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There has recently been an explosion of studies implicating the human microbiome in playing a critical role in many disease and wellness states. The etiology of abnormal semen analysis (SA) parameters is not identified in 30% of cases; investigations involving the semen microbiome may bridge this gap. Here, we explore the relationship between the semen microbiome and alterations of sperm parameters. We recruited men presenting for fertility evaluation or vasectomy consultation with proven biological paternity. SA and next generation sequencing was performed. Differential abundance testing using Analysis of composition of Microbiota with Bias Correction (ANCOM-BC) was performed along with canonical correlational analysis for microbial community profiling. Men with abnormal (N = 27) sperm motility showed a higher abundance of Lactobacillus iners compared to those with normal (N = 46) sperm motility (mean proportion 9.4% versus 2.6%, p = 0.046). This relationship persisted on canonical correlational analysis (r = 0.392, p = 0.011). Men with abnormal sperm concentration (N = 20) showed a higher abundance of Pseudomonas stutzeri (2.1% versus 1.0%, p = 0.024) and Pseudomonas fluorescens (0.9% versus 0.7%, p = 0.010), but a lower abundance of Pseudomonas putida (0.5% versus 0.8%, p = 0.020), compared to those with normal sperm concentration (N = 53). Major limitations are related to study design (cross-sectional, observational). Our results suggest that a small group of microorganisms may play a critical role in observed perturbations of SA parameters. Some of these microbes, most notably Lactobacillus iners, have been described extensively within other, fertility-related, contexts, whereas for others, this is the first report where they have potentially been implicated. Advances in our understanding of the semen microbiome may contribute to potentially new therapeutic avenues for correcting impairments in sperm parameters and improving male fertility.

Fertilization occurs before the completion of oocyte meiosis in the majority of animal species and sperm contents move long distances within the zygotes of mouse and C. elegans. If incorporated into the meiotic spindle, paternal chromosomes could be expelled into a polar body resulting in lethal monosomy. Through live imaging of fertilization in C. elegans, we found that the microtubule disassembling enzymes, katanin and kinesin-13 limit long-range movement of sperm contents and that maternal ataxin-2 maintains paternal DNA and paternal mitochondria as a cohesive unit that moves together. Depletion of katanin or double depletion of kinesin-13 and ataxin-2 resulted in the capture of the sperm contents by the meiotic spindle. Thus limiting movement of sperm contents and maintaining cohesion of sperm contents within the zygote both contribute to preventing premature interaction between maternal and paternal genomes.

Despite rapid evolution across eutherian mammals, the X-linked MIR-506 family miRNAs are located in a region flanked by two highly conserved protein-coding genes (SLITRK2 and FMR1) on the X chromosome. Intriguingly, these miRNAs are predominantly expressed in the testis, suggesting a potential role in spermatogenesis and male fertility. Here, we report that the X-linked MIR-506 family miRNAs were derived from the MER91C DNA transposons. Selective inactivation of individual miRNAs or clusters caused no discernible defects, but simultaneous ablation of five clusters containing 19 members of the MIR-506 family led to reduced male fertility in mice. Despite normal sperm counts, motility, and morphology, the KO sperm were less competitive than wild-type sperm when subjected to a polyandrous mating scheme. Transcriptomic and bioinformatic analyses revealed that these X-linked MIR-506 family miRNAs, in addition to targeting a set of conserved genes, have more targets that are critical for spermatogenesis and embryonic development during evolution. Our data suggest that the MIR-506 family miRNAs function to enhance sperm competitiveness and reproductive fitness of the male by finetuning gene expression during spermatogenesis.

Context: Prostate-specific antigen (PSA) is important for semen liquefaction and sperm motility. Anti-PSA antibodies may lead to immune infertility. Aims: The present study was conducted to evaluate the impact of seminal anti-PSA antibodies on semen parameters in fertile and infertile men. Methods: A cross-sectional analytic study was conducted on 105 fertile men (≥21–50 years) having biological children (within last 2 years) with normal semen analysis as controls and 105 infertile men with abnormal semen analysis as cases. All semen samples were cryopreserved till 210 samples were collected, followed by estimation of anti-PSA antibodies using an indirect enzyme-linked immunosorbent assay technique. Key results: Mean ± standard deviation (s.d.) age of 210 participants was 30.0 ± 4.65 years. Mean ± s.d. levels of seminal anti-PSA antibodies in infertile men were 27.82 ± 102.19 ng/mL and in fertile men −30.45 ± 49.49 ng/mL (P = 0.001). A significant negative correlation was observed between anti-PSA antibody levels and sperm concentration (P = 0.013), rapid progressive motility (P = 0.001), slow progressive motility (P = 0.006), progressive sperm motility (P = 0.001), and normal morphology (P = 0.001), and significant positive correlation was observed with immotile sperms (P = 0.001). The overall accuracy of anti-PSA antibody for differentiating infertile from fertile men was 63.33%. Conclusions: Seminal anti-PSA antibodies were significantly correlated with semen parameters in fertile and infertile men with an accuracy of 63.33%. A negative correlation was observed between antibody levels and progressive sperm motility. Implications: Seminal anti-PSA antibodies can be used as a biomarker for male infertility assessment. Globally, male factor infertility is increasing and so is the need for newer and effective tests for its diagnosis and management. The present study was conducted to evaluate the impact of seminal anti-prostate-specific antibodies (PSA) antibodies on semen parameters in fertile and infertile men. A significant negative correlation was observed with sperm concentration, motility, and morphology, and a positive correlation was observed with grade D motility. The overall accuracy of anti-PSA antibodies in predicting infertile from fertile men was 63.33%. Image by Naina Kumar. [ABSTRACT FROM AUTHOR]

Cryopreservation is a pivotal technique in safeguarding genetic material across diverse species, despite its inherent challenges linked to induced spermatozoa damage, notably apoptosis and lipid peroxidation (LPO). Given the insufficient antioxidant defense of spermatozoa against LPO, there is a rising interest in integrating additional additives into extenders to ameliorate mammalian semen quality. Among these additives, flavonoids have garnered considerable attention due to their potent antioxidative properties. Hence, our study aimed to assess the efficacy of flavone (FL) and 3-hydroxyflavone (3-OH =) supplementation in the cryopreservation medium to protect canine sperm against the damaging impacts of freezing and ensure the preservation of their reproductive potential. Semen was collected from five Beagle stud dogs and then pooled. Then, the sample was divided into 7 groups, each treated with 1) 0 mM, 2) 0.1 mM FL, 3) 0.2 mM FL, 4) 0.4 mM FL, 5) 0.1 mM 3-OH = , 6) 0.2 mM 3-OH = , 7) 0.4 mM 3-OH =. Semen samples were subjected to cryopreservation in French straws and glycerol as a cryoprotectant. In the frozen thawed semen, sperm motility parameters by CASA system and sperm membrane integrity, acrosome status, mitochondrial activity, DNA fragmentation, early apoptosis with capacitation, and LPO were assessed using flow cytometry just after thawing (0 h) and 4 h post thaw. Results reveal significant increase in the proportion of live spermatozoa with undamaged acrosomes in the FL 0.1 and 3-OH = 0.2 groups at 0 h post thaw. At this time point, 3-OH = 0.1 significantly reduced the DNA fragmentation index (DFI) compared to the FL 0.1 and 0.2 groups. However, after the next 4 h, 3-OH = 0.4 exhibited the lowest (P < 0.05) DFI compared to FL 0.2 and 3-OH = 0.1. Additionally, 3-OH = 0.4 showed the highest (P < 0.05) proportion of non apoptotic and non capacitated spermatozoa compared to FL 0.1 0 h post-thaw. Simultaneously, the same group demonstrated significant reduction in apoptotic and capacitated sperm cells, at 0 h and 4 h post-thaw. Moreover, 3-OH = at 0.1 (0 h and 4 h) and 0.2 mM (4 h) significantly enhances the proportion of live sperm without LPO post thaw. Whitin the FL groups, only 0.4 FL significantly increased the percentage of live sperm without LPO. No significant effect of the tested substances was observed on sperm motility, cell membrane integrity, or mitochondrial activity. These findings highlight the promising role of flavone and 3-hydroxyflavone in enhancing sperm resilience during cryopreservation, suggesting their protective function against acrosome damages, capacitation, apoptosis and lipid peroxidation. [Display omitted] • lFlavone (FL) and 3-hydroxyflavone (3-OH =) were used to supplement the extender for canine semen cryopreservation for the first time. • Using FL 0.1 mM and 3-OH = 0.2 mM increased (P < 0.05) the proportion of live spermatozoa with undamaged acrosomes. • Using 3-OH = 0.1 and 0.4 mM significantly reduced the DNA fragmentation index of spermatozoa. • Using 3-OH = 0.4 mM significantly reduced apoptosis and capacitation of sperm cells. • Using FL 0.4 mM and 3-OH = 0.1 and 0.2 mM significantly protected the sperm against lipid peroxidation. [ABSTRACT FROM AUTHOR]

