Sperm Characteristics Research Articles

Ameliorative impacts of carvacrol on DNA integrity, oxidative stress, and sperm quality in asthenozoospermic infertile individuals during cryopreservation.

Nowadays, the cryopreservation process can produce reactive oxygen species (ROS), which cause damage to the motility, cell membrane, and DNA integrity of sperm. This study is aimed at evaluating the impact of carvacrol, as a powerful antioxidant, on sperm features and improving oxidative stress throughout the cryopreservation procedure. In this prospective study, semen samples from 25 patients with asthenozoospermia were separated into three groups: fresh, freezing, and freezing + carvacrol (a dose of 100µM). The subsequent parameters were evaluated using standard methods in all three groups: sperm motility according to WHO criteria, sperm morphology using Papanicolaou staining, sperm viability with eosin-nigrosin staining, DNA integrity with acridine orange staining, levels of antioxidant enzymes (catalase, glutathione, and superoxide dismutase), total antioxidant capacity (TAC), and malondialdehyde (MDA) using ELISA. Also, DNA fragmentation was analyzed by the SDFA kit, and mitochondrial membrane potential (MMP) was assessed by rhodamine staining. Average sperm viability, motility, mitochondrial membrane potentiality, integrity of sperm membrane, and antioxidant enzyme levels are meaningfully reduced in the freezing group in contrast to the control group. The freezing group showed a meaningful rise in the mean MDA levels and DNA fragmentation compared to the control group. In the freezing group supplemented with carvacrol, a meaningful rise could be visible in mean percentages of viability, motility, and antioxidant enzyme levels, whereas mean levels of MDA and DNA fragmentation meaningfully declined in contrast to the freezing group. The results demonstrate that carvacrol, a potent antioxidant, offers significant protection against the loss generated by the freeze-thaw procedure, thereby improving sperm quality.

Evaluating the Effects of Heavy Metals on Seminal Fluid Anti Oxidants Status and Semen Parameters among Males with Infertility at Tertiary Health Centre in Nigeria

Abstract Introduction: There are evidences of a reduction in male fertility potential globally. This, coupled with the limitation of the gold standard investigative tool of male infertility, has become necessary that focus should be on elucidating the aetiopathogenic causes of male fertility. This includes the impact of environmental toxic products such as levels of heavy metals that have been proposed to impact both the sperm quality and seminal antioxidant status. The study assessed how the seminal concentrations of specific heavy metals such as cadmium, lead and Iron affect the seminal fluid analysis (SFA) parameters and seminal antioxidant status of males exhibiting abnormal parameters of sperm, analysing infertile male patients consecutively in a cross-sectional manner, with at least one abnormal SFA parameter, at a tertiary health centre. Materials and Methods: One hundred and thirty consenting males who have one or several anomalies with sperm characteristics were selected as subjects. Semen samples were collected, processed and centrifuged. The supernatants were analysed for the heavy metals using Buck Scientific 210/211VGP Flame Atomic Absorption Spectrophotometers 220GF Graphite Furnace and 220AS Autosampler and seminal Vitamins C and E, using spectrophotometric method and glutathione peroxidase, whereas catalase was analysed using ELISA method. Results: Subjects’ seminal levels of Cd, Fe and Pb were notably greater than those of controls, respectively. In addition, individuals’ levels of seminal antioxidants were noticeably lower than controls. Seminal antioxidant status and Seminal Cd, Fe and Pb were positive and negative significantly correlated with sperm count and active motility, respectively. Conclusion: Laboratory quantification of the seminal concentrations of these heavy metals in the evaluation of males with infertility, especially those still regarded as idiopathic, because of limitations of investigative tools and biomarkers, will be of clinical utility, in both the management and prevention of sperm quality deterioration.

