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Background Objectives Materials and methods Results Discussion and conclusion The potential impact of diabetes mellitus type 1 (DM1) on male fertility is currently poorly defined. Hyperglycaemia and insulin deficiency may affect spermatogenesis. Some evidence suggests that men with DM1 have a significant reduction in progressive sperm motility, sperm morphology and semen volume, without significant changes in sperm concentration and count, but definite data are lacking.To evaluate the impact of DM1 on clinical parameters related to male fertility and semen analysis.We compared a court of 42 male DM1 patients with 43 nondiabetic subjects overlapping in age and remaining clinical data in an observational case‐control study. All subjects underwent a comprehensive andrological reproductive evaluation, including medical history, physical examination, and semen analysis. We collected biochemical data in all patients with DM1, while diabetic patients with any alteration in semen parameters underwent sperm culture and scrotal ultrasound. In addition, all men completed the IIEF‐5 questionnaire (International Index of Erectile Function‐5) and the AMS (Aging Male Symptom score) questionnaire.Patients with DM1 had a higher prevalence of infertility, erectile dysfunction and worse semen parameters compared with controls. In particular, semen volume, total sperm count, and total and progressive sperm motility were significantly lower (p < 0.001, p = 0.003, p = 0.048, and p = 0.022 respectively). In addition, the rate of semen anti‐sperm antibody positivity, the AMS score and FSH levels were higher.Several mechanisms may contribute to these semen alterations in DM1 patients, such as oxidative damage to spermatogenesis, seminal infections and pelvic neurological changes. These data suggest that patients with DM1 should be counselled from an andrological‐reproductive point of view. [ABSTRACT FROM AUTHOR]

Background Objectives Material and methods Results Discussion and conclusion The WHO manual for basic semen analysis and ISO 23162 describe sperm morphology assessment as a standard part of semen analysis. Older studies showed a correlation between morphology results and (artificial) conception. In more recent studies this relationship was less apparent and there is more emphasis on sperm morphology as a marker for healthy spermatogenesis (and general male health). Meantime, many laboratories ceased morphology assessment, probably due to unfamiliarity with this paradigmatic shift and to technical difficulties in the assessment, like the interpretation of morphological criteria.The aim of this study was to identify morphological criteria with high variability in results in the Dutch External Quality Control (EQC) program.Over the period 2015–2020, a total of 72 photos of sperm cells along with dichotomous propositions based on 14 criteria as defined in WHO5 (2010) were distributed in the Dutch EQC program for semen analysis. The EQC results were evaluated for variability per criterion and for trends in time.Between 2015 and 2020, 40 to 60 laboratories assessed the photos. Criteria with low variability between participants were related to acrosomal vacuoles, excessive residual cytoplasm, and tail metrics. In contrast, head ovality, regularity of head and midpiece contours, and alignment of the major axis of the midpiece and head led to the highest variability in outcomes. In general, there was a slightly positive trend (lower variability) in time, except for the criteria with the highest variability (stable or declining trend).This study indicates that there are (high) variabilities in the interpretation of the morphological criteria, leading to inconsistent outcomes of morphology assessment. The results are discussed from the perspective of imperfections in definitions and examples of the criteria as given in the WHO manuals. [ABSTRACT FROM AUTHOR]

Precise regulation of chromosome dynamics in the germline is essential for reproductive success across species. Yet, the mechanisms underlying meiotic chromosomal events such as homolog pairing and chromosome segregation are not fully understood in many species. Here, we employ Oligopaint DNA FISH to investigate mechanisms of meiotic homolog pairing and chromosome segregation in the holocentric pantry moth, Plodia interpunctella, and compare our findings to new and previous studies in the silkworm moth, Bombyx mori, which diverged from P. interpunctella over 100 million years ago. We find that pairing in both Bombyx and Plodia spermatogenesis is initiated at gene-rich chromosome ends. Additionally, both species form rod shaped cruciform-like bivalents at metaphase I. However, unlike the telomere-oriented chromosome segregation mechanism observed in Bombyx, Plodia can orient bivalents in multiple different ways at metaphase I. Surprisingly, in both species we find that kinetochores consistently assemble at non-telomeric loci toward the centers of chromosomes regardless of where chromosome centers are located in the bivalent. Additionally, sister kinetochores do not seem to be paired in these species. Instead, four distinct kinetochores are easily observed at metaphase I. Despite this, we find clear end-on microtubule attachments and not lateral microtubule attachments connecting these separated kinetochores to the meiotic spindle. These findings challenge the classical view of segregation where paired, poleward-facing kinetochores are required for accurate homolog separation in meiosis I. Our studies here highlight the importance of exploring fundamental processes in non-model systems, as employing novel organisms can lead to the discovery of novel biology. Author summary: In order for genetic information to be accurately passed on from one generation to the next, maternal and paternal chromosome copies (homologs) must be separated from each other when making egg and sperm cells in a process called meiosis. To facilitate this equal division, homologous partners pair up from end to end, undergo an exchange of genetic information, and then large protein complexes called kinetochores are formed to enable chromosome segregation. Here, we investigate how homologs pair and how kinetochores form during the formation of sperm in two different moth species: Bombyx mori (the silkworm moth) and Plodia interpunctella (the pantry moth). Using a combination of microscopy approaches, we observe that homologs seem to pair via interactions at chromosome ends. Conversely, we found that kinetochores form at chromosome centers, away from chromosome ends. This is true even when chromosomes are folded into different configurations. These central kinetochores are functional and able to recruit microtubules, the structures that physically pull chromosomes apart for segregation. How chromosome centers are specified to be kinetochore-recruitment sites in meiosis, and what causes the formation of different chromosome folding configurations, are areas of future study. [ABSTRACT FROM AUTHOR]