Dielectric barrier discharge (DBD) plasma regulates the levels of reactive oxygen species (ROS), which are critical for sperm quality. MicroRNAs (miRNAs) are non-coding single-stranded RNA molecules encoded by endogenous genes, which regulate post-transcriptional gene expression in animals. At present, it is unknown whether DBD plasma can regulate sperm ROS levels through miRNAs. To further understand the regulatory mechanism of DBD plasma on sperm ROS levels, miRNAs in fresh boar spermatozoa were detected using Illumina deep sequencing technology. We found that 25 known miRNAs and 50 novel miRNAs were significantly upregulated, and 14 known miRNAs and 74 novel miRNAs were significantly downregulated in DBD plasma-treated spermatozoa. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that target genes of differentially expressed miRNAs were involved in many activities and pathways associated with antioxidants. We verified that DBD plasma significantly increased boar sperm quality and reduced ROS levels. These results suggest that DBD plasma can improve sperm quality by regulating ROS levels via miRNAs. Our findings provide a potential strategy to improve sperm quality through miRNA-targeted regulation of ROS, which helps to increase male reproduction and protect cryopreserved semen in clinical practice. • DBD plasma improves boar sperm quality by regulating ROS levels. • DBD plasma affects sperm ROS levels via differentially expressed miRNAs. • DBD plasma regulates sperm ROS levels via miRNA-targeted antioxidant enzyme genes. • DBD plasma regulates sperm ROS levels via miRNAs targeting the KEAP1/NRF2 pathway. [ABSTRACT FROM AUTHOR]

Melatonin (MLT) has strong antioxidant capacity and can reduce the damage caused by oxidative stress in sperm, but there is still little content in the field we have studied. In this study, we are committed to scientific research on adding melatonin to Belgian blue bull semen diluent for cryopreservation. Different concentrations (0, 0.1, 0.3, 0.5 or 0.7 mg/mL) of MLT were added diluent. Sperm kinetic parameters, enzyme activity, antioxidant gene expression and fertility were analyzed after thawing. The results showed that MLT concentration of 0.3 mg/mL exerted positive effects on post-thaw kinetic parameters. Compared with other groups, 0.3 mg/mL MLT treated sperm acrosome and plasma membrane integrity, catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels significantly increased. Meanwhile, the mRNA expression of antioxidant genes SOD2 , CAT and GPx increased in the 0.3 mg/mL MLT treatment group, and the mRNA expression of apoptosis genes Caspase-3 and Bax were significantly reduced. In addition, in vitro fertilization (IVF) embryo cleavage, blastocyst rate and artificial insemination (AI) pregnancy rate were higher in 0.3 mg/mL MLT. Therefore, MLT showed cryoprotective capacity to the freezing diluent used for Belgian blue bull sperm during the process of freezing-thawing, and the optimal concentration of MLT for the frozen diluent was 0.3 mg/mL. • Optimizing our laboratory's semen diluent formulation by adding MLT. • Belgian blue bulls have good research and production advantages. • Adding 0.3 mg/mL MLT significantly improves bull semen storage efficiency. [ABSTRACT FROM AUTHOR]

Although under appropriate laboratory conditions, sperm from different mammalian species can be capacitated in vitro , the optimal conditions for sperm capacitation in the stallion have been elusive. This study evaluated the effect of different capacitating inducers in Whitten and Tyrode media and assessed their impact on capacitation-related factors. Stallion sperm were incubated with different combinations of capacitating inducers at 38.5 °C in an air atmosphere. Sperm quality variables such as motility, mitochondrial membrane potential, and lipid peroxidation were assessed. Membrane fluidity and intracellular calcium levels were evaluated as early markers of capacitation, while tyrosine phosphorylation events and the sperm's ability to perform acrosomal exocytosis were used as late capacitation markers. Finally, these sperm were evaluated using a heterologous zona pellucida binding assay. The findings confirm that capacitating conditions evaluated increase intracellular calcium levels and membrane fluidity in both media. Similarly, including 2 or 3 inducers in both media increased tyrosine phosphorylation levels and acrosomal exocytosis after exposure to progesterone, confirming that stallion sperm incubated in these conditions shows cellular and molecular changes consistent with sperm capacitation. Furthermore, the zona pellucida binding assay confirmed the binding capacity of sperm incubated in capacitation conditions, a key step for stallion in vitro fertilization success. Further studies are needed to evaluate the effect of these conditions on in vitro fertilization in the horse. • Different treatments for equine sperm capacitation were evaluated in two media. • Combination of capacitating inducers boosts phosphorylation and acrosomal exocytosis. • Capacitating conditions improve sperm's zona pellucida binding capacity. [ABSTRACT FROM AUTHOR]

Backgroud: Before fertilization, spermatozoa undergo a crucial maturation step called capacitation, which is a unique event regulates the sperm's ability for successful fertilization. The capacitation process takes place as the spermatozoa pass through the female reproductive tract (FRT). Dihydrolipoamide dehydrogenase (DLD) protein is a post-pyruvate metabolic enzyme, exhibiting reactive oxygen species (ROS) production which causes capacitation. Additionally, other vital functions of DLD in buffalo spermatozoa are hyperactivation and acrosome reaction. DLD produces the optimum amount of ROS required to induce capacitation process in FRT. Depending on physiological or pathophysiological conditions, DLD can either enhance or attenuate the production of reactive oxygen species (ROS). Aim of this study was to investigate whether changes in the production of ROS in sperm cells can impact their ability to fertilize by triggering the capacitation and acrosome reaction. Results: In this study, abundance of DLD protein was quantified between high (n = 5) and low fertile bull (n = 5) spermatozoa. It was found that compared to high-fertile (HF) bulls, low-fertile (LF) bulls exhibited significantly (P < 0.05) higher DLD abundances. Herein, we optimised the MICA concentration to inhibit DLD function, spermatozoa were treated with MICA in time (0, 1, 2, 3, 4, and 5 h) and concentrations (1, 2.5, 5, and 10 mmol/L) dependent manner. Maximum DLD inhibition was found to be at 4 h in 10 mmol/L MICA concentration, which was used for further experimentation in HF and LF. Based on DLD inhibition it was seen that LF bull spermatozoa exhibited significantly (P < 0.05) higher ROS production and acrosome reaction in comparison to the HF bull spermatozoa. The kinematic parameters of the spermatozoa such as percent total motility, velocity parameters (VCL, VSL, and VAP) and other parameters (BCF, STR, and LIN) were also decreased in MICA treated spermatozoa in comparison to the control (capacitated) spermatozoa. Conclusions: The present study provides an initial evidence explaining the buffalo bull spermatozoa with higher DLD abundance undergo early capacitation, which subsequently reduces their capacity to fertilize. [ABSTRACT FROM AUTHOR]

Objective: This work aimed to investigate the potential correlation between chromosomal polymorphisms and various reproductive abnormalities. Methods: We examined 21,916 patients affected by infertility who sought care at the Department of Reproductive Medicine, Affiliated Hospital of Shandong Second Medical University between January 2018 and December 2022. A total of 2227 individuals identified as chromosomal polymorphism carriers constituted the polymorphism group, and 2245 individuals with normal chromosome karyotypes were randomly selected to form a control group. Clinical manifestations, histories of spontaneous miscarriage, abnormal reproductive developments, fetal abnormalities, and male sperm quality anomalies were statistically compared between these two groups. Results: Of the 21,916 patients analyzed, 2227 displayed chromosomal polymorphism, representing a 10.16% detection rate. Amongst the male patients, 1622 out of 10,827 exhibited polymorphisms (14.98%), whereas 605 out of 11,089 females showed polymorphisms (5.46%). Female carriers in the polymorphism group, showed statistically significant increased rates of spontaneous abortion (29.75% vs. 18.54%), fetal anomalies (1.32% vs. 0.81%), and uterine abnormalities compared with the control group (1.32% vs. 0.81%). Male carriers in the polymorphism group had higher rates of spontaneous abortion in partners (22.87% vs. 10.37%), fetal anomalies (1.97% vs. 0.25%), compromised sperm quality (41.74% vs. 7.18%), testicular underdevelopment (2.28% vs. 0.92%), and hypogonadotropic hypogonadism (0.62% vs. 0.37%) compared with the control group. Conclusion: Chromosomal polymorphisms may have a certain negative effect on reproductive irregularities, including spontaneous abortions, fetal anomalies, and reduced sperm quality in males. Their clinical effects deserve further investigation. [ABSTRACT FROM AUTHOR]

Introduction: Over the past few decades, there has been growing concern about the concurrent trends of increasing Autism Spectrum Disorder (ASD) prevalence and declining sperm quality. These trends represent significant public health challenges that warrant thorough investigation of their underlying causes and implications. Objectives: The primary objectives of this study are to analyze trends in ASD prevalence and sperm quality parameters from 2000 to 2024, assess the statistical significance and effect size of these trends, explore potential correlations between ASD prevalence and sperm quality parameters, and identify significant predictors among sperm quality parameters that influence ASD prevalence. Methods: This study employed a longitudinal approach using multiple regression, time series analysis, ANOVA, Principal Component Analysis (PCA), hierarchical clustering, logistic regression, and cross-correlation analysis. Data on ASD prevalence were sourced from the CDC Autism and Developmental Disabilities Monitoring Network, while sperm quality data were collected from various published studies. Results: The findings reveal significant negative associations between ASD prevalence and sperm quality parameters such as sperm concentration and motility, suggesting that better sperm quality is linked to lower ASD rates. Conversely, parameters like sperm DNA fragmentation (SDF), volume of ejaculate, pH level, and semen viscosity show positive associations with ASD prevalence, indicating higher values in these parameters correlate with higher ASD rates. Conclusion: The study highlights the importance of maintaining reproductive health to potentially mitigate ASD risk and calls for further research to elucidate the underlying mechanisms driving these trends. These findings support the hypothesis that reproductive health factors play a crucial role in ASD etiology and suggest potential biological markers for assessing ASD risk. [ABSTRACT FROM AUTHOR]