Global DNA methylation level in spermatozoa is not associated with ICSI fertilization outcome and embryo quality in donor oocyte programme

Epigenetic mechanisms, including DNA methylation and histone modifications, govern chromatin arrangement in sperm, enhancing motility and safeguarding DNA integrity for accurate epigenetic inheritance. Abnormal methylation is linked to poor sperm quality and fertility issues, underscoring the need to study sperm DNA methylation and its impact on sperm function and embryo development in assisted reproductive technology. In this study, processed spermatozoa from 75 normozoospermic and 15 abnormal ejaculates were examined for sperm global DNA methylation levels using a colourimetric absorbance method. Although semen characteristics were poor in abnormal ejaculates, no significant correlation was found between sperm global DNA methylation levels and sperm characteristics in either normozoospermic or abnormal cohorts. However, mean global DNA methylation levels were significantly lowered in abnormal sperm samples compared to normozoospermic samples (p < 0.05). Furthermore, injecting spermatozoa from these patients (N = 50) into donor oocytes did not show a significant relationship between sperm global DNA methylation and embryo developmental competence. These findings highlight the limitation of sperm global DNA methylation as a biomarker for embryo development and quality.

Enhancing of Rabbit Sperm Cryopreservation with Antioxidants Mito-Tempo and Berberine

Cryopreservation plays a critical role in animal breeding and the conservation of endangered species, but it often compromises sperm characteristics such as morphology, motility, and viability due to oxidative stress. This study explores the antioxidative effect of Mito-Tempo (MT) and Berberine (BER) to enhance post-thaw sperm quality in rabbits. Pooled rabbit sperm samples were supplemented with different concentrations (0.0, 0.5, 5, 10, 50 µmol/L) of MT and BER. Sperm motility was evaluated using computer-assisted semen analysis, while viability, apoptosis, reactive oxygen species (ROS) levels, acrosome integrity, and mitochondrial function were assessed through flow cytometry. The results revealed that MT at 5 and 10 µmol/L and BER at 10 µmol/L significantly improved total and progressive motility, mitochondrial activity, and sperm viability compared to the control group. Furthermore, 10 µmol/L BER enhanced acrosome integrity, while both 5 µmol/L MT and 10 µmol/L BER effectively reduced ROS levels and apoptosis. This study is the first to demonstrate the protective effects of MT and BER on rabbit sperm during cryopreservation. By mitigating oxidative stress and reducing apoptosis, these antioxidants markedly improved post-thaw sperm quality, positioning MT and BER as promising agents for improving sperm cryosurvival.

Crude Garden Cress Seed Oil (Lepidium sativum Linn.) Enhances Post-Thawed Boar Sperm Quality

This study aimed to examine the effects of crude garden cress seed oil (CGCSO) on frozen–thawed boar sperm qualities. Semen ejaculates (n = 12) were collected and further divided into six equal aliquots based on CGCSO concentrations (0, 0.5, 1, 1.5, 2, and 2.5% v/v) in the freezing extender. Semen samples were processed and cryopreserved utilizing the traditional liquid nitrogen vapor technique. Subsequently, semen samples were thawed in a thermos with warm water at 50 °C for 12 s and evaluated for sperm morphology using scanning electron microscopy, sperm motility using a CASA, sperm viability, acrosome integrity, mitochondrial function, MDA level, total antioxidant capacity (TAC), glutathione peroxidase (GSH-Px), and catalase (CAT) activity. The results indicated that 1% CGCSO resulted in superior post-thaw sperm characteristics, including enhanced sperm morphology, motility, viability, acrosome integrity, and mitochondrial function. Particularly, the total motile sperm increased by 16.5%, progressive motile sperm increased by 13.0%, viability improved by 15.1%, acrosome integrity increased by 14%, and mitochondrial function improved by 14.1% compared to the control group. CGCSO treatment at 1% and 1.5% exhibited the lowest level of MDA (45.73 ± 11.2 and 45.73 ± 11.3 µmol/L, respectively) compared to the other groups. The CGCSO-supplemented groups showed higher values of TAC, GSH-Px, and CAT than the control group but not significantly.