Single gamete cell sequencing together with long-read sequencing can reliably produce chromosome-level phased genomes. In this study, we employed PacBio HiFi and Hi-C sequencing on a male Landrace pig, coupled with single-sperm sequencing of its 102 sperm cells. A haplotype assembly method was developed based on long-read sequencing and sperm-phased markers. The chromosome-level phased assembly showed higher phasing accuracy than methods that rely only on HiFi reads. The use of single-sperm sequencing data enabled the construction of a genetic map, successfully mapping the sperm motility trait to a specific region on chromosome 1 (105.40–110.70 Mb). Furthermore, with the assistance of Y chromosome-bearing sperm data, 26.16 Mb Y chromosome sequences were assembled. We report a reliable approach for assembling chromosome-level phased genomes and reveal the potential of sperm population in basic biology research and sperm phenotype research. An improved pig genome assembly is reported, using single sperm sequencing to reconstruct chromosome haplotypes, and with improved Y chromosome sequence. [ABSTRACT FROM AUTHOR]

In internal fertilisers, the precise timing of ovulation with the arrival of sperm at the site of fertilisation is essential for fertilisation success. In birds, mating is often not synchronised with ovulation, but instead females utilise specialised sperm storage tubules (SSTs) in the reproductive tract, which can ensure sperm are always available for fertilisation at the time of ovulation, whilst simultaneously providing a mechanism of post‐copulatory sexual selection. Despite the clear importance of SSTs for fertilisation success, we know little about the mechanisms involved in sperm acceptance, storage, and release. Furthermore, most research has been conducted on only a small number of species, based on which SSTs are usually assumed to look and function in the same way across all species. Here, we conduct a comparative exploration of SST morphology across 26 species of Galliformes. We show that SSTs, and the surrounding tissue, can vary significantly in morphology across species. We provide observational evidence that Galliformes exhibit at least 5 distinct categories of tubule types, including distinctive coiled and multi‐branched tubules, and describe 2 additional features of the surrounding tissue. We suggest functional explanations for variation in tubule morphology and propose next steps for future research. Our findings indicate that SSTs are likely to be far more variable than has previously been assumed, with potentially important consequences for our understanding of sperm storage in birds and post‐copulatory sexual selection in general. [ABSTRACT FROM AUTHOR]

In this cross-sectional prospective study, advanced next-generation sequencing technology was used to compare the molecular karyotyping of individual human sperm cells in infertile couples with severe oligoteratozoospermia (i.e., low sperm count and motility) to those of infertile couples with normal semen. Fourteen infertile couples who were patients at Ramathibodi Hospital in Bangkok, Thailand, were recruited from January to November 2023, and they were categorized into two groups based on semen analysis results. The study group comprised couples with severe oligoteratozoospermia, whereas the control group exhibited normal semen. Individual sperm cells from the semen samples were isolated by the micromanipulation technique for subsequent whole-genome amplification and next-generation sequencing, where the primary outcome was the aneuploidy rate. Seventy individual sperm cells were isolated with a 90% success rate for amplification. The next-generation sequencing results showed that the aneuploidy rate was 25%–75%, with a mean of 48.28% in the study group. In contrast, the control group exhibited aneuploidy rates of 0–75%, with a mean of 15.15%. The difference between the two groups was statistically significant (odds ratio: 5.8, 95% confidence interval: 1.30–26.03). Sperm cells of the study group showed a threefold higher aneuploidy rate than those in the control group, even though the sperm cells were selected by micromanipulation for their normal morphology. Comprehensive counseling is recommended to address elevated aneuploidy rates that potentially surpass those of the general infertile population. Guidance on preimplantation genetic testing is also recommended to ensure the transfer of embryos with normal chromosomes. [ABSTRACT FROM AUTHOR]

Odad3 gene loss-of-function mutation leads to Primary Ciliary Dyskinesia (PCD), a disease caused by motile cilia dysfunction. Previously, we demonstrated that knockout of the Odad3 gene in mice replicates several features of PCD, such as hydrocephalus, defects in left–right body symmetry, and male infertility, with a complete absence of sperm in the reproductive tract. The majority of Odad3 knockout animals die before sexual maturation due to severe hydrocephalus and failure to thrive, which precludes fertility studies. Here, we performed the expression analysis of the Odad3 gene during gonad development and in adult testes. We showed that Odad3 starts its expression during the first wave of spermatogenesis, specifically at the meiotic stage, and that its expression is restricted to the germ cells in the adult testes, suggesting that Odad3 plays a role in spermatozoa formation. Subsequently, we conditionally deleted the Odad3 gene in adult males and demonstrated that even partial ablation of the Odad3 gene leads to asthenoteratozoospermia with multiple morphological abnormalities of sperm flagella (MMAF) in mice. The analysis of the seminiferous tubules in Odad3-deficient mice revealed defects in spermatogenesis with accumulation of seminiferous tubules at the spermiogenesis and spermiation phases. Furthermore, analysis of fertility in heterozygous Odad3+/− knockout mice revealed a reduction in sperm count and motility as well as abnormal sperm morphology. Additionally, Odad3+/− males exhibited a shorter fertile lifespan. Overall, these results suggest the important role of Odad3 and Odad3 gene dosage in male fertility. These findings may have an impact on the genetic and fertility counseling practice of PCD patients carrying Odad3 loss-of-function mutations. [ABSTRACT FROM AUTHOR]

This comprehensive review explores the evolving landscape of sperm selection techniques within the realm of Assisted Reproductive Technology (ART). Our analysis delves into a range of methods from traditional approaches like density gradient centrifugation to advanced techniques such as Magnetic-Activated Cell Sorting (MACS) and Intracytoplasmic Morphologically Selected Sperm Injection (IMSI). We critically assess the efficacy of these methods in terms of sperm motility, morphology, DNA integrity, and other functional attributes, providing a detailed comparison of their clinical outcomes. We highlight the transition from conventional sperm selection methods, which primarily focus on physical characteristics, to more sophisticated techniques that offer a comprehensive evaluation of sperm molecular properties. This shift not only promises enhanced prediction of fertilization success but also has significant implications for improving embryo quality and increasing the chances of live birth. By synthesizing various studies and research papers, we present an in-depth analysis of the predictability of different sperm selection procedures in ART. The review also discusses the clinical applicability of these methods, emphasizing their potential in shaping the future of assisted reproduction. Our findings suggest that the integration of advanced sperm selection strategies in ART could lead to more cost-effective treatments with reduced duration and higher success rates. This review aims to provide clinicians and researchers in reproductive medicine with comprehensive insights into the current state and future prospects of sperm selection technologies in ART. [ABSTRACT FROM AUTHOR]

At present, liquid storage is the most efficient method for pig semen preservation. This approach relies upon reducing sperm metabolism, allowing for the maintenance of cell lifespan. In this context, the study of proteins that could protect sperm during liquid storage is of high relevance. The 70 kDa Heat Shock Protein (HSP70) is an anti-apoptotic protein that has been reported to be relevant to sperm survival. Thus, we explored the role of HSP70 during prolonged storage of pig semen at 17 °C. Six semen pools were incubated with YM-1 (0, 0.05, 0.1 and 0.2 μM), an HSP70 inhibitor, and stored at 17 °C for 21 days. On days 0, 4, 10, 14 and 21, sperm quality and function were evaluated through flow cytometry and Computer-Assisted Sperm Analysis (CASA), and HSP70 activity and chromatin condensation were also determined. While inhibition of HSP70 increased progressive motility, Ca2+ and Reactive Oxygen Species (ROS) levels, and mitochondrial activity during the first 10 days of storage, it had a detrimental effect on sperm motility after 14 and 21 days. In spite of this, sperm viability was not altered. We can conclude that HSP70 contributes to the liquid storage of pig semen because it keeps mitochondrial activity low, which is needed for the maintenance of sperm function. [ABSTRACT FROM AUTHOR]

Sperm-associated antigen 5 (SPAG5) regulates cancer cell invasion and is involved in the progression of many cancers. However, the role of SPAG5 in endometrial carcinoma (EC) is still unknown. The purpose of this study was to explore the role of SPAG5 in EC and its potential molecular mechanism. The UALCAN tool and cBioPortal were used to analyze the expression and alterations of SPAG5 in EC, respectively. OncoLnc was used for survival analysis. We analyzed the effects of SPAG5 on immune cell infiltration and the expression levels of immune checkpoints. We also overexpressed and knocked down SPAG5 in EC cells to explore the effect of SPAG5 regulation on migration, invasion, apoptosis, and the cell cycle of EC cells. We found that SPAG5 was overexpressed and the SPAG5 gene was often mutated in EC. High SPAG5 expression was significantly associated with poor overall survival in patients with EC. SPAG5 also affected the level of immune cell infiltration in the TIME and the expression of immune checkpoints lymphocyte activating 3 (LAG3) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) in patients with EC. It may also be involved in the immunotherapy response in these patients. In vitro experiments showed that SPAG5 promotes cancer cell migration and invasion. In conclusion, this study lays the foundation for further understanding the molecular mechanisms of EC involving SPAG5 and contributes to diagnosing and managing this disease. [ABSTRACT FROM AUTHOR]

Preserving boar semen at 5°C instead of the conventional storage temperature of 17°C would enable a reduction of antibiotic use in pig insemination. To protect the chilling-sensitive boar spermatozoa, holding the extended semen at a higher temperature before cooling could be beneficial and facilitate the implementation of the innovative preservation concept in practice, provided that bacterial growth is kept at a low level. The aim of this study was to introduce a holding time (HT) at 17°C before cooling and to examine the effect on sperm quality and bacterial growth compared to the original cooling protocol for antibiotic-free 5°C semen storage. A series of experiments with semen doses from eight boars extended in Androstar® Premium without conventional antibiotics revealed that sperm kinematics and the integrity of sperm plasma membranes and acrosomes were improved with HT between 16 and 24 h followed by delayed cooling with 0.04°C/min when compared to the original protocol for semen preservation at 5°C (p < 0.05). Both a shorter HT of 6 h and a faster cooling rate of 0.07°C/min reduced sperm quality (p < 0.05). The HT for 24 h did not compromise the inhibitory effect on bacterial growth during long-term semen storage at 5°C, not even in semen doses spiked with Serratia marcescens. In conclusion, semen storage at 5°C with the modified cooling protocol improved sperm quality and is antimicrobially efficient. It thus presents a ready-to-use tool for a reduction or replacement of antibiotics in pig insemination. [ABSTRACT FROM AUTHOR]

Background: The issue of male fertility is becoming increasingly common due to genetic differences inherited over generations. Gene expression and evaluation of non-coding RNA (ncRNA), crucial for sperm development, are significant factors. This gene expression can affect sperm motility and, consequently, fertility. Understanding the intricate protein interactions that play essential roles in sperm differentiation and development is vital. This knowledge could lead to more effective treatments and interventions for male infertility. Materials and methods: Our research aim to identify new and key genes and ncRNA involved in non-obstructive azoospermia (NOA), improving genetic diagnosis and offering more accurate estimates for successful sperm extraction based on an individual's genotype. Results: We analyzed the transcript of three NOA patients who tested negative for genetic sperm issues, employing comprehensive genome-wide analysis of approximately 50,000 transcript sequences using microarray technology. This compared gene expression profiles between NOA sperm and normal sperm. We found significant gene expression differences: 150 genes were up-regulated, and 78 genes were down-regulated, along with 24 ncRNAs up-regulated and 13 ncRNAs down-regulated compared to normal conditions. By cross-referencing our results with a single-cell genomics database, we identified overexpressed biological process terms in differentially expressed genes, such as "protein localization to endosomes" and "xenobiotic transport." Overrepresented molecular function terms in up-regulated genes included "voltage-gated calcium channel activity," "growth hormone-releasing hormone receptor activity," and "sialic acid transmembrane transporter activity." Analysis revealed nine hub genes associated with NOA sperm: RPL34, CYB5B, GOL6A6, LSM1, ARL4A, DHX57, STARD9, HSP90B1, and VPS36. Conclusions: These genes and their interacting proteins may play a role in the pathophysiology of germ cell abnormalities and infertility. [ABSTRACT FROM AUTHOR]

Background: The involvement of ATP and cAMP in sperm function has been extensively documented, but the understanding of the role of adenosine and adenosine receptors remains incomplete. This study aimed to examine the presence of adenosine A2A receptor (A2AR) and study the functional role of A2AR in human sperm. Methods: The presence and localization of A2AR in human sperm were examined by western blotting and immunofluorescence assays. The functional role of A2AR in sperm was assessed by incubating human sperm with an A2AR agonist (regadenoson) and an A2AR antagonist (SCH58261). The sperm level of A2AR was examined by western blotting in normozoospermic and asthenozoospermic men to evaluate the association of A2AR with sperm motility and in vitro fertilization (IVF) outcomes. Results: A2AR with a molecular weight of 43 kDa was detected in the tail of human sperm. SCH58261 decreased the motility, penetration ability, intracellular Ca2+ concentration, and CatSper current of human sperm. Although regadenoson did not affect these sperm parameters, it alleviated the adverse effects of SCH58261 on these parameters. In addition, the mean level of A2AR in sperm from asthenozoospermic men was lower than that in sperm from normozoospermic men. The sperm level of A2AR was positively correlated with progressive motility. Furthermore, the fertilization rate during IVF was lower in men with decreased sperm level of A2AR than in men with normal sperm level of A2AR. Conclusions: These results indicate that A2AR is important for human sperm motility and is associated with IVF outcome. [ABSTRACT FROM AUTHOR]

Background: Asthenozoospermia, a type of male infertility, is primarily caused by dysfunctional sperm mitochondria. Despite previous bioinformatics analysis identifying potential key lncRNAs, miRNAs, hub genes, and pathways associated with asthenospermia, there is still a need to explore additional molecular mechanisms and potential biomarkers for this condition. Methods: We integrated data from Gene Expression Omnibus (GEO) (GSE22331, GSE34514, and GSE160749) and performed bioinformatics analysis to identify differentially expressed genes (DEGs) between normozoospermia and asthenozoospermia. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to gain insights into biological processes and signaling pathways. Weighted Gene Co-expression Network Analysis (WGCNA) identified gene modules associated with asthenozoospermia. Expression levels of key genes were assessed using datasets and experimental data. Gene Set Enrichment Analysis (GSEA) and correlation analysis identified pathways associated with the hub gene and explore the relationship between the ZNF764 and COQ9 and mitochondrial autophagy-related genes. Competitive endogenous RNA (ceRNA) networks were constructed, and in vitro experiments using exosome samples were conducted to validate this finding. Results: COQ9 was identified as a marker gene in asthenozoospermia, involved in autophagy, ATP-dependent chromatin remodeling, endocytosis, and cell cycle, etc. The ceRNA regulatory network (LINC00893/miR-125a-5p/COQ9) was constructed, and PCR demonstrated that LINC00893 and COQ9 were downregulated in asthenozoospermia, while miR-125a-5p and m6A methylation level of LINC00893 were upregulated in asthenozoospermia compared to normozoospermic individuals. Conclusion: The ceRNA regulatory network (LINC00893/miR-125a-5p/COQ9) likely plays a crucial role in the mechanism of asthenozoospermia. However, further functional experiments are needed to fully understand its significance. [ABSTRACT FROM AUTHOR]

This study is the first to identify bovine blastocysts through in vitro fertilization (IVF) of matured oocytes with a large quantity of highquality sperm separated from a biomimetic cervix environment. We obtained high-quality sperm in large quantities using an IVF sperm sorting chip (SSC), which could mimic the viscous environment of the bovine cervix during ovulation and facilitates isolation of progressively motile sperm from semen. The viscous environment-on-a-chip was realized by formulating and implementing polyvinylpyrrolidone (PVP)-based solutions for the SSC medium. Sperm separated from the IVF-SSC containing PVP 1.5% showed high motility, normal morphology and high DNA integrity. As a result of IVF, a higher rate of hatching blastocysts, which is the pre-implantation stage, were observed, compared to the conventional swim-up method. Our results may significantly contribute to improving livestock with superior male and female genetic traits, thus overcoming the limitation of artificial insemination based on the superior genetic traits of existing males. [ABSTRACT FROM AUTHOR]

Recent studies have shifted the spotlight from adult disease to gametogenesis and embryo developmental events, and these are greatly affected by various environmental chemicals, such as drugs, metabolites, pollutants, and others. Growing research has highlighted the critical importance of identifying and understanding the roles of chemicals in reproductive biology. However, the functions and mechanisms of chemicals in reproductive processes remain incomplete. We developed a comprehensive database called the Reproductive Chemical Database (RCDB) (https://yu.life.sjtu.edu.cn/ChenLab/RCDB) to facilitate research on chemicals in reproductive biology. This resource is founded on rigorous manual literature extraction and precise protein target prediction methodologies. This database focuses on the delineation of chemicals associated with phenotypes, diseases, or endpoints intricately associated with four important reproductive processes: female and male gamete generation, fertilization, and embryo development in human and mouse. The RCDB encompasses 93 sub-GO processes, and it revealed 1447 intricate chemical–biological process interactions. To date, the RCDB has meticulously cataloged and annotated 830 distinct chemicals, while also predicting 614 target proteins from a selection of 3800 potential candidates. Additionally, the RCDB offers an online predictive tool that empowers researchers to ascertain whether specific chemicals play discernible functional roles in these reproductive processes. The RCDB is an exhaustive, cross-platform, manually curated database, which provides a user-friendly interface to search, browse, and use reproductive processes modulators and their comprehensive related information. The RCDB will help researchers to understand the whole reproductive process and related diseases and it has the potential to promote reproduction research in the pharmacological and pathophysiological areas. [ABSTRACT FROM AUTHOR]

Despite efforts to explore the genome of the malaria vector Anopheles gambiae, the Y chromosome of this species remains enigmatic. The large number of repetitive and heterochromatic DNA sequences makes the Y chromosome exceptionally difficult to fully assemble, hampering the progress of gene editing techniques and functional studies for this chromosome. In this study, we made use of a bioinformatic platform to identify Y-specific repetitive DNA sequences that served as a target site for a CRISPR/Cas9 system. The activity of Cas9 in the reproductive organs of males caused damage to Y-bearing sperm without affecting their fertility, leading to a strong female bias in the progeny. Cytological investigation allowed us to identify meiotic defects and investigate sperm selection in this new synthetic sex ratio distorter system. In addition, alternative promoters enable us to target the Y chromosome in specific tissues and developmental stages of male mosquitoes, enabling studies that shed light on the role of this chromosome in male gametogenesis. This work paves the way for further insight into the poorly characterised Y chromosome of Anopheles gambiae. Moreover, the sex distorter strain we have generated promises to be a valuable tool for the advancement of studies in the field of developmental biology, with the potential to support the progress of genetic strategies aimed at controlling malaria mosquitoes and other pest species. Author summary: Genetic elements known as sex ratio meiotic drive can manipulate the sex ratio of offspring, favouring the male or female sex. This fascinating phenomenon has inspired the development of synthetic sex ratio distorter systems in several organisms. In species where females and males harbour XX and XY sex chromosomes respectively, the X-chromosome can be 'shredded' during male gametogenesis, leading to the production of non-functional X-bearing sperm, while Y-bearing sperm are left intact and able to fertilise the eggs. These systems can produce offspring that are extremely biased towards males, which can be used as genetic tools to control harmful insect populations. In our study, we applied this molecular strategy to target the Y chromosome of Anopheles gambiae. Our aim was to investigate the cellular consequences of the shredding of this chromosome, the impact on meiosis and sperm selection, and the potential to achieve strong female bias in the offspring. The outcome of this study enhances our understanding of the molecular and biological mechanisms behind synthetic sex-ratio distorters in Anopheles mosquitoes, which could inform the development of vector control strategies that target sex ratio. Additionally, we present a genetic sexing strain able to produce mostly females, providing a valuable genetic tool for fundamental studies on this deadly vector. [ABSTRACT FROM AUTHOR]

Background: Several previous animal and human studies have found a strong association between asthma and spermatozoa quality, but whether these associations are causal or due to bias remains to be elucidated. Methods: We performed a two-sample Mendelian randomization (MR) analysis to assess the causal effect of genetically predicted asthma on the risk of abnormal spermatozoa. Asthma, childhood-onset asthma (COA), and adult-onset asthma (AOA) (sample sizes ranging from 327,670 to 408,442) were included as the exposures. Genetic information for abnormal spermatozoa was obtained from a genome-wide association study (GWAS) comprising 209,921 participants. In univariable MR (UVMR) analysis, the inverse variance weighted (IVW) method was conducted as the primary method, with the MR Egger and weighted median used as supplementary methods for causal inference. Sensitivity analyses, including the Cochran Q test, Egger intercept test, MR-PRESSO, and leave-one-out analysis, were performed to verify the robustness of the MR results. Multivariable MR (MVMR) was conducted to evaluate the direct causal effects of asthma on abnormal spermatozoa risk. Results: UVMR detected causal associations between genetically predicted asthma and an increased risk of abnormal spermatozoa (OR: 1.270, 95% CI: 1.045-1.545, p = 0.017). Moreover, we found that AOA (OR: 1.46, 95% CI: 1.051, 2.018, p = 0.024) has positive causal effects on the risk of abnormal spermatozoa rather than COA (p = 0.558). Sensitivity analysis found little evidence of bias in the current study (p > 0.05). MVMR further confirmed that asthma directly affected the risk of abnormal spermatozoa. Conclusion: Our MR study suggested that genetically predicted asthma could be associated with an increased risk of abnormal spermatozoa, and similar results were obtained in AOA. Further studies are warranted to explain the underlying mechanisms of this association and may provide new avenues for prevention and treatment. [ABSTRACT FROM AUTHOR]

To understand the seasonality of spermatogenesis in cultured males of Mytilus edulis from a cold‐ocean environment, we investigated the cellular transitions occurring within the spermatogenic epithelium of the testicular acini during early and advancing spermatogenesis, with specific reference to the histology of the epithelium, gene specific spermatogenic response, condition, culture environment, and season. A combination of histological evaluation, qPCR analysis, and in situ hybridization was used to examine the cellular transitions taking place in the germinal epithelium from late winter through to a seasonal spawn in summer. We observed clear seasonal transitionary changes in the spermatogenic cell population making up the germinal epithelium (i.e., spermatogonial stem cells, spermatocytes, spermatids, and spermatozoa) extending from February to July. These seasonal transitions in spermatogenic cell type coincided with significant variation in the spatiotemporal expression of two molecular markers for spermatogenesis (i.e., Kelch‐like protein 10 [KLHL10] and Armadillo repeat‐containing protein 4 isoform X2 [ARMC4]) but not for expression of a gamete‐specific Mitochondrial cytochrome oxidase I (MT‐COI). The spatiotemporal expression of these genes is directly linked to the cellular changes taking place in the germinal epithelium during spermatogenesis. These observations not only corresponded to seasonal changes in physiological condition but also environmental temperature and chlorophyll a, thus further supporting the link between male gametogenesis and environment in higher latitude regions. [ABSTRACT FROM AUTHOR]

In most ectotherms, several life‐history traits, including body size, respond to environmental conditions through the temperature–size rule (TSR). The mechanisms underlying the TSR are still being debated, but studying idiobiont insect parasitoids, which develop with a fixed amount of resources, may shed light on this relationship. In this study, we conducted experiments to determine how the developmental temperature affects various characteristics of male and female Trichogramma euproctidis (Girault) (Hymenoptera: Trichogrammatidae), an idiobiont egg parasitoid of Lepidoptera. The study tested various hypotheses, including the cellular or oxygen diffusion hypotheses and the resource acquisition hypothesis, to understand whether T. euproctidis follows the TSR. The developmental time of both male and female T. euproctidis decreased with increasing temperature. Both males and females displayed a unimodal distribution for size, dry mass, and lipid content, with individuals at lower and higher temperatures being smaller, weighing less, and containing fewer lipids. Female lifetime fecundity increased from 13 to 24 °C and then decreased at 31 °C. Additionally, the number and size of gametes in male and female T. euproctidis displayed a unimodal distribution with increasing temperature. Trichogramma euproctidis deviates from the TSR as it follows a non‐linear reaction norm with an optimal developmental temperature. This result supports the hypothesis that for species following TSR and having unlimited access to food resources, the resource acquisition hypothesis is a significant mechanism explaining the TSR. With climate change affecting temperature, understanding the TSR is crucial, and research on insect parasitoids may help reveal how the interplay between environmental temperature and resource allocation affects the TSR in natural populations. [ABSTRACT FROM AUTHOR]

This pioneering study aimed to evaluate the cryopreservation of semen from P. falkneri (n = 4) and P. motoro (n = 4), maintained ex situ at the Sao Paulo Aquarium, Brazil. For this purpose, the animals were physically restrained, biometric data of the disc and clasper were obtained, and semen was collected through manual massage. Total motility and progressive motility parameters were evaluated using Computer-Assisted Sperm Analysis (CASA) with IVOS II equipment and Animal Breeders II software. The semen extenders INRA 96 and OptiXcell were used to assess their efficacy in sperm cryopreservation. INRA required the addition of 5% dimethyl sulfoxide (DMSO) as a cryoprotectant. The results indicated that there was no difference in semen motility values before and after freezing with INRA + DMSO (p = 0.6226). On the other hand, samples cryopreserved with OptiXcell showed a difference in semen motility post-thaw (p = 0.0156). These findings contribute to a broader study on optimizing cryopreservation protocols to ensure long-term viability and fertility of semen, enhancing genetic diversity and supporting wild population restoration. A multidisciplinary approach integrating reproductive biology, ecology, physiology, and assisted reproduction technologies, aligned with the One Conservation concept, is essential for advancing conservation and management strategies for these threatened species. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to investigate the effect of Codonopsis pilosula polysaccharide (CPP) on the motility, mitochondrial integrity, acrosome integrity rate, and antioxidant ability of sheep sperm after preservation at 4°C. Methods: Semen from healthy adult rams were collected and divided into four groups with separate addition of 0, 200, 400, and 1,000 mg/L CPP. Sperm motility was analyzed using the Computer-Assisted Semen Analysis software after preservation at 4°C for 24, 72, 120, and 168 h. Sperm acrosome integrity rate was analyzed by Giemsa staining at 24, 72, and 120 h, and mitochondrial membrane integrity was analyzed by Mito-Tracker Red CMXRos. The total antioxidant capacity (T-AOC) and malondialdehyde (MDA) content of spermatozoa were measured after 120 h of preservation. Results: The sperm viability and forward-moving sperm under 200 mg/L CPP were significantly higher than that in the control group at 72 h (61.28%±3.89% vs 52.83%± 0.70%, 51.53%±4.06% vs 42.84%±1.14%), and 168 h (47.21%±0.85% vs 41.43%±0.37%, 38.68%±0.87% vs 31.68%±0.89%). The percentage of fast-moving sperm (15.03%±1.10% vs 11.39%±1.03%) and slow-moving sperm (23.63%±0.76% vs 20.29%±1.11%) in the 200 mg/L group was significantly higher than control group at 168 h. The mitochondrial membrane integrity of the sperm in the group with 200 mg/L CPP was significantly higher than those in the control group after storage at 4°C for 120 h (74.76%±2.54% vs 65.67% ±4.51%, p<0.05). The acrosome integrity rate in the group with 200 mg/L (87.66%±1.26%) and 400 mg/L (84.00%±2.95%) was significantly higher than those in the control group (80.65%±0.16%) after storage for 24 h (p<0.05). CPP also increased T-AOC and decreased the MDA concentration after preservation at 4°C (p<0.05). Conclusion: Adding CPP could improve the T-AOC of sperm, inhibit lipid peroxidation, and facilitate semen preservation. [ABSTRACT FROM AUTHOR]

The in vitro storage of stallion spermatozoa for use in artificial insemination leads to oxidative stress and imbalances in calcium homeostasis that trigger the formation of the mitochondrial permeability transition pore (mPTP), resulting in premature cell death. However, little is understood about the dynamics and the role of mPTP formation in mammalian spermatozoa. Here, we identify an important role for mPTP in stallion sperm Ca2+ homeostasis. We show that stallion spermatozoa do not exhibit "classical" features of mPTP; specifically, they are resistant to cyclosporin A‐mediated inhibition of mPTP formation, and they do not require exogenous Ca2+ to form the mPTP. However, chelation of endogenous Ca2+ prevented mPTP formation, indicating a role for intracellular Ca2+ in this process. Furthermore, our findings suggest that this cell type can mobilize intracellular Ca2+ stores to form the mPTP in response to low Ca2+ environments and that under oxidative stress conditions, mPTP formation preceded a measurable increase in intracellular Ca2+, and vice versa. Contrary to previous work that identified mitochondrial membrane potential (MMP) as a proxy for mPTP formation, here we show that a loss of MMP can occur independently of mPTP formation, and thus MMP is not an appropriate proxy for the detection of mPTP formation. In conclusion, the mPTP plays a crucial role in maintaining Ca2+ and reactive oxygen species homeostasis in stallion spermatozoa, serving as an important regulatory mechanism for normal sperm function, thereby contraindicating the in vitro pharmacological inhibition of mPTP formation to enhance sperm longevity. [ABSTRACT FROM AUTHOR]

The study investigated if gonad maturation in triploid brown trout, Salmo trutta, was entirely suppressed or only delayed, and if triploids could interbreed with diploid counterparts. Ten percent of the total number of 3‐year‐old triploid S. trutta, 15% of 4‐year‐old fish, and 17% of 5‐year‐old fish produced semen. Three and 4 years old triploid fish did not produce eggs, but 15% of the 5‐year‐old fish did so. The quantity and sperm motility of triploid semen did not differ from diploids, but the sperm concentration was significantly lower. When diploid eggs were fertilized with triploid semen (3n × 2n crosses), the percentage of eyed stage embryos, of hatched larvae, and of normal‐shaped larvae did not differ from the diploid controls. Circa 90% of 3n × 2n crosses had a ploidy level of 2.4n. In the remaining percentage of 3n × 2n crosses, the ploidy level was ≥2n and <2.4n. In sperm competition experiments where diploid eggs were fertilized with a mixture of diploid and triploid semen, 52% of the originating larvae had a ploidy level of 2n, 43% of 2.4n, and 5% of the fish were not exactly classified. From the start of feeding to an age of 248 days, the mortality rate of 3n × 2n interploid crosses and of 2n × 2n controls was similar. The growth of interploid crosses was significantly higher than that of controls. In triploid mature females, the egg mass per kilogram of body weight was significantly lower than in diploids. The mass of the non‐hardened eggs and the percentile weight increase during hardening did not differ from diploid eggs. When triploid eggs were fertilized with diploid semen (2n × 3n crosses), the development rate to normal hatched larvae was less than 10%. All originating larvae had a ploidy level of 3n. From the start of feeding to an age of 248 days, 2n × 3n crosses had a higher mortality rate (15%) than diploid controls (<5%). Growth of this type of interploid crosses was reduced in comparison to controls. Therefore, triploids introduced into natural waters for recreational fisheries or escaping from farms may interbreed with diploid counterparts. This not only alters the genotypes of local populations but also changes the ploidy levels. [ABSTRACT FROM AUTHOR]

pH homeostasis is crucial for spermatogenesis, sperm maturation, sperm physiological function, and fertilization in mammals. HCO3− and H+ are the most significant factors involved in regulating pH homeostasis in the male reproductive system. Multiple pH-regulating transporters and ion channels localize in the testis, epididymis, and spermatozoa, such as HCO3− transporters (solute carrier family 4 and solute carrier family 26 transporters), carbonic anhydrases, and H+-transport channels and enzymes (e.g., Na+-H+ exchangers, monocarboxylate transporters, H+-ATPases, and voltage-gated proton channels). Hormone-mediated signals impose an influence on the production of some HCO3− or H+ transporters, such as NBCe1, SLC4A2, MCT4, etc. Additionally, ion channels including sperm-specific cationic channels for Ca2+ (CatSper) and K+ (SLO3) are directly or indirectly regulated by pH, exerting specific actions on spermatozoa. The slightly alkaline testicular pH is conducive to spermatogenesis, whereas the epididymis's low HCO3− concentration and acidic lumen are favorable for sperm maturation and storage. Spermatozoa pH increases substantially after being fused with seminal fluid to enhance motility. In the female reproductive tract, sperm are subjected to increasing concentrations of HCO3− in the uterine and fallopian tube, causing a rise in the intracellular pH (pHi) of spermatozoa, leading to hyperpolarization of sperm plasma membranes, capacitation, hyperactivation, acrosome reaction, and ultimately fertilization. The physiological regulation initiated by SLC26A3, SLC26A8, NHA1, sNHE, and CFTR localized in sperm is proven for certain to be involved in male fertility. This review intends to present the key factors and characteristics of pHi regulation in the testes, efferent duct, epididymis, seminal fluid, and female reproductive tract, as well as the associated mechanisms during the sperm journey to fertilization, proposing insights into outstanding subjects and future research trends. [ABSTRACT FROM AUTHOR]

Simple Summary: Boar sperm is a highly sensitive biological product, and its quality is susceptible to various environmental factors during transportation. Presently, vibration emissions were observed and documented during the transportation of artificial insemination doses. Vibrations, serving as an environmental stressor, harbor the potential to exert an influence on sperm quality. However, the precise mechanism underlying the detrimental effects of vibration emissions on sperm functionality remains elusive. In the present research, we discerned that vibration emissions impair the quality of boar sperm by modulating their metabolic processes, ultimately resulting in a diminished fertilization capability. Artificial insemination (AI) with liquid-preserved semen has recently become common in pig breeding. The semen doses are produced in a centralized manner at the boar stud and then subsequently distributed and transported to sow farms. However, vibration emissions during transportation by logistic vehicles may adversely affect the quality of boar sperm. Therefore, this study aimed to explore the impact of vibration-induced emissions on sperm quality and function under simulated transportation conditions. Each time, ejaculates from all 15 boars were collected and then pooled together to minimize individual variations, and the sample was split using an extender for dilution. Different rotational speeds (0 rpm, 80 rpm, 140 rpm, 200 rpm) were utilized to simulate varying intensities of vibration exposure using an orbital shaker, considering different transportation times (0 h, 3 h, and 6 h). Subsequently, evaluations were conducted regarding sperm motility, plasma membrane integrity, acrosome integrity, mitochondrial function, adenosine triphosphate (ATP) levels, mitochondrial reactive oxygen species (ROS) levels, pH, glycolytic pathway enzyme activities, and capacitation following exposure to vibration emissions. Both vibration time and intensity impact sperm motility, plasma membrane integrity, and acrosomal integrity. Vibration exposure significantly reduced sperm ATP levels, mitochondrial membrane potential, and the levels of mitochondria-encoded proteins (MT-ND1, MT-ND6) (p < 0.05). After vibration emission treatment, the pH value and mitochondrial ROS levels significantly increased (p < 0.05). Inhibition of sperm glycolysis was observed, with reduced activities of hexokinase (HK), pyruvate kinase (PK), and lactate dehydrogenase (LDH), along with decreased lactate levels (p < 0.05). Additionally, sperm tyrosine phosphorylation levels were significantly reduced by vibration emissions compared to the control group (p < 0.05). After the vibration emission treatment, the number of sperm bound to each square millimeter of oviduct explants decreased significantly compared to the control group (p < 0.05). Similarly, compared to the control group, using semen subjected to vibration stress for AI results in significantly reduced pregnancy rates, total born litter size, live-born litter size, and healthy born litter size (p < 0.05). [ABSTRACT FROM AUTHOR]

There has been emerging research linking per- and poly-fluoroalkyl substances (PFAS) to gamete viability and fertility. PFAS, prevalent in the environment and water supplies, undergo slow degradation due to their C-F bond and a long half-life (2.3–8.5 years). In females, PFAS inhibit the hypothalamic–pituitary–gonadal (HPG) axis, reducing follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels, leading to the inhibition of androgen and estradiol production. PFAS have been found to cause detrimental effects on egg quality through impairing folliculogenesis. In males, PFAS can impair sperm motility and morphology: two fundamental qualities of successful fertilization. PFAS exposure has been proven to inhibit testosterone production, sperm capacitation, and acrosomal reaction. After fertilization, the results of PFAS exposure to embryos have also been investigated, showing reduced development to the blastocyst stage. The aim of this review is to report the main findings in the literature on the impact of PFAS exposure to gamete competency and fertilization capability by highlighting key studies on both male and female fertility. We report that there is significant evidence demonstrating the negative impacts on fertility after PFAS exposure. At high doses, these environmentally abundant and widespread compounds can significantly affect human fertility. [ABSTRACT FROM AUTHOR]