Background: In polygynous species, the development of secondary sexual characters is usually decisive for male reproductive success. However, our understanding about the links between the growth of these traits and reproductive efficiency is still elusive. Most research efforts in this topic have been also focused on adult males, although the development of some secondary sexual characters, like bovid horns, typically starts after birth, continues during the puberty and in some species, such as the common eland, slows or even stops during adulthood. In this study, we investigated the relationships between horn size and testicular function during sexual development in common elands using a comprehensive approach that considers both spermatogenic and sperm parameters. Methods: Twenty-two non-sexually mature common elands were used for the present study. Horn size, body mass, testes mass, and gonadosomatic index were assessed. Spermatogenic activity was determined by cytological and histological analyses. Sperm concentration, morphology, morphometry, and intramale variation in sperm size were evaluated on epididymal sperm samples. Cluster analysis was performed to explore the influence of age on relationships between horn size and reproductive function. Results: We found that bigger horns are associated with increased Sertoli cell efficiency and reduced intramale variation in sperm size. Both parameters were not related to one another while they have shown to be associated with enhanced sperm quality in ungulates. Moreover, horn size was positively linked to the testis mass, sperm concentration, and testicular investment in the seminiferous epithelium. Spiral length and basal circumference were the horn traits most strongly correlated with spermatogenic and sperm parameters as well as those responsible for the sexual dimorphism in this species. Cluster analysis rendered two groups: the first one including males ≤30 months old, while the second one those >30 months old. Horn development and reproductive function were still correlated within age groups, with the strongest relationship found between horn size and sperm size homogeneity in males >30 months old. Conclusion: Taken together, our results indicate that horn size can be regarded as a good index of male reproductive potential during sexual development and provide insights into the role of secondary sexual characters in sexual selection dynamics. [ABSTRACT FROM AUTHOR]

Objective: The aim of the current study was to assess the effects of Nigella sativa essential oil on testicular toxicity in mice induced by dexamethasone. Materials and Methods: Forty NMRI mice were randomly divided into four groups. The first group (Sham) received 1 ml per day of normal saline by intraperitoneal (i.p.) injection for 7 days. The second group (Control) received (i.p) injection of 5 mg/kg dexamethasone for 7 days. The third group (Dexa+ N.S 5 mg/kg) received dexamethasone (5 mg/kg) and gavaged 5 mg/kg N. sativa essential oil for 7 days. The fourth group (N.S 5 mg/kg) for 7 days was gavaged 5 mg/kg N. sativa. Histopathology of testis, spermatogenesis, and sperm fertility rate were assessed. Results: The results of histopathology assessments showed that in the third group, all histopathology criteria were decreased compared to the second group. The number of seminiferous tubules that had abnormal spermatogenesis in Johnsen’s score was slightly decreased in the third group compared to the second group. Furthermore, in the third group, embryo formation criteria were increased. Conclusion: The data of this research demonstrate that N. sativa improves spermatogenesis defects and sperm fertility in mice treated with dexamethasone. [ABSTRACT FROM AUTHOR]

Background: Currently, high polyspermy remains a significant obstacle to achieving optimal efficiency in in vitro fertilization (IVF) and in vitro embryo production (IVP) systems in pigs. Developing strategies that would prevent polyspermy is essential in overcoming this challenge and maximizing the potential of this reproductive biotechnology. Previous results have demonstrated that using boar spermatozoa subjected to a high‐extension and reconcentration procedure and then cryopreserved resulted in significant improvements in IVF/IVP systems with high rates of monospermy and penetration. Objective: The aim of the present study was to unveil the molecular mechanisms that may underlie the changes in fertilization patterns exhibited by highly extended and cryopreserved boar spermatozoa. Materials and methods: To achieve this goal, we used quantitative proteomic analysis (LC‒ESI‒MS/MS SWATH) to identify differentially abundant proteins (DAPs) between highly extended (HE) and conventionally (control; CT) cryopreserved boar spermatozoa. Prior to the analysis, we evaluated the in vitro post‐thawing fertilizing ability of the sperm samples. The results demonstrated a remarkable improvement in monospermy and IVF efficiency when using HE spermatozoa in IVF compared with CT spermatozoa. Results: At the proteomic level, the combination of high‐extension and cryopreservation had a significant impact on the frozen–thawed sperm proteome. A total of 45 proteins (24 downregulated and 21 upregulated) were identified as DAPs (FC > 1 or ≤1; p < 0.05) when compared with CT spermatozoa. Some of these proteins were primarily linked to metabolic processes and the structural composition of sperm cells. The dysregulation of these proteins may have a direct or indirect effect on essential sperm functions and significantly affect spermatozoa–oocyte interaction and, therefore, the sperm fertilization profile under in vitro conditions. While these findings are promising, further research is necessary to comprehend how the disturbance of specific proteins affects sperm fertilization ability. [ABSTRACT FROM AUTHOR]

Goat bucks are seasonal breeders that show variation in sperm quality, endogenous melatonin (MLT), and presumably in the expression of MLT receptors on the sperm throughout the year, which may modify sperm freezability. The aim of this study was to determine whether sperm freezability is associated with (i) endogenous melatonin levels in seminal plasma and (ii) the expression of sperm plasma membrane melatonin receptors (MT1, MT2). To evaluate this, spermatozoa from seven Saanen goat bucks were cryopreserved throughout the year in Mexico using a standard freezing protocol. Seminal plasma MLT concentrations were determined by ELISA and the expression and localization of MT1 and MT2 were detected by immunocytochemistry and confirmed by western blotting. The recovery rate of progressive motility after thawing was higher in spring than autumn and winter; in contrast, the F pattern (CTC assay) was higher in winter than in the other seasons. A proportional increase in the AR pattern (CTC assay) was smaller in winter than in the other seasons and the proportion of sperm showing high plasma membrane fluidity was higher in spring than in summer and autumn. The seminal plasma MLT concentrations showed no significant interseasonal differences. The MT1 receptor was immunolocalised at the apical region of the sperm head, while MT2 was mainly localised in the neck. The relative expression of MLT receptors showed significant differences between summer and winter for all bands, except at 75 kDa of MT2. In conclusion, there was an association between the relative expression of MT1 and MT2 receptors throughout the year and sperm freezability in goat bucks in México. Post-thaw sperm quality is enhanced in semen samples collected during breeding season. • MLT concentration in goats' seminal plasma does not show interseasonal variation. • Expression of MLT receptors in goat spermatozoa shows variation throughout the year. • Goat buck sperm freezability is associated with the expression of MLT receptors. [ABSTRACT FROM AUTHOR]

Fourier harmonic analysis (FHA) is a robust method for identification of minute changes in sperm nuclear shape that are indicative of reduced fertility. The current study was designed to develop a fertility prediction model for Nili-Ravi buffalo bulls through FHA of sperm. In experiment I, FHA technique was standardized, average sperm nuclear perimeter was measured and sperm nuclear shape plot of buffalo bull was constructed. Sperm of buffalo bulls (n = 10) were stained with YOYO-1 and Hoechst-33342 to differentiate live and dead, and digital images were captured using phase contrast and fluorescent microscopy. The images were analyzed by ImageJ software and 100 sperm/bull were evaluated. The results are described as mean ± SEM values of mean harmonic amplitude (mharm), skewness harmonic amplitude (skharm), kurtosis harmonic amplitude (kurharm) and variance harmonic amplitude (varharm) at Fourier frequencies 0–5 along with the cartesian and polar coordinate plots of buffalo bull sperm. In experiment II, a fertility prediction model was developed based on FHA of buffalo bull sperm. Semen samples of low (n = 6), medium (n = 3) and high (n = 8) fertility bulls were investigated for FHA of sperm and harmonic amplitudes (HA) were generated. Firstly, to determine if live and dead sperm population have unique nuclear shape distribution; the mean, skewness, kurtosis and variance HA 0–5 of 1700 live and 1294 dead spermatozoa of 17 bulls were evaluated. T-test signified a difference in the mharm0 (2.363 ± 0.01 vs. 2.439 ± 0.02), skharm0 (−0.0002 ± 0.07 vs. −0.266 ± 0.09), kurharm0 (−0.156 ± 0.07 vs. 0.260 ± 0.18), kurharm2 (0.142 ± 0.11 vs. 1.031 ± 0.32) and varharm4 (0.109 ± 0.00 vs. 0.082 ± 0.00) of live vs. dead sperm population (p < 0.05). Therefore, 100 live sperm/bull were further evaluated for mean, skewness, kurtosis and variance HA 0–5 values among high (n = 6) and low-fertility (n = 6) groups. Results of T-test showed higher values of mharm2 (0.739 ± 0.01 vs. 0.686 ± 0.00), mharm4 (0.105 ± 0.001 vs. 0.007 ± 0.001), and skharm0 (0.214 ± 0.109 vs. −0.244 ± 0.097) in high vs. low-fertility group (p < 0.05). In next step, five significantly different combinations of discriminant measures between high and low-fertility groups were obtained by discriminant analysis. In conclusion, mharm4, skharm0 and varharm2 correctly identified 91.7 % of bulls into their respective fertility groups, and upon cross validation the value of the canonical correlation was 0.928. • Fourier harmonic analysis (FHA) technique was standardized for Nili-Ravi buffalo sperm. • Average sperm nuclear perimeter was measured & sperm nuclear shape plot constructed. • FHA detected difference in live and dead sperm nuclear shape. • Sperm from high and low fertility bulls statistically differed in nuclear shape. • Discriminant analysis based on FHA defined an excellent fertility prediction model. [ABSTRACT FROM AUTHOR]

Sexual dimorphism and male combat are observed in many species. Often, the outcome of male combat affects the outcome of subsequent combats, mating success, number of sperm, and fitness of the male's offspring. Also, the quantity and quality of sperm may be regulated by winning or losing, depending on species ecology and mating system. However, very few studies have experimentally examined the influence of fight outcomes on male offspring fitness. We studied male combat in the giant mealworm (Zophobas atratus) in which males bite each other's hind legs. We hypothesized that subsequent fitness could differ between winners and losers in the escalated male combat of this species. We measured several fitness traits including the number of eggs laid by mated females, and the number of hatches sired by uncontested males, winners, and losers in escalated and non-escalated combat, and compared the fitness of each winner and loser to that of an uncontested male. We also measured mating duration. The numbers of eggs and the percentages of hatched eggs of losers in the escalated combat were significantly reduced compared to that of the uncontested males. This reduction may be due to injuries from escalated leg- biting fights and a result of the sperm amount of the uncontested males being greater than that of the loser males. [ABSTRACT FROM AUTHOR]