Mammalian Ste-20-like Kinase 1/2 (MST1/2) Inhibitor XMU-MP-1: A Potential Compound to Improve Spermatogenesis in Mouse Model of Diabetes Mellitus

Background/Objectives: Spermatogenesis is a key process in male reproduction that, if it does not happen correctly, can lead to infertility, with diabetes being one of the most prevalent causes of spermatogenesis disruption. Currently, there is a lack of research examining the potential benefits of targeting cell proliferation to enhance spermatogenesis in this condition. XMU-MP1 has been identified as an inhibitor of MST1, a core component of the Hippo pathway, which is anticipated to promote proliferation and regeneration. This study aims to evaluate the effects of XMU-MP1 treatment on sperm and testicular characteristics in mice. Methods: We used the STZ-induced diabetic mouse model to investigate the impact of administering XMU-MP1 on testicular tissue and sperm parameters. This study compared the seminiferous tubules, specifically focusing on the diameter of the seminiferous tubule, the thickness of the seminiferous tubule epithelium, the ratio of the thickness of the seminiferous tubule epithelium to the diameter of the seminiferous tubules, and the lumen diameter of the seminiferous tubules. We also conducted a comparison of sperm parameters, including sperm concentration, progressive motility, total motility, total motility, and morphology. Results: XMU-MP1-treated mice had a larger spermatogenesis area and better sperm motility than control mice. Diabetic mice treated with XMU-MP1 also showed a trend toward improvements in the spermatogenesis area, sperm concentration, sperm motility, and sperm morphology, although these improvements were not statistically significant. Conclusions: XMU-MP1 serves as a potential compound to improve spermatogenesis in mice.

Protective effects of Sphaeranthus indicus floral extract against BPS-induced testicular damage in rats occurs through downregulation of RIPK1/3-MLK-driven necroptosis and Fas-FasL-mediated apoptosis

AbstractBisphenol S (BPS) is one of the monomers preferred in the manufacturing of polycarbonate plastics. Unfortunately, its estrogenic and genotoxic effects are similar to those of bisphenol A. The protective effects of Sphaeranthus indicus floral extract (SFE) against reprotoxic effects of BPS (50 µg/kg per body weight) in rats exposed to it via drinking water was investigated. Different SFE doses (25, 50, and 100 mg/kg) were administered via oral gavage for 10 weeks. High-performance liquid chromatography (HPLC) results indicated that SFE was rich in polyphenols, with rutin and quercetin being important bioactive molecules modulating BPS-induced necroptosis and apoptosis. Biochemical analyses unveiled that rats administered BPS only exhibited considerable elevation of biomarkers of nitro-oxidative stress, necroptotic (RIPK1/RIPK3 and MLKL), and apoptotic mediators (Fas/FasL and caspase 3/caspase-8). These events caused changes in sperm characteristics (sperm motility, sperm head, and sperm viability), sperm count, and hormonal profile (thyroid stimulating hormone, luteinizing hormone, and follicle-stimulating hormone) of the rats. Histological analysis suggested that there was pronounced sloughing of Sertoli cells, reduced spermatogenic cell density, increased levels of seminiferous tubules, and disorganized morphometric parameters related to seminiferous tubules. The SFE supplementation in rats with BPS-containing water restored nitro-oxidative stress biomarkers, which led to the reduction of necroptosis and apoptosis. Reinstatement of all the biomarkers of oxidative stress, inflammation, necroptosis, and apoptosis after SFE supplementations restored the hormonal profile and normal histoarchitecture of the testes. Virtual screening elucidated that the key regulators of the necroptosis are RIPK3-rutin and RIPK1-quercetin complexes. Further studies are needed to assess its pharmacodynamics, kinetics, and effective concentration for daily use in humans.

Novel bi-allelic DNAH3 variants cause oligoasthenoteratozoospermia

BackgroundOligoasthenoteratozoospermia (OAT) is a widespread cause of male infertility. One of the usual clinical manifestations of OAT is multiple morphological abnormalities of the sperm flagella (MMAF), which are frequently associated with mutations and defects in the dynein family. However, the relationship between the newly identified Dynein Axonemal Heavy Chain 3 (DNAH3) mutation and oligonasthenospermia in humans has not yet been established.MethodsWhole exome sequencing, pathogenicity analysis, and species conservation analysis of mutation sites were conducted on two patients from different unrelated families with DNAH3 mutations. We identified representative mutation sites and predicted the protein structure following these mutations. The sperm characteristics of the two patients with DNAH3 mutations were verified using Papanicolaou staining, scanning electron microscopy, and transmission electron microscopy. Additionally, mRNA and protein levels were assessed through RT-qPCR and Western blotting.ResultsThe biallelic mutations in the first progenitor included a heterozygous deletion and insertion, c.6535_6536 delinsAC (to infect mutation (p.Asp2179Thr), and stop codon premutation, c.3249G &amp;gt; A (p.Trp1083Ter). In Family II, the patient (P2) harbored a DNAH3 heterozygous missense mutation, c. 10439G&amp;gt; A(p.Arg3480Gln), along with a stop codon premutation, (c.10260G &amp;gt; A; p.Trp3420Ter). Patients with premature termination of transcription or translation due to DNAH3 mutations exhibit OAT phenotypes, including fibrous sheath dysplasia and multiple tail malformations. We identified the representative sites after mutation, predicted the protein structure, and assessed changes in the protein levels of DNAH3 and related genes following mutations. Notably,a significant reduction in DNAH3 protein expression was validated in these patients. We may explore in the future how DNAH3 affects sperm motility and quality through regulatory mechanisms involving protein structural changes.ConclusionNovel biallelic mutations in DNAH3, especially those resulting in a premature stop codon, may alter protein expression, structure, and active site, leading to spermatogenic failure and potentially inducing OAT. The discovery of new mutations in DNAH3 may be the key to the diagnosis and treatment of OAT.

Human papillomavirus carriage in the semen of men consulting for infertility: prevalence and correlations with sperm characteristics.

We aim to study the semen carriage of human papillomavirus (HPV) and evaluate its association with patient characteristics. We conduct a single-center cohort study at Amiens University Hospital Center (Amiens, France). From May 1 to October 31, 2021, 461 men consulting for infertility and with semen analysis data were included. Each participant gave his written informed consent for the use of laboratory, demographic, clinical, and lifestyle data. A proportion of the semen samples were sent to a virology laboratory for HPV screening in a polymerase chain reaction (PCR) assay. In univariate and multivariate analyses with a logistic regression model, HPV+ and HPV- groups were compared with regard to semen characteristics (including the DNA fragmentation index and the sperm decondensation index) and demographic, clinical, and lifestyle variables. Semen HPV carriage was detected in 22.3% of the patients. High-oncogenic-risk HPV genotypes were predominant (57.6%). Multivariate analysis showed that HPV carriage was significantly associated with the presence of at least one abnormal spermogram dinging (according to the 6th World Health Organization criteria), with an adjusted odds ratio (OR) of 4.10 (95% confidence interval [CI]: 2.32-7.25, P < 0.001). A statistically significant association was also found for the type of infertility (OR: 1.61, 95% CI: 1.00-2.57, P = 0.05), the presence of varicocele (OR: 3.99, 95% CI: 1.48-10.71, P = 0.01), and a history of cryptorchidism, testicular ectopia, or monorchidism (OR: 3.54, 95% CI: 1.07-11.66, P = 0.04). Infection with a single HPV genotype or multiple HPV genotypes was significantly associated with at least one abnormal spermogram finding for all HPV oncogenic risk groups (OR: 3.93, 95% CI: 2.08-7.41, P < 0.001; and OR: 4.11, 95% CI: 1.58-10.68, P = 0.01, respectively). The association between sperm HPV carriage and the risk of infertility was statistically significant in a multivariate analysis (OR: 5.63, 95% CI: 3.16-10.01, P < 0.001) and after adjustment for the propensity score (OR: 6.10, 95% CI: 3.33-11.21, P < 0.001). Our results suggest that semen HPV carriage has an impact on male fertility. Sperm screening for HPV might be a useful addition to the work-up for male infertility.

The Effect of Cyperus esculentus on Sperm Function Parameters in Prepubertal Mice as a Model for Human

The objective of this work was to study the effect of oral administration of Cyperus esculentus (CE) and its alcoholic extract on sperm function parameters in prepubertal mice as a model for human .The animals were divided into three groups each contains 6 animals .Group 1 was treated with 150 mg/ kg body weight /day of crude CE, group 2 was treated with same dose of alcohol extract of CE and group 3 regarded as control throughout six weeks period. The results showed a significant (p&gt; 0.05) increase in the mean of sperm concentration ,sperm motility percent and progressive sperm motility between treated groups and control . There was no differences among groups in the mean of sperm normal morphology and sperm viability . No significant differences was recorded in the mean of body weight among groups throughout the study. The results revealed that the administration of Cyperus esculentus may enhance certain sperm characters in prepubertal mice without affecting body weight.

Correlation of Seminal Plasma EMMPRIN Levels with Sperm Quality and the Efficacy of Activation Techniques

Background: Infertility is a major global health challenge, impacting 15% to 20% of couples globally. Male infertility, which is typically associated with issues such as low sperm motility, has increased during the last two decades. Male fertility relies heavily on the development of extracellular matrix metalloproteinase inducer (EMMPRIN), a glycosylated transmembrane protein involved in sperm function and embryo implantation. Understanding how sperm preparation techniques affect sperm quality and EMMPRIN expression is crucial for achieving optimal results in assisted reproductive technologies (ART). Objective: To investigate EMMPRIN expression in human sperm and compare the impact of two in vitro sperm preparation procedures, “direct swim-up (DSU) and pellet swim-up (PSU), on various sperm characteristics. To determine the optimum technique for improving sperm quality in ART. Materials and Methods: Masturbation was employed to collect sperm samples from 44 participants (aged 21 to 45). Following liquefaction, samples were evaluated macroscopically and microscopically in accordance with WHO 2021 guidelines. The samples were separated into three aliquots for ELISA assessment of EMMPRIN levels and sperm processing with DSU and PSU techniques. Sperm chromatin immaturity (SCI) was assessed using Aniline Blue staining. Statistical investigations were carried out to determine the correlations between EMMPRIN levels, semen parameters, and the impacts of the two in vitro sperm preparation techniques. Results: Sperm concentration significantly decreased from control (40.25 ± 2.30) to DSU (15.51 ± 1.74) and PSU (9.99 ± 1.16) conditions. However, motility, morphology, and SCI significantly improved, particularly with PSU, which showed a greater reduction in SCI % compared to DSU, indicating its effectiveness in selecting higher-quality sperm. There were no significant associations detected between EMMPRIN levels with semen metrics, although PSU exhibited a slightly higher association with improved sperm parameters. Conclusion: Seminal plasma EMMPRIN levels do not appear to directly influence semen parameters or sperm chromatin maturity. However, the choice of sperm activation technique, particularly PSU, significantly impacts sperm quality. More research is needed to better understand the importance of EMMPRIN in ART outcomes and its potential as a sperm quality indicator.

Pumpkin seed oil lessens the colchicine-induced altered sex male hormone balance, testicular oxidative status, sperm abnormalities, and collagen deposition in male rats via Caspase3/Desmin/PCNA modulation

This study examined the efficiency of pumpkin seed oil (PSO) to rescue the colchicine (CHC)-induced adverse impacts on sperm characteristics, male sex hormones, testicular architecture, oxidative status, DNA content, collagen deposition, and immune expression of desmin and PCNA. Male Sprague Dawley rats were divided into four experimental groups (n = 10 each): control (distilled water), CHC (0.6 mg/kg b.wt), PSO (4 mL/kg b.wt), and CHC + PSO. After 60 days of dosing, CHC significantly reduced sperm motility (19%), sperm concentration (38%), estradiol (52%), testosterone (37%), luteinizing hormone (54%), and follicle-stimulating hormone (29%) compared to the control. Yet, the testicular tissues of CHC-administered rats exhibited elevated abnormal sperms (156%), malondialdehyde (354%), lactate dehydrogenase (73%), Caspase-3 (66%), and 8-hydroxyguanosine (65%) but lower reduced glutathione (74%), catalase (73%), and superoxide dismutase (78%) compared to the control group. Moreover, CHC induced testicular degeneration, distorted seminiferous tubules, apoptotic cells, exfoliated spermatogenic cells, reduced DNA content, decreased PCNA and desmin immune-expression, and increased collagen deposition. PSO effectively reversed the CHC-induced alterations in sperm quality and testicular function and architecture, likely through its antioxidant, antifibrotic, anti-apoptotic, and DNA-protective properties. These results suggest that PSO may be a beneficial intervention for long-term CHC users and may protect against CHC-induced male reproductive toxicity.

Ultrastructural Characteristics of the Mature Spermatozoon of Artyfechinostomum malayanum (Digenea: Echinostomatidae), an Intestinal Parasite of Rattus norvegicus (Rodentia: Muridae) in Vietnam.

The study of sperm characteristics has proven useful for elucidating interrelationships in several groups of Platyhelminthes, such as digeneans. Thus, in the present work, the ultrastructural organization of the mature spermatozoon of the digenean Artyfechinostomum malayanum (Echinostomatidae), a parasite of Rattus norvegicus (Rodentia: Muridae) from Dong Thap Province, Vietnam, was investigated for the first time using transmission electron microscopy. The male gamete of A. malayanum exhibits two axonemes of different lengths, showing the 9 + '1' pattern of the Trepaxonemata, a nucleus, two mitochondria, two lateral expansions, two bundles of parallel cortical microtubules, external ornamentation, spine-like bodies, and granules of glycogen. Thus, the mature spermatozoon follows a Type V sperm model proposed for digeneans. We also highlight some noteworthy characteristics in Echinostomatidae with possible phylogenetic implications, such as two lateral expansions in the anterior region of the spermatozoon and two mitochondria.

Effect of Incorporating Soursop (Annona muricata) Leaves Powder on Reproductive Performance of Japanese Quail (Coturnix coturnix japonica)

Introduction: Soursop leaves are rich in various molecules, including total phenols, terpenes, and steroids, which possess a range of pharmacological properties that can be utilized in animal production to enhance both growth and reproduction of animals. The present study aimed to evaluate the effect of incorporating soursop (Annona muricata; A. muricata) leaves powder into feed on the reproductive performance of Japanese quail. Materials and methods: A total of 80 Japanese quails (64 females and 16 males) aged two weeks were randomly divided into four experimental groups, labeled T0, T1, T2, and T3, and received feed additives with 0 mg/kg body weight (bw), 250 mg/kg bw, 500 mg/kg bw, and 750 mg/kg bw of powdered soursop leaves, respectively. Additionally, water was provided ad libitum, and the quails' weights were measured every 7 days for a period of 75 days. At the end of the period, 12 female quails from each group were sacrificed after 24 hours fasting period. Blood was collected for hematological (Leukocyte, Erythrocyte, and Platelet indices) and serum biochemical (total serum cholesterol, total proteins, albumin, and globulin, Aspartate aminotransferase, Alanine aminotransferase, Urea, and Creatinine) analysis. The males were also sacrificed to evaluate the spermatozoa characteristics (mobility, concentration, and viability). Results: No statistical significant changes in growth characteristics or hematological parameters were observed. However, biochemical parameters increased significantly with the inclusion of Soursop (A. muricate) in quail feed, including increased total cholesterol, total protein, and globulin levels, and decreased malondialdehyde levels. This effect was most significant at dosage of 500 mg/kg bw. Serum levels of urea, Alanine aminotransferase, and Aspartate aminotransferase were not significantly affected by A. muricata whatever the concentration considered. A significant increase in fast progressive spermatozoa, along with a decrease in immotile spermatozoa, was observed with A. muricate at dosage of 500 mg/kg bw compared to the control. Sperm viability also increased significantly, particularly in live at dosage of 500 mg/kg bw. Significant increase was observed in fertility parameters, including increased fertility rate, hatchability rate of fertile eggs, total hatchability rate, and chick weight, along with decreased embryonic mortality at 500 mg/kg bw treatment compared to the control. Conclusion: In conclusion, the findings indicated that incorporating A. muricata leaf powder at 500 mg/kg bw into quail feed positively influences reproductive cells and boosts fertility growth promoters.

The Effect of Metallic Nanoparticles Supplementation in Semen Extender on Post-thaw Quality and Fertilizing Ability of Egyptian Buffalo (Bubalus bubalis) Spermatozoa.

Nanomaterials offer several promising prospects in the field of farm animal reproduction, encompassing a broad range of applications such as transgenesis and the precise delivery of substances to sperm cells, antimicrobial, antioxidants properties as well as their potent role in improving cryopreservation methods. The aim of the present study is to explore the effect of supplementing the semen extender with 10µg/mL nano gold (Au-NPs10), 10µg/mL nano silver (Ag-NPs10), 1µg/mL nano selenium (Se-NPs1), and 100µg/mL nano zinc oxide (ZnO-NPs100) on sperm characteristics and kinematics parameters, acrosome integrity, oxidative biomarkers, morphological and apoptosis-like changes of frozen-thawed buffalo bull sperm, and, ultimately, their fertilizing capacity. The results revealed that all aforementioned nano materials significantly improved viability, progressive motility, membrane integrity, acrosome integrity, and kinematic parameters as well as apoptosis-like changes of post-thawed buffalo bull sperm compared to the control (p < 0.05). No discernible effects were observed on sperm ultrastructure morphology measures as a response to the addition of these metallic nanoparticles to the extender. The values of caspase 3 significantly decreased by 64.22, 45.99, 75.59, and 49.39% in Au-NPs10, Ag-NPs10, Se-NPs1, and ZnO-NPs100treated groups, respectively, compared to the control. The addition of 100µg ZnO-NPs to the extender significantly decreased the total count of bacteria, fungi, and yeast compared to the control (p < 0.05). The AuNPs10 and SeNPs1 treated groups showed lower content of malondialdehyde, hydrogen peroxide, and nitric oxide concentrations and higher values of total antioxidant capacity of post-thawed extended semen (p < 0.05). Pregnancy rates increased by 17.5, 20, and 30% in buffalo cows inseminated with sperm treated with ZnO-NPs100, Se-NPs1, and Au-NPs10, respectively, compared to the control group. The present results indicate that the freezing extender supplemented with metallic nanoparticles can be an effective strategy to enhance the cryotolerance and fertility potential of buffalo bull sperm.

How do taurine and ergothioneine additives improve the parameters of high- and low-quality turkey semen during liquid preservation?

Abstract The study aimed to examine the parameters of turkey ejaculates (n=40) of high and low quality (HQ and LQ, respectively), preserved with the addition of taurine (TAU; 5 mM, 10 mM) and ergothioneine (EGT; 5 mM, 10 mM) for 48 h at 5°C in a liquid state. The motility, plasma membrane integrity (PMI), mitochondrial function, apoptotic and necrotic spermatozoa percentage, and sperm cells that generated NO were determined after 2, 24, and 48 h of storage. The preserved semen was also analysed for the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), glutathione (GSH) content, and malondialdehyde (MDA) production. Taurine, in both doses, may improve the antioxidant status of stored turkey semen as well as sperm motility, viability and functionality regardless of ejaculate quality, as manifested by increased SOD and CAT activities, reduced MDA levels, and enhanced sperm characteristics i.e. plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), total motility (TMOT), and progressive motility (PMOT). In turn, the addition of EGT increased GSH content in the external environment and suppressed lipid peroxidation in turkey spermatozoa, in particular those from low-quality (LQ) ejaculates. However, it appears that EGT could deliver more satisfactory results when added at lower concentrations.

Long-term cryopreservation of bull semen as a main strategy for the ex situ conservation of endangered Polish Red cattle

Abstract Regular verification of the quality of cryopreserved semen derived from native cattle is one of the tasks performed at the bank as recommended by the Food and Agriculture Organization. The purpose of the present study was to evaluate the quality of semen from PR bulls stored for 40, 50 and 60 years in the BMB using standard evaluation parameters such as sperm motility as well as structural-functional parameters such as plasma membrane integrity, transmembrane mitochondrial potential and sperm chromatin damage. Semen pellets from 27 PR bulls (3 ejaculates/bull) were tested. The data were analysed by one-way and two-way ANOVA, and the significance of the difference (P≤ 0.01) between the means was determined using Duncan’s test. Our study results revealed that the long-term storage of semen had no effect on sperm characteristics after thawing. However, statistically significant differences (P≤0.01) in sperm plasma membrane integrity and transmembrane mitochondrial potential, following storage in liquid nitrogen were noted between bulls at all time points. However, there were no significant differences (P&gt;0.01) in sperm chromatin damage between breeds or between different storage times, and the degree of DNA fragmentation ranged from 0.4 to 0.8%.

Enhancing evaluation of bull fertility through multivariate analysis of sperm

Computer-assisted sperm analysis (CASA) has become the predominant tool for assessing bull semen in artificial insemination programs. Despite such popularity CASA's ability to predict fertility has been limited, especially when emphasis is based upon single motion characteristics. Our hypothesis is that numerical sets of CASA measures provide a more effective method to differentiate the potential fertilization capacity of bulls and that bulls can be clustered based upon sets of CASA measures. Therefore, we used CASA to evaluate frozen-thawed semen samples from 307 Holstein and 152 Jersey bulls sourced from USDA-ARS's National Animal Germplasm Program gene bank. Sperm was evaluated immediately after thawing and 30 min later. We evaluated sperm kinetic and morphometric means and variances to capture the structure of CASA data in relation to various sources of variation. These data were subjected to univariate and multivariate statistical methods to investigate animal and management factors affecting sperm characteristics measured by CASA. Clustering with K-means identified 4 clusters of bulls based upon each cluster's set of CASA parameters after thawing. There was little overlap among clusters for sets of CASA measures. At the extremes, bull cluster 1 (BC1, n = 180) and BC3 (n = 101) had different sire conception rates (SCR) -0.07 vs -1.29, respectively and sets of CASA measures. Interestingly, BC2 had CASA measures that could be perceived as negative, e.g., cell size at 8.18mm2 vs 6.37mm2 for BC4 and total motility of 29.7% vs 48.7% for BC3, but SCR for BC2 was higher (-0.79) than BC3 (-1.29). Despite such discrepancies for some BC2 CASA values it appears the potentially negative effects were offset by the levels of other CASA values. Our findings suggest improved approaches for using CASA could lie in evaluating multiple CASA measures as sets within specific numerical ranges rather than as independent measures.

The quality and characteristics of bovine sperm are compromised by Toxoplasma gondii antigens, impacting in in vitro bull fertility

Studies in various species have demonstrated different results on the effects of T. gondii infection on sperm quality. It has also been demonstrated that in some stages of the disease, there is elimination of cellular debris or even the intact parasite in the semen. The present work aimed to evaluate the in vitro effects of the presence of soluble T. gondii antigens in bovine semen on sperm integrity. The spermatozoa were treated with T. gondii antigens in double serial dilutions classified as high, medium and low doses (8, 4, 2 µg/ml) in "TALP-Sperm” and “TALP-Fert" media. The results showed that T. gondii antigens affect sperm motility and mitochondrial activity, and cause changes in sperm chromatin integrity, as well as damage to the sperm membrane and acrosome. Finally, spermatozoa treated with T. gondii antigens were evaluated in the in vitro production of embryos (IVEP). The use of semen contaminated with antigens in IVEP routines did not lead to a decrease in the fertilization of oocytes, as sperm undergo selection before in vitro fertilization, which eliminates the most altered sperm. However, early embryonic development was affected, probably by structural changes that were not eliminated in the selection process. The results demonstrated that the presence of soluble T. gondii antigens in bovine semen alters sperm integrity and vital characteristics for the fertilization process and embryonic development and therefore causes fertility problems in males.

Effect of Sperm Extender and Dilution Ratio on The Sperm Motility, Fertility, and Hatching Rates of Depik Fish Rasbora Tawarensis (Pisces: Cyprinidae) Eggs

Abstract There is no known artificial breeding method for depik fish Rasbora tawarensis. Sperm extender and the dilution ratio are crucial factors in the artificial breeding of fish. The current study aimed to assess the effect of several common fish sperm extenders and its dilution ratio on sperm characteristics and fertilization of depik fish eggs. Two series of experiments were performed to find the best extender and its dilution ratio. In the first experiment, five types of extenders were tested: Ringer’s solution, physiological solution, coconut water, sugarcane water, and roomie water. The second experiment studied four dilution levels of sperm and extender (v/v): 1:20, 1:30, 1:40, and 1:50. Every treatment was conducted in four replicates and the experiments were conducted consequently so that the findings in the first experiment were applied in the second. The first experiment showed that Ringer’s is the best extender for depik sperm with sperm motility, fertilization and hatching rates of 71.00%, 69.30%, and 53.66%, respectively; and the second experiment revealed that Ringer’s at dilution ratio 1:40 gave higher sperm motility, fertilization, and hatching rates of 74.66, 70.33, and 59.00%, respectively. In conclusion, Ringer’s solution was the most favorable extender for sperm which yielded best fertilization results at 1:40 dilution ratio (sperm:extender, v/v) in depik fish.