, increasing the fluidity of the hydrophobic portion of the bilayer and the lipid order in the hydrophilic portion. Because lower membrane fluidity in fresh semen was linked with better viability and motility after cryopreservation, this parameter may be considered an important factor in determination of sperm cryoresistance.

Evaluation of equine semen frozen in extenders free of egg yolk using two different freezing curves

A chemically defined cryopreservation extender that maintains seminal parameters is relevant. Fifteen ejaculates from 5 stallions (n= 5; r=3) were diluted in 5 extenders: 1) EDTA-glucose based extender with egg-yolk and dimethylformamide (EY); 2) commercial equine extender (CE); 3) CE with dimethylformamide (CE-3); 4) bovine commercial extender with liposomes (OP); 5) bovine commercial extender with soybean lecithin (BIO), and frozen using a slow and a rapid temperature descent curve. Post-thaw evaluations were: sperm kinematic parameters, viability and acrosome status, membrane lipoperoxidation and DNA fragmentation. Sperm data were analysed using an ANOVA or Friedman test (results mean ± SD). Paired comparison between the two freezing curves was analysed using the Wilcoxon test. Total and progressive motility were significantly higher (P<0.05) in the EY and CE-3 samples using the slow curve, whereas for the fast curve, total and progressive motility were significantly higher (P<0.05) in the EY samples compared to all the extenders and the samples frozen in CE-3 were significantly higher than the remaining extenders (P<0.05). The percentages of live acrosome intact sperm and of live non-peroxidized sperm were significantly higher (P<0.05) in the EY extender when using either of the freezing curves and in turn, were significantly higher (P<0.05) in samples frozen in CE-3 compared to the remaining extenders. Intact DNA was significantly lower (P<0.05) in the BIO extender, using the rapid curve. To conclude, the commercial equine extender with 3% dimethylformamide, without egg-yolk, could be a suitable alternative for extenders with egg-yolk.

High percentage of midpiece defects in Brangus bull sperm with no reduction in sperm kinematics.

The study investigated midpiece defects in sperm from a 5-year-old Brangus bull with a high rate of semen batch rejection, due to morphologically abnormal sperm, with no reduction in sperm kinematics. A comprehensive evaluation was conducted over a 16-month period, involving 28 ejaculates. Notably, despite the high proportion of midpiece defects (average 37.73%, from 3% to 58%), the study revealed stable sperm production, with no discernible differences in the kinematic data before and after cryopreservation. Electron microscopy identified discontinuities in the mitochondrial sheath, characteristic of midpiece aplasia (MPA). The anomalies were attributed to be of genetic origin, as other predisposing factors were absent. Additionally, the electron microscopy unveiled plasma membrane defects, vacuoles and chromatin decondensation, consistent with previous findings linking acrosome abnormalities with midpiece defects. The findings underscored the necessity of conducting thorough laboratory evaluations before releasing cryopreserved semen for commercialization. Despite substantial morphological alterations, the initial semen evaluation data indicated acceptable levels of sperm kinematics, emphasizing the resilience of sperm production to severe morphological changes. This case report serves as a contribution to the understanding of midpiece defects in bull sperm, emphasizing the need for meticulous evaluation and quality control in semen processing and commercialization.

Kualitas Semen Babi yang ditambahkan Antioksidan Semangka Merah (Citrullus Lanatus)

This study is aimed at recognizing the effect that the addition of juice from watermelon (Citrulus lanatus) as antioxidants against motility and semen viability of pigs. The source of cement used in research came from the landrace boar. The semen of the pig was collected using the method of massage with the help of dummy sow. Cement is macroscopic and microscopic. The natural dilution material used is lontar. The treatment of the juice of 100 µL/10 mL, 200 µL/10 mL, 300 µL/10 mL, 400 µL/10 mL, 500 µL/10 mL. Liquid semen evaluations are performed every two hours after storage and sperm motility and viability evaluations of spermatozoa. The addition of watermelon juice (Citrulus lanatus) P3 treatment group at a dose of 300 µL/10mL has the best possible result in sustaining the motility of spermatozoa.

Cumulus matrix selection leads to isolation of spermatozoa with better motility, morphology, and lower DNA fragmentation.

The objective of this study was to compare the motility, morphology, and levels of DNA fragmentation of spermatozoa subjected to conventional swim-up or cumulus matrix (CM) sperm selection. Semen samples were collected from 60 normozoospermic men at a private hospital between December 2021 and March 2022. After liquefaction, semen samples were separated into two portions - one part was subjected to conventional swim-up preparation and the remaining spermatozoa were subjected to CM selection. The CM was obtained by mechanical isolation from healthy donor oocytes. Semen analysis and evaluation of sperm were performed according to WHO 6th Edition Laboratory Manual and Kruger's strict criteria, respectively. Sperm DNA fragmentation (SDF, %) of the two preparations was evaluated using Halosperm G2 detection kit (Halotech, Madrid, Spain). Wilcoxon rank-sum test was used to compare the characteristics of spermatozoa obtained by the two preparations. Spermatozoa selected by CM showed significantly better rapidly progressive motility (43.5% vs. 30.6%, respectively, p<0.001), higher percentage of morphologically normal forms (14.0% vs. 9.0%, respectively, p<0.05), and lower levels of SDF (26.0% vs. 45.0%, p<0.05) in comparison to those prepared by conventional swim-up. Moreover, the incidence of multiple sperm defects was considerably lower in the samples that underwent CM selection compared to those that did not (30.0% vs. 49.0%, respectively, p<0.05).The selection by CM significantly increases sperm motility and reduces morphologically abnormal spermatozoa and DNA fragmentation rates in comparison to the conventional swim-up preparation. The application of this selection technique may increase chances of successful IVF outcomes.

Therapeutic effects of vitamin B17 against anabolic steroid Trenorol induced testicular toxicity, injury, DNA damage and apoptosis in male rats.

Several anabolic androgenic steroids (ASSs) are a group of synthetic molecules derived from testosterone and developed mainly for veterinary use that classed as a Schedule III and sometimes utilized by athletes to enlarge their muscles. Abuse of anabolic androgenic steroids can result in severe organ damage that cannot be repaired. Therefore; the objective of the current investigation was to examine the therapeutic effects of vitamin B17 (VitB17) on the testicular toxicity caused by the anabolic steroid Trenorol in male rats. Rats were randomly assigned into control, VitB17 (50mg/kg b.wt./day, orally), Trenorol (received 10mg/kg b.wt./week, IM) and Trenorol+VitB17 treated groups. At the end of experiment, hormonal assay, semen evaluation, testicular enzymes, and DNA damage were assessed. Besides, the histopathological and immunohistochemical investigations of the P53 expression were performed. Current results revealed that; Trenorol induced significant depletion in relative weights of testis (RWT), total testosterone follicle stimulating hormone (FSH), and luteinizing hormone (LH), sperm count, morphology index, viability, progressive motility, and testicular injury and a significant increase sperm abnormalities, testicular DNA damage and P53 experssions. Treatment of rats with Trenorol+VitB17 decreased the testicular toxicity, sperm parameters, DNA damage and apoptosis. We can conclude that; Trenorol induced toxicity, DNA damage and apoptosis in rat testis and treatments with VitB1 improved these parameters.

Evaluation of the effect of the addition of an olive oil-derived antioxidant (Pectoliv-80A) in the extender for cryopreservation of rooster sperm through the use of a discriminant statistical tool

During the poultry sperm cryopreservation process, an excess of reactive oxygen species is generated resulting in oxidative stress which harms the quality of avian spermatozoa. To counteract this effect, the addition of exogenous antioxidants, such as Pectoliv-80A (a by-product of olive oil), to the cryopreservation diluent is interesting. For this purpose, 16 roosters belonging to the Utrerana avian breed were used. Six semen pools (from the 6 different replicates) were divided into 4 aliquots corresponding to different concentrations of Pectoliv-80A that were tested (0, 300, 400, and 500 μg/mL), and the cryopreservation process was carried out. To evaluate post-thawing semen quality, different parameters such as motility, membrane functionality, reactive oxygen species production, lipid peroxidation, and acrosome integrity were studied. A discriminant canonical analysis was used to determine both the differences between the Pectoliv-80A concentration groups and the discriminant power of the aforementioned parameter used for semen evaluation. Total motility and membrane functionality were reported to be the most discriminant variables for differentiating the different antioxidant enrichment groups and concluded that concentrations of 300 μg/mL showed the most desirable quality of post-thawing semen. The present study could lead to the optimization of both cryopreservation and quality evaluation techniques of the sperm of rooster species, that support the conservation program of endangered local breeds.