The use of herbal bio-actives and phyto-contents to increase livestock productivity is a revived interest in veterinary research. The effect of wild strawberry {Fragaria vesca (FV)} pulp extract (PE) as antioxidant supplement in bovine semen cryopreservation diluent was explored. The experiment involved three fertile bulls. Three semen ejaculates per animal were collected. Each sample was diluted with Semen Transportation Diluent (STD) in 1:2 ratio. The (semen + STD) sample was vortexed and centrifuged (at 32g for 15 min) to remove 50 % supernatant. The remaining sample was re-diluted with semen cryopreservation diluent (SCD) in the ratio 1:4. Three aliquots of 0.3 ml were set aside from each (semen + STD + SCD) sample and were further treated with; 1: 0.7 mL of SCD (SCD group); 2: 0.7 mL SCD and 0.8 % FV-PE (FV-PE 0.8) and; 3: 0.7 mL SCD and 1.6 % FV-PE (FV-PE 1.6). Sperm quality parameters were evaluated before and after 48 h of cryopreservation. Results showed significantly enhanced mean percentage of sperm viability, progressive motility, velocity and fertilizability in FV-PE 0.8 and FV-PE 1.6 groups as compared with SCD group. The findings propose the protective and ameliorative potential of FV-PE for cryopreserved bovine sperms. [ABSTRACT FROM AUTHOR]

Context: Integrated omics studies hold a crucial role in improving our understanding of reproductive biology. However, the complex datasets so generated are often only accessible via supplementary data files, which lack the capacity for interactive features to allow users to readily interrogate and visualise data of interest. Aims: The intent of this technical note was to develop an interactive web-based application that enables detailed interrogation of a representative sperm proteome, facilitating a deeper understanding of the proteins identified, their relative abundance, classifications, functions, and associated phenotypes. Methods: We developed a Shiny web application, ShinySperm (), utilising R and several complementary libraries for data manipulation (dplyr), interactive tables (DT), and visualisation (ggplot2, plotly). ShinySperm features a responsive user interface, dynamic filtering options, interactive charts, and data export capabilities. Key results: ShinySperm allows users to interactively search, filter, and visualise sperm proteomics data based on key features (e.g. protein classification, sperm cell domain, presence, or absence at different maturation stages). This application responds live to filtering options and enables the generation of interactive plots and tables, thus enhancing user engagement and understanding of the data. Conclusions: ShinySperm provides a robust platform for the dynamic exploration of epididymal sperm proteome data. It significantly improves accessibility and interpretability of complex datasets, allowing for effective data-driven insights. Implications: This technical note highlights the potential of interactive web applications in reproductive biology and provides a plug and play script for the field to produce applications for meaningful researcher interaction with complex omic datasets. Understanding complex biological data is crucial for advancements in reproductive biology. Our study developed ShinySperm, a web application that allows researchers to easily explore, interrogate, and visualise intricate sperm proteomic data. This tool sets out a blueprint for future research, enabling researchers to bring their intricate datasets to life, allowing others to interact with and interpret more effectively, ultimately driving innovation in reproductive biology. Image by David Skerrett-Byrne. This article belongs to the Collection Frontiers in Reproduction Science – Rising Stars at the Society for Reproductive Biology 2023. [ABSTRACT FROM AUTHOR]

The effects of honey addition to Tris media on osmolality, pH values and on the quality characteristics of fresh and chilled goat spermatozoa were studied. Semen samples were collected from five Shami bucks and incubated at 20 °C for the fresh samples and at 5 °C for the chilled ones in Tris-Based Medium (TBM) and Tris-Egg Yolk (TEY) medium without (controls) and with honey at ratios of 1%, 2% and 4% (v/v) respectively. Osmolality and pH levels were determined for all used media. Spermatozoa motility was analysed using the Computer Assisted Sperm Analyzer (CASA) system and the integrity of sperm plasma membrane was tested by the Hypo-Osmotic Swelling Test (HOST). Osmolality values were significantly increased (P<0.05) by honey addition reaching 558 mOsm/kg in TBM and 675 mOsm/kg in TEY with 4% of honey compared with 307 mOsm/kg and 312 mOsm/kg for the controls respectively. In contrast, pH values decreased (P<0.05) from 6.93 in TBM control medium to 6.4 in TBM with 4% of honey and from 6.83 in TEY control medium to 6.1 in TEY with 4% of honey. CASA motility parameters including the percent motility (MOT%) and the percent of sperm showing progressive motility (PMOT%) increased after adding 1% and 2% of honey while the addition of 4% led to a clear decrease (P<0.05) for the fresh and chilled samples. The percentages of spermatozoa subpopulations including the rapid % and the static % categories for fresh and chilled samples were significantly decreased and increased respectively after adding 4% honey. Moreover, for all media and for all incubation temperatures, the HOST percentage decreased significantly (P<0.05) by adding 4% honey. It was concluded that honey addition to Tris media influenced the osmolality and pH values, which in turn affected the quality characteristics of buck spermatozoa based on honey concentration regardless of the incubation temperature or the used media. [ABSTRACT FROM AUTHOR]

Aging in men is associated with diminished sperm quality and a higher incidence of altered fetal development and miscarriage in resultant pregnancies. We used a mouse model of advanced paternal age to characterize embryonic development in older male mice and tested whether pre-conception treatment with the mitochondrial activator BGP-15 improves reproductive outcomes in old males. Like older men, reproductively old male mice had higher levels of sperm DNA damage and delayed pre-implantation development, associated with a reduced fetal weight and placental weight. Analysis of neonatal outcomes of in vivo-conceived offspring found that pups sired by old males were smaller, had delayed locomotor development, and increased mortality. BGP-15 treatment for 5 days prior to conception reduced sperm DNA oxidation levels and improved on-time embryo development after IVF and pup survival. BGP-15 treatment for 3 weeks prior to conception improved on-time pre-implantation embryo development and fetal viability and increased fetal size in pregnancies sired by old males. These results validate that ageing negatively affects male fertility and offspring physiology and indicates that pre-conception treatment with BGP-15 has the potential to improve sperm quality as well as early embryo development and post-natal health. [ABSTRACT FROM AUTHOR]

Zinc finger MYND‐type containing 15 (ZMYND15) has been documented to play important roles in spermatogenesis, and mutants contribute to recessive azoospermia, severe oligozoospermia, non‐obstructive azoospermia, teratozoospermia, even male infertility. ZMYND10 is involved in sperm motility. Whether environmental pollutants impair male fertility via regulating the expression of ZMYND15 and ZMYND10 has not been studied. Arsenic exposure results in poor sperm quality and male infertility. In order to investigate whether arsenic‐induced male reproductive toxicity is related to the expression of ZMYND15, ZMYND10 and their target genes, we established a male rat model of sodium arsenite exposure‐induced reproductive injury, measured sperm quality, serum hormone levels, mRNA and protein expressions of intratesticular ZMYND15 and ZMYND10 as well as their target genes. The results showed that, in addition to the increased mRNA expression of Tnp1, sodium arsenite exposure reduced sperm quality, serum hormone levels, and mRNA and protein expression of intratesticular ZMYND15 and ZMYND10 and their target genes in male rats compared with the control group (p <.05). Therefore, our study first showed that the environmental pollutant arsenic impairs sperm quality in male rats by reducing the expression of ZMYND10 and ZMYND15 and their regulatory genes, which provides a possible diagnostic marker for environmental pollutants‐induced male infertility. [ABSTRACT FROM AUTHOR]

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The INO80D protein, a component of the INO80 chromatin remodeling complex, plays a pivotal role in chromatin remodeling, gene expression, and DNA repair within mammalian sperm. In contrast to the condensed nuclear structure of mammalian sperm, Chinese mitten crab, Eriocheir sinensis, exhibits a distinctively decondensed sperm nucleus. The distribution and function of INO80D during the E. sinensis spermatogenesis were previously enigmatic. Our research endeavored to elucidate the distribution and function of INO80D, thereby enhancing our comprehension of sperm decondensation and the process of spermatogenesis in this species. Employing transcriptome sequencing, RT-qPCR, western blot analysis, and immunofluorescence techniques, we observed a pronounced upregulation of INO80D in the adult E. sinensis in comparison to the juvenile. The protein predominantly resides in the cellular nucleus, with high levels in spermatogonia and spermatocytes, less in stage I and III spermatids, and lowest in mature sperm. The results indicated that INO80D is initially instrumental in chromatin decondensation to facilitate gene accessibility and DNA repair during the early phases of spermatogenesis. Its role subsequently shifts to maintaining decondensed chromatin stability and genetic integrity during spermiogenesis. The sustained presence of INO80D during spermiogenesis is essential for the ultimate maturation of the decondensed sperm nucleus, imperative for preserving the unique decondensed state and the protection of genetic material in E. sinensis. Our study concludes that INO80D exerts a multifaceted influence on the spermatogenesis of E. sinensis, impacting chromatin decondensation, genetic integrity, and the regulation of early gene expression. This understanding could potentially improve crab breeding in aquaculture. [ABSTRACT FROM AUTHOR]

Voltage-sensing phosphatase (VSP) exhibits voltage-dependent phosphatase activity toward phosphoinositides. VSP generates a specialized phosphoinositide environment in mammalian sperm flagellum. However, the voltage-sensing mechanism of VSP in spermatozoa is not yet characterized. Here, we found that VSP is activated during sperm maturation, indicating that electric signals in immature spermatozoa are essential. Using a heterologous expression system, we show the voltage-sensing property of mouse VSP (mVSP). The voltage-sensing threshold of mVSP is approximately −30 mV, which is sensitive enough to activate mVSP in immature spermatozoa. We also report several knock-in mice in which we manipulate the voltage-sensitivity or electrochemical coupling of mVSP. Notably, the V312R mutant, with a minor voltage-sensitivity change, exhibits abnormal sperm motility after, but not before, capacitation. Additionally, the V312R mutant shows a significant change in the acyl-chain profile of phosphoinositide. Our findings suggest that electrical signals during sperm maturation are crucial for establishing the optimal phosphoinositide environment in spermatozoa. Membrane potentials in living cells can function as electrical signals. Here, the authors report that spermatozoa utilize their electrical signals during the maturing stage to create appropriate membrane lipids environment that is important for sperm motility. [ABSTRACT FROM AUTHOR]

Ribonucleoprotein (RNP) granules are membraneless electron-dense structures rich in RNAs and proteins, and involved in various cellular processes. Two RNP granules in male germ cells, intermitochondrial cement and the chromatoid body (CB), are associated with PIWI-interacting RNAs (piRNAs) and are required for transposon silencing and spermatogenesis. Other RNP granules in male germ cells, the reticulated body and CB remnants, are also essential for spermiogenesis. In this study, we disrupted FBXO24, a testis-enriched F-box protein, in mice and found numerous membraneless electron-dense granules accumulated in sperm flagella. Fbxo24 knockout (KO) mice exhibited malformed flagellar structures, impaired sperm motility, and male infertility, likely due to the accumulation of abnormal granules. The amount and localization of known RNP granule-related proteins were not disrupted in Fbxo24 KO mice, suggesting that the accumulated granules were distinct from known RNP granules. Further studies revealed that RNAs and two importins, IPO5 and KPNB1, abnormally accumulated in Fbxo24 KO spermatozoa and that FBXO24 could ubiquitinate IPO5. In addition, IPO5 and KPNB1 were recruited to stress granules, RNP complexes, when cells were treated with oxidative stress or a proteasome inhibitor. These results suggest that FBXO24 is involved in the degradation of IPO5, disruption of which may lead to the accumulation of abnormal RNP granules in sperm flagella. [ABSTRACT FROM AUTHOR]

Fine particulate matter (PM 2.5), a significant component of air pollution particulate matter, is inevitable and closely associated with increasing male reproductive disorder. However, the testicular targets of PM 2.5 and its toxicity related molecular mechanisms are still not fully understood. In this study, the conditional knockout (cKO) mice and primary Leydig cells were used to explore the testicular targets of PM 2.5 and the related underlying mechanisms. First, apparent the structure impairment of seminiferous tubules, Leydig cells vacuolization, decline of serum testosterone and sperm quality reduction were found in male wild-type (WT) and Sirt1 knockout mice after exposure to PM 2.5. Enrichment analyses revealed that differentially expressed genes (DEGs) were enriched in steroid hormone biosynthesis, ferroptosis, and HIF-1 signaling pathway in the mice testes after exposure to PM 2.5 , which were subsequently verified by the molecular biological analyses. Notably, similar enrichment analyses results were also observed in primary Leydig cells after treatment with PM 2.5. In addition, Knockdown of Sirt1 significantly increased PM 2.5 -induced expression and activation of HIF-1α, which was in parallel to the changes of cellular iron levels, oxidative stress indicators and the ferroptosis markers. In conclusion, this highlights that PM 2.5 triggers ferroptosis via SIRT1/HIF-1α signaling pathway to inhibit testosterone synthesis in males. These findings provide a novel research support for the study that PM 2.5 causes male reproductive injury. [Display omitted] • PM 2.5 induced impairment of testicular structures and sperm parameters in mice. • Ferroptosis involved in PM 2.5 -induced male reproductive toxicity. • PM 2.5 inhibited testosterone synthesis-related enzymes. • PM 2.5 activated ferroptosis through SIRT1/HIF-1α signaling pathway. [ABSTRACT FROM AUTHOR]

Background: Alcohol consumption is widely known to have detrimental effects on various organs and tissues. The effects of ethanol on male reproduction have been studied at the physiological and cellular levels, but no systematic study has examined the effects of ethanol on male reproduction-related gene expression. Results: We employed a model of chronic ethanol administration using the Lieber-DeCarli diet. Ethanol-fed mice showed normal testicular and epididymal integrity, and sperm morphology, but decreased sperm count. Total RNA sequencing analysis of testes from ethanol-fed mice showed that a small fraction (∼ 2%) of testicular genes were differentially expressed in ethanol-fed mice and that, of these genes, 28% were cell-type specific in the testis. Various in silico analyses were performed, and gene set enrichment analysis revealed that sperm tail structure-related genes, including forkhead box J1 (Foxj1), were down-regulated in testes of ethanol-fed mice. Consistent with this result, ethanol-fed mice exhibited decreased sperm motility. Conclusion: This study provides the first comprehensive transcriptomic profiling of ethanol-induced changes in the mouse testis, and suggests gene expression profile changes as a potential mechanism underlying ethanol-mediated reproductive dysfunction, such as impaired sperm motility. [ABSTRACT FROM AUTHOR]

Introduction: The epididymis is important for sperm transport, maturation, and storage. Methods: The head and tail of the epididymis of 5-week-old and 10-week-old C57 BL/6J male mice were used for single-cell sequencing. Results: 10 cell types including main, basal, and narrow/clear cells are identified. Next, we performed cell subgroup analysis, functional enrichment analysis, and differentiation potential prediction on principal cells, clear cells, and basal cells. Our study indicates that the principal cells are significantly involved in sperm maturation, as well as in antiviral and anti-tumor immune responses. Clear cells are likely to play a crucial role in safeguarding sperm and maintaining epididymal pH levels. Basal cells are implicated in the regulation of inflammatory and stress responses. The composition and functions of the various cell types within the epididymis undergo significant changes before and after sexual maturity. Furthermore, pseudo-temporal analysis elucidates the protective and supportive roles of epididymal cells in sperm maturation during sexual maturation. Discussion: This study offers a theoretical framework and forecasts for the investigation of epididymal sperm maturation and epididymal immunity. [ABSTRACT FROM AUTHOR]

Simple Summary: Capacitation confers competency to spermatozoa to fertilize the oocyte, yet its regulatory mechanisms are not fully understood. Thus, we aimed to investigate the comparative proteomic profiling in ram spermatozoa under non-capacitating (NC) and capacitating (CAP) conditions in vitro using a liquid chromatography–tandem mass spectrometry (LCMS/MS) combined with tandem mass tag (TMT) labeling strategy. Functional enrichment analysis indicated that the differentially abundant proteins Prune Exopolyphosphatase 1, Galactose-1-Phosphate Uridylyltransferase, and ATP Citrate Lyase were strictly related to energy production and conversion, and Phosphoglycolate phosphatase, Glucosamine-6-Phosphate Deaminase 1 and 2 were related to metabolism, RNA processing, and vesicular transport pathways. Furthermore, the networks of protein–protein interaction indicated a strong interaction among these differential proteins in annotated pathways such as ubiquitin and transport metabolism. Together, our results provided the database for studying the differentially expressed proteins during ram sperm capacitation. Ram sperm undergo a sequence of physiological and biochemical changes collectively termed as capacitation to perform oocyte fertilization. However, the protein changes induced by capacitation remain in need of further exploration. Thus, the present study investigated the comparative proteomic profiling in ram spermatozoa under non-capacitating (NC) and capacitating (CAP) conditions in vitro using a liquid chromatography–tandem mass spectrometry combined with tandem mass tag labeling strategy. As a results, 2050 proteins were identified and quantified; 348 of them were differentially abundant, with 280 of the proteins upregulated and 68 of the proteins downregulated between the CAP and NC spermatozoa, respectively. Functional enrichment analysis indicated that the differentially abundant proteins Prune Exopolyphosphatase 1, Galactose-1-Phosphate Uridylyltransferase, and ATP Citrate Lyase were strictly related to energy production and conversion, and Phosphoglycolate phosphatase, Glucosamine-6-Phosphate Deaminase 1 and 2 were related to metabolism, RNA processing, and vesicular transport pathways. Furthermore, the networks of protein–protein interaction indicated a strong interaction among these differential proteins in annotated pathways such as ubiquitin and transport metabolism. Our findings indicate that capacitation progress might be regulated through different pathways, providing insights into mechanisms involved in ram sperm capacitation and fertility. [ABSTRACT FROM AUTHOR]

Zinc (Zn) is an essential trace element; it exhibits a plethora of physiological properties and biochemical functions. It plays a pivotal role in regulating the cell cycle, apoptosis, and DNA organization, as well as in protein, lipid, and carbohydrate metabolism. Among other important processes, Zn plays an essential role in reproductive health. The ZIP and ZnT proteins are responsible for the mobilization of Zn within the cell. Zn is an inert antioxidant through its interaction with a variety of proteins and enzymes to regulate the redox system, including metallothioneins (MTs), metalloenzymes, and gene regulatory proteins. The role of Zn in the reproductive system is of great importance; processes, such as spermatogenesis and sperm maturation that occur in the testicle and epididymis, respectively, depend on this element for their development and function. Zn modulates the synthesis of androgens, such as testosterone, for these reproductive processes, so Zn deficiency is related to alterations in sperm parameters that lead to male infertility. [ABSTRACT FROM AUTHOR]

AbstractThe objective of this study was to compare the results of semen analysis using the manual method and the SQA-Vision sperm analyser after four years of practice and with a large cohort of patients. This was a comparative study of 1130 cases collected for semen analysis between October 2019 and October 2023, which were analysed simultaneously and independently by different operators using the manual microscopic method and an SQA-V automated analyser. For each sample, sperm concentration, progressive motility, motility, normal morphology, and round cells count were performed. There was no significant difference between the SQA-V method and manual assessment for all sperm parameters (Mann–Whitney test p > 0.05). According to the parameter studied, there was a strong correlation (rho = 0.81) and a very high correlation (rho = 0.98) between manual assessment and the SQA-V method. In the analysis of sperm concentration, the sensitivity and specificity were 0.90 and 0.99, respectively. The sensitivity and specificity for the analysis of sperm progressive motility were 0.98 and 0.99, respectively, while the sensitivity and specificity for the analysis of sperm motility were 0.87 and 0.99, respectively. The sensitivity and specificity for the analysis of normal morphology were 0.88 and 0.99, respectively. Regarding the analysis of round cells, the sensitivity and specificity were 0.98 and 0.99, respectively. The results of this retrospective study indicate that the SQA-V system offers satisfactory performance for routine sperm analysis. [ABSTRACT FROM AUTHOR]

This study aimed to investigate if washing ram sperm from seminal plasma (SP) could be an effective tool to extend sperm lifespan in medium-term preservation in liquid form to optimize ovine artificial insemination protocols. To this end, in Experiment 1 SP was added to a sperm model without previous contact with this substance (ram epididymal sperm) at the beginning or the end of a 48-hour preservation protocol at 5 °C (n = 13). Sperm motility and kinetic parameters and sperm functionality in terms of sperm viability, apoptosis, mitochondrial activity and reacted acrosomes were assessed after 6 h of storage at 15 °C (standard liquid preservation method) and 24 and 48 h at 5 °C. Extended sperm showed better results after 48 h when stored in the absence than in the presence of SP in most sperm quality parameters. Moreover, the final SP supplementation of this experimental group resulted in the highest sperm motility and kinetic parameters, viability and mitochondrial activity. These results suggested that initial SP deprivation could be beneficial in a medium-term ram sperm preservation protocol in liquid form, as well as a final supplementation. Therefore, we conducted Experiment 2 to evaluate the effect of SP removal from freshly ejaculated ram semen under the same storage conditions as in Experiment 1 (n = 12). Surprisingly, SP withdrawal impaired sperm functionality, leading to increased apoptosis and decreased mitochondrial activity after 24 and 48 h at 5 °C. Conversely, SP supplementation at the end of the preservation protocol of the ejaculate processed as usual had a positive effect on sperm quality and fertility. To summarize, SP absence was beneficial for a medium-term preservation protocol (up to 48 h at 5 °C) of ram epididymal sperm, but the same preservation protocol for ram ejaculated sperm revealed a possible failure of the SP removal method in avoiding the sperm-SP interaction effect. Meanwhile, SP supplementation of ram semen at the end of the preservation protocol increased in vitro sperm quality and fertility after artificial insemination. [ABSTRACT FROM AUTHOR]

Background Objectives Materials and methods Results Discussion Conclusion Small RNAs interacting with PIWI (piRNAs) play a crucial role in regulating transposable elements and translation during spermatogenesis and are essential in male germ cell development. Disruptions in the piRNA pathway typically lead to severe spermatogenic defects and thus male infertility. The HENMT1 gene is a key player in piRNAs primary biogenesis and dysfunction of HENMT1 protein in meiotic and haploid germ cells resulted in the loss of piRNA methylation, piRNA instability, and TE de‐repression. Henmt1‐knockout mice exhibit a severe oligo‐astheno‐teratozoospermia (OAT) phenotype, whereas patients with HENMT1 variants display more severe azoospermia phenotypes, ranging from meiotic arrest to hypospermatogenesis. Through whole‐exome sequencing (WES) of infertile patient cohorts, we identified two new patients with variants in the HENMT1 gene presenting spermatozoa in their ejcaulate, providing us the opportunity to study spermatozoa from these patients.Investigate the spermatozoa of two patients harboring an HENMT1 variant to determine whether or not these scarce spermatozoa could be used with assisted reproductive technologies.HENMT1 variants identified by WES were validated through Sanger sequencing. Comprehensive semen analysis was conducted, and sperm cells were subjected to transmission electron microscopy for structural examination, in situ hybridization for aneuploidy assessment, and aniline blue staining for DNA compaction status. Subsequently, we assessed their suitability for in vitro fertilization using intracytoplasmic sperm injection (IVF‐ICSI).Our investigations revealed a severe OAT phenotype similar to knockout mice, revealing altered sperm concentration, mobility, morphology, aneuploidy and nuclear compaction defects. Multiple IVF‐ICSI attempts were also performed, but no live births were achieved.We confirm the crucial role of HENMT1 in spermatogenesis and highlight a phenotypic continuum associated with HENMT1 variants. Unfortunately, the clinical outcome of these genetic predispositions remains unfavorable, regardless of the patient's phenotype.The presence of spermatozoa is insufficient to anticipate ICSI pregnancy success in HENMT1 patients. [ABSTRACT FROM AUTHOR]

Arboviruses can be paternally transmitted by male insects to offspring for long-term persistence, but the mechanism remains largely unknown. Here, we use a model system of a destructive rice reovirus and its leafhopper vector to find that insect ribosome-rescuer Pelo-Hbs1 complex expressed on the sperm surface mediates paternal arbovirus transmission. This occurs through targeting virus-containing tubules constituted by viral nonstructural protein Pns11 to sperm surface via Pns11-Pelo interaction. Tubule assembly is dependent on Hsp70 activity, while Pelo-Hbs1 complex inhibits tubule assembly via suppressing Hsp70 activity. However, virus-activated ubiquitin ligase E3 mediates Pelo ubiquitinated degradation, synergistically causing Hbs1 degradation. Importantly, Pns11 effectively competes with Pelo for binding to E3, thus antagonizing E3-mediated Pelo-Hbs1 degradation. These processes cause a slight reduction of Pelo-Hbs1 complex in infected testes, promoting effective tubule assembly. Our findings provide insight into how insect sperm-specific Pelo-Hbs1 complex is modulated to promote paternal virus transmission without disrupting sperm function. In this study, Sun et al identify the leafhopper Pelo-Hbs1 complex in protein translational quality control as a sperm factor that mediates paternal arbovirus transmission to offspring without impairing sperm function, which contributes to long-term virus persistence in its natural vector. [ABSTRACT FROM AUTHOR]

The karyotype of an organism is the set of gross features that characterize the way the genome is packaged into separate chromosomes. It has been known for decades that different taxonomic groups often have distinct karyotypic features, but whether selective forces act to maintain these differences over evolutionary timescales is an open question. In this paper we analyze a database of karyotype features and sperm head morphology in 103 mammal species with spatulate sperm heads and 90 sauropsid species (birds and non-avian reptiles) with vermiform heads. We find that mammal species with a larger head area have more chromosomes, while sauropsid species with longer heads have a wider range of chromosome lengths. These results remain significant after controlling for genome size, so sperm head morphology is the relevant variable. This suggest that post-copulatory sexual selection, by acting on sperm head shape, can influence genome architecture. [ABSTRACT FROM AUTHOR]

This document, titled "The Multifaceted Impact of Environmental Pollutants on Health and Ecosystems," discusses the pervasive effects of environmental pollutants on both ecosystems and human health. The document synthesizes findings from multiple studies and articles to highlight the intricate mechanisms and broader implications of various contaminants, including heavy metals, microplastics, and antibiotic resistance genes. The studies reviewed in this document emphasize the need for comprehensive risk reduction strategies to protect both the environment and public health, including reducing exposure to heavy metals, controlling the spread of antibiotic resistance, and addressing the proliferation of microplastics and endocrine disruptors. [Extracted from the article]

Background Objective Methods Results Conclusion Sleep deprivation (SD) can cause damage to the male reproductive system. However, the duration required for such damage and the specific sequence and severity of damage to the testis and epididymis remain unclear.To investigate the effects of different durations of SD on different parts of the testis and epididymis caput, corpus, and cauda.Adult ICR mice were randomly assigned to five groups: the SD group (SD for 18 h/day for 1, 2, 3, or 4 weeks), the SD + Vit E group (supplemented with Vit E 50 mg/kg/d during 4 weeks of SD, the SD+NS group (saline supplementation during 4 weeks of SD), the SD + RS group (5 weeks of recovery sleep after 4 weeks of SD), and a normal sleep control (Ctrl) group. Following the interventions, sperm parameters, testicular and epididymal histopathology, inflammatory response, and oxidative stress markers were compared between the groups.Compared to the Ctrl group, the SD group showed a decrease in sperm motility and concentration from SD 2 W and SD 3 W, respectively. Decreases in sperm concentration and motility were more pronounced in the cauda compared to the caput and corpus. Pathological damage was less severe in the epididymis caput than in the corpus and cauda. After 4 weeks of SD, inflammation and oxidative stress increased in both testes and epididymis. Both sleep recovery and vitamin E supplementation showed significant improvements, though they did not fully reach the level of the Ctrl group.Chronic SD for more than 2 weeks causes varying degrees of damage to the testis, epididymis caput, corpus, and cauda in male mice. This damage is not fully reversible after 5 weeks of sleep recovery and antioxidant stress treatment. These findings help us to identify and prevent SD damage to the male reproduction at an early stage. [ABSTRACT FROM AUTHOR]

The sperm epigenome is thought to affect the developmental programming of the resulting embryo, influencing health and disease in later life. Age‐related methylation changes in the sperm of old fathers may mediate the increased risks for reproductive and offspring medical problems. The impact of paternal age on sperm methylation has been extensively studied in humans and, to a lesser extent, in rodents and cattle. Here, we performed a comparative analysis of paternal age effects on protein‐coding genes in the human and marmoset sperm methylomes. The marmoset has gained growing importance as a non‐human primate model of aging and age‐related diseases. Using reduced representation bisulfite sequencing, we identified age‐related differentially methylated transcription start site (ageTSS) regions in 204 marmoset and 27 human genes. The direction of methylation changes was the opposite, increasing with age in marmosets and decreasing in humans. None of the identified ageTSS was differentially methylated in both species. Although the average methylation levels of all TSS regions were highly correlated between marmosets and humans, with the majority of TSS being hypomethylated in sperm, more than 300 protein‐coding genes were endowed with species‐specifically (hypo)methylated TSS. Several genes of the glycosphingolipid (GSL) biosynthesis pathway, which plays a role in embryonic stem cell differentiation and regulation of development, were hypomethylated (<5%) in human and fully methylated (>95%) in marmoset sperm. The expression levels and patterns of defined sets of GSL genes differed considerably between human and marmoset pre‐implantation embryo stages and blastocyst tissues, respectively. [ABSTRACT FROM AUTHOR]

To assess the accuracy of the YO home sperm test by real-world/amateur users. This multisite study investigated whether amateur users could use the FDA-cleared YO Home Sperm Test to obtain accurate motile sperm concentration (MSC) results when compared to trained laboratory technicians. The qualitative MSC results of amateur and professional YO users were compared to each other as well as to the results of an established automated sperm quality analyzer (SQA-V) above and below a 6 m/mL MSC cut-off. This was a blinded, prospective study of 316 amateur users and 3 professional laboratory technicians across 3 study sites. Amateur vs Professional YO users demonstrated an accuracy of >97%. Qualitative results of amateurs and professionals vs SQA-V results showed a >95.7% accuracy. Amateur users with no prior training were able to follow the YO test directions to receive highly accurate qualitative motile sperm concentration (MSC) results. The YO test is user-friendly and can be used as an effective initial home screening tool to gain preliminary insight into the fertility status of the male in a real-world setting. Furthermore, the YO test results correlated with those obtained by the FDA-cleared SQA-V laboratory analyzer, confirming that the YO test delivers accurate MSC results in the hands of amateur users. [ABSTRACT FROM AUTHOR]

• Extracts of Mondia whitei improve semen parameters in vitro in a dose dependent manner. • Extracts of Mondia whitei reduce the levels of sperm DNA fragmentation in vitro in a dose dependent manner. • Extracts of Mondia whitei increase the percentage of sperm with intact mitochondrial membrane potential in vitro. • Extracts of Mondia whitei increase the percentage of capacitated and acrosome reacted sperm in vitro. Mondia whitei (Mw) is a prominent African medicinal plant that is widely used as traditional treatment in the management of sexual dysfunction and male infertility. Yet, there is a paucity of scientific evidence to support its medicinal use. Therefore, the aim of this study was to investigate the in vitro effect of aqueous root extract of Mw on human semen parameters and sperm function. Semen samples were obtained from healthy donors (n = 30) and infertile patients (n = 30) for in vitro experiments. Samples were washed using human tubular fluid medium supplemented with bovine serum albumin (HTF-BSA), and incubated with increasing concentrations (0.0185, 0.185, 1.85, 18.5 and 185 μg/mL) of aqueous Mw rhizome extracts for 1 h at 37 °C. HTF-BSA washed sperm served as a control for the experiments. Sperm total and progressive motility, kinematic parameters, vitality, mitochondrial membrane potential (MMP), sperm DNA fragmentation (SDF), intracellular reactive oxygen species (ROS), capacitation and acrosome reaction were assessed following the termination of experiments. Extracts of Mw increased spermatozoa total motility, vitality, percentage of spermatozoa with intact MMP and intracellular ROS in both the donor and infertile groups. Mw increased progressive motility, SDF, capacitation and acrosome reaction in the infertile group only. Sperm kinetic motility parameters and hyperactivation were not significantly affected in any group. Although there was increased intracellular ROS generation, this did not negatively affect MMP, SDF, hyperactivation or acrosome reaction and capacitation. Mw improves functional human sperm parameters in vitro , increasing total and progressive motility, and intact MMP in a dose dependent manner. Further studies instigating the effect of Mw on male fertility are warranted. [ABSTRACT FROM AUTHOR]

Purpose: To identify the sperm preparation procedure that selects the best sperm population for medically assisted reproduction. Methods: Prospective observational study comparing the effect of four different sperm selection procedures on various semen parameters. Unused raw semen after routine diagnostic analysis was split in four fractions and processed by four different methods: (1) density gradient centrifugation (DGC), (2) sperm wash (SW), (3) DGC followed by magnetic activated cell sorting (MACS), and (4) using a sperm separation device (SSD). Each fraction was analyzed for progressive motility, morphology, acrosome index (AI), and DNA fragmentation index (DFI). Results: With DGC as standard of care in intraclass correlation coefficient analysis, only SSD was in strong disagreement regarding progressive motility and DFI [0.26, 95%CI (− 0.2, 0.58), and 0.17, 95%CI (− 0.19, 0.45), respectively]. When controlling for abstinence duration, DFI was significantly lower after both MACS and SSD compared to DGC [− 0.27%, 95%CI (− 0.47, − 0.06), p = 0.01, and − 0.6%, 95%CI (− 0.80, − 0.41), p < 0.001, respectively]. Further comparisons between SSD and MACS indicate significantly less apoptotic cells [Median (IQR) 4 (5), 95%CI (4.1, − 6.8) vs Median (IQR) 5 (8), 95%CI (4.9, − 9.2), p < 0.001, respectively] and dead cells [Median (IQR) 9.5 (23.3), 95%CI (13.2, − 22.4) vs Median (IQR) 22 (28), 95%CI (23.1, − 36.8), p < 0.001, respectively] in the SSD group. Conclusion: The selection of a population of highly motile spermatozoa with less damaged DNA from unprocessed semen is ideally performed with SSD. Question remains whether this method improves the embryological outcomes in the IVF laboratory. [ABSTRACT FROM AUTHOR]

Aim: The aim of this study is the evaluation effect of nanoliposome-loaded Mito-Tempo on sperm parameters during human sperm cryopreservation. Methods: Semen samples of 50 Asthenoteratozoospermia men (random) were collected. Sperm parameters were analyzed based on World Health Organization (WHO, 2010) criteria (2021) and each sample was divided into 5 groups (E1–E5). E1 (control group): the sperm was cryopreserved without nanoliposome, and Mito-Tempo. E2: sperm cryopreservation with Mito-Tempo-loaded nanoliposome (Mito-Tempo 0.1 mM) + freezing medium. E3: sperm cryopreservation with Mito-Tempo-loaded nanoliposome (Mito-Tempo 0.2 mM) + freezing medium. E4: in this group, the cryopreservation sperm with Mito-Tempo 0.3 mM + freezing medium. E5: the cryopreservation sperm with Mito-Tempo 0.2 mM + freezing medium. Results: The result of this study indicated that sperm parameters and total antioxidant capacity (TAC) significantly increase in E3 and E4 groups, compared to E1, E2, and E5 groups respectively (P < 0.05). The percentage of abnormal morphology, DNA fragmentation index (DFI), malondialdehyde (MDA), and the levels of ROS significantly decrease in E3 and E4 groups, compared to E1, E2, and E5 groups (P < 0.05). In addition, the sperm parameters and stress oxidative factors significantly improve in E3 group compared to other groups (P < 0.05). Conclusions: In conclusion, the combination of Mito-Tempo with nanoliposome due to its ability to cooperate with lipid layers may lead to significant performance in reducing oxidative stress damage and increasing the quality of sperm parameters. [ABSTRACT FROM AUTHOR]

Sperm ultrastructure is frequently employed as a source for phylogenetic inference due to the ease of accessing spermatozoa. Despite being unicellular, sperm cells exhibit a relatively high number of diverse characters and character states. Spermatozoa are subject to strong sexual selection as they are finely tuned for maximizing male reproductive success. Given this strong functional constraint, one might anticipate the emergence of convergent characters in line with similar modes of reproduction. As a result, it might be expected that sperm cells do not possess substantial phylogenetic signal, with functional constraints overshadowing any evolutionary heritage. To test this assumption, we conducted a study on sperm ultrastructure in 11 nemertean species, representing closely related groups and major nemertean lineages. We analyzed these data for their phylogenetic signal within the context of the most recent nemertean phylogenies. Our findings demonstrate that, at all systematic levels, functional constraints imposed by sexual selection, or the mode of reproduction do not supersede the influence of historical constraints on sperm ultrastructure. [ABSTRACT FROM AUTHOR]

The ultrastructural spermatozoon organisation of the acanthocolpid digenean Stephanostomum bicoronatum have been described by micrographs of transmission electron microscopy (TEM). Live digeneans were isolated from the digestive tract of the Brown meagre Sciaena umbra (Teleostei: Sciaenidae), obtained from artisanal fishing off the coast of La Chebba (Tunisia Mediterranean Sea). The ultrastructural study reveals that the male gamete of S. bicoronatum is a filamentous cell tapered at both extremities, which present typical characteristics of digenean spermatozoa of the type III, namely two axonemes of the 9 + '1' pattern of trepaxonematan Platyhelminthes, nucleus, one mitochondrion, external ornamentation of the plasma membrane associated with cortical microtubules and located in the posterior part of the anterior region of the sperm cell, two parallel bundles of cortical microtubules, and maximum number of cortical microtubules in the anterior part of the spermatozoon. In addition, the male gamete of S. bicoronatum shows spine-like bodies, a large amount of glycogen granules and a posterior extremity containing only the nucleus. The results are compared and discussed according to the existing data in other digeneans, particularly with those of the family Acanthocolpidae. [ABSTRACT FROM AUTHOR]

Background: Systematic reviews and meta‐analyses are instrumental in shaping clinical practice. However, their findings can sometimes be marred by discrepancies and potential biases, thereby diluting the strength of the evidence presented. Umbrella reviews serve to comprehensively assess and synthesise these reviews, offering a clearer insight into the quality of the evidence presented. In the context of the relationship between sperm DNA fragmentation (SDF) and assisted conception outcomes, there is a divergence in the literature. Some reviews suggest a clear cause‐and‐effect linkage, whereas others present conflicting or inconclusive results. Objectives: In this umbrella review we aimed to synthesise the evidence collated in systematic reviews and meta‐analyses summarising the association of SDF with assisted reproductive technology (ART) outcomes. Search strategy: After preregistration (https://doi.org/10.17605/OSF.IO/6JHDP), we performed a comprehensive search of the PubMed, Scopus, Cochrane Library, Web of Science and Embase databases. We conducted a search for systematic reviews on the association between SDF and ART without any restrictions on language or publication date. Selection criteria: Systematic reviews and meta‐analyses assessing the association between SDF and ART outcomes were eligible. Data collection and analysis: We assessed the quality of the included reviews using AMSTAR 2 and ROBIS, and determined the degree of overlap of primary studies between reviews estimating the corrected covered area (CCA), adjusted for structural missingness. We evaluated the most recent reviews assessing the association of SDF with live birth, pregnancy, miscarriage, implantation, blastulation and fertilisation. The synthesis of evidence was harmonised across all included quantitative syntheses, re‐estimating the odds ratio (eOR) in random‐effects meta‐analyses with 95% confidence intervals (95% CIs) and 95% prediction intervals (95% PIs). We categorised the evidence strength into convincing, highly suggestive, suggestive, weak or nonsignificant, according to the meta‐analysis re‐estimated P‐value, total sample size, I2 statistic for heterogeneity, small study effect, excess significance bias and the largest study significance. Main results: We initially captured and screened 49 332 records. After excluding duplicate and ineligible articles, 22 systematic reviews, 15 of which were meta‐analyses, were selected. The 22 reviews showed a moderate degree of overlap (adjusted CCA 9.2%) in their included studies (overall n = 428, with 180 unique studies). The 15 meta‐analyses exhibited a high degree of overlap (adjusted CCA = 13.6%) in their included studies (overall n = 274, with 118 unique studies). AMSTAR 2 categorised the quality of evidence in 18 reviews as critically low and the quality of evidence in four reviews as low. ROBIS categorised all the reviews as having a high risk of bias. The re‐estimated results showed that the association of SDF with live birth was weak in one and nonsignificant in four meta‐analyses. Similarly, the association of SDF with pregnancy, miscarriage, implantation, blastulation and fertilisation was also weak or nonsignificant. The association of high SDF with different ART outcomes was also weak or nonsignificant for different interventions (IVF, ICSI and IUI) and tests. Conclusions: This umbrella review did not find convincing or suggestive evidence linking SDF with ART outcomes. Caution should be exercised in making any claims, policies or recommendations concerning SDF. [ABSTRACT FROM AUTHOR]

Pigs are usually bred through artificial insemination with liquid semen preserved at 15–20 °C. While this method of preservation brings many benefits, including a greater reproductive performance compared to frozen-thawed sperm, the period of storage is a limiting factor. As the mitochondrion regulates many facets of sperm physiology, modulating its activity could have an impact on their lifespan. Aligned with this hypothesis, the present study sought to investigate whether inhibition of voltage-dependent anion channels (VDACs), which reside in the outer mitochondrial membrane and regulate the flux of ions between mitochondria and the cytosol in somatic cells, influences the resilience of pig sperm to liquid preservation at 17 °C. For this purpose, semen samples (N = 7) were treated with two different concentrations of TRO19622 (5 μM and 50 μM), an inhibitor of VDACs, and stored at 17 °C for 10 days. At days 0, 4 and 10, sperm quality and functionality parameters were evaluated by flow cytometry and computer-assisted sperm analysis (CASA). The effects of inhibiting VDACs depended on the concentration of the inhibitor. On the one hand, the greatest concentration of TRO19622 (50 μM) led to a decrease in sperm motility, viability and mitochondrial membrane potential, which could be related to the observed intracellular Ca2+ increase. In contrast, total sperm motility was higher in samples treated with 5 μM TRO19622 than in the control, suggesting that when VDACs channels are inhibited by the lowest concentration of the blocking agent the resilience of pig sperm to liquid storage increases. In conclusion, the current research indicates that mitochondrial function, as regulated by ion channels in the outer mitochondrial membrane like VDACs, is related to the sperm resilience to liquid preservation and may influence cell lifespan. • The effects of TRO19622 on pig sperm depend on the concentration of the inhibitor. • Inhibition of VDACs with 50 μM TRO19622 decreases sperm motility, viability and mitochondrial membrane potential. • Blockage of VDACs with 5 μM TRO19622 increases sperm motility, and has no detrimental impact on viability. • Mitochondrial function, as regulated by VDACs among other ion channels, is related to the liquid preservation of pig semen. [ABSTRACT FROM AUTHOR]

Objectives: The current study estimated the levels of anti mullerian hormone (AMH) and inhibin-B and reproductive hormones in non obstructive azoospermic (NOA) cases and obstructive azoospermic (OA) cases as well as comparing between them as regards the sensitivity and specificity in determining the sperm retrieval hope in the NOA cases. Finally, we analyzed any potential correlation between all the hormones measured in the current study. Methods: The current case control study was conducted at the andrology outpatient clinic from June (2021) to March (2022). The study recruited 135 participants divided equally into three groups NOA cases, OA cases and controls, respectively. Results: The mean inhibin-B was significantly lower in the NOA cases compared to the OA cases and the controls. There was a positive moderate correlation between AMH and Inhibin-B. Also, there was a positive moderate correlation between inhibin-B and free testosterone (FT) and positive weak correlation between beta Inhibin and leutinizing hormone (LH). Conclusions: The current study asserts the observation that inhibin-B is also expressed by Leydig cells as it has demonstrated positive correlations with FT and LH. [ABSTRACT FROM AUTHOR]

Drosophila melanogaster is an ideal model organism for investigating spermatogenesis due to its powerful genetics, conserved genes and visible morphology of germ cells during sperm production. Our previous work revealed that ocnus (ocn) knockdown resulted in male sterility, and CG9920 was identified as a significantly downregulated protein in fly abdomen after ocn knockdown, suggesting a role of CG9920 in male reproduction. In this study, we found that CG9920 was highly expressed in fly testes. CG9920 knockdown in fly testes caused male infertility with no mature sperms in seminal vesicles. Immunofluorescence staining showed that depletion of CG9920 resulted in scattered spermatid nuclear bundles, fewer elongation cones that did not migrate to the anterior region of the testis, and almost no individualization complexes. Transmission electron microscopy revealed that CG9920 knockdown severely disrupted mitochondrial morphogenesis during spermatogenesis. Notably, we found that CG9920 might not directly interact with Ocn, but rather was inhibited by STAT92E, which itself was indirectly affected by Ocn. We propose a possible novel pathway essential for spermatogenesis in D. melanogaster , whereby Ocn indirectly induces CG9920 expression, potentially counteracting its inhibition by the JAK-STAT signaling pathway. [Display omitted] • CG9920 knockdown in germline caused male sterility with no mature sperms. • Depletion of CG9920 resulted in scattered spermatid nuclear bundles. • CG9920 knockdown led to fewer elongation cones and almost no ICs. • CG9920 knockdown disrupted mitochondrial morphogenesis during spermatogenesis. • CG9920 might be regulated by Ocnus through the JAK-STAT pathway. [ABSTRACT FROM AUTHOR]

Simple Summary: This study identified the protein expression profiles of horse and donkey sperm and investigated the effect of sperm proteins on sperm viability. In short, we first assessed the viability of horse and donkey sperm, revealing higher viability in donkey sperm compared to horse sperm. Subsequently, 4D-DIA protein-sequencing technology was employed to identify 3436 proteins expressed in horse sperm and 3404 proteins expressed in donkey sperm, with 73 proteins specific to horse sperm, and 41 specific to donkey sperm. Further analysis established a correlation between sperm proteins and sperm viability. These findings are significant in elucidating the reproductive variances and evolutionary relationships between horses and donkeys. The reproductive performance of horse sperm and donkey sperm has been reported to differ. Sperm proteins play a crucial role in sperm viability and fertility. Although differences between species are known, no prior study has investigated disparities in the sperm proteome between horses and donkeys. Therefore, this study characterized and compared the sperm proteomes of horses and donkeys using 4D-DIA mass spectrometry technology. We identified 3436 proteins in horse sperm and 3404 proteins in donkey sperm. Of these, 3363 proteins were expressed in both horse and donkey sperm, with 73 proteins being specifically expressed in horse sperm, and 41 in donkey sperm. According to data analysis, donkeys exhibited a greater percentage of motility and progressive movement in straight-line sperm than horses, as well as lower percentages of static and slow sperm than horses. Joint analysis of the results from the horse and donkey sperm proteomes and their CEROS II-read parameters demonstrated a possible association between sperm proteins and their sperm viability patterns. These findings suggest that there are discrepancies in the expression levels and protein compositions of horse and donkey sperm and that certain specific proteins may be responsible for the differences in performance between these two species. [ABSTRACT FROM AUTHOR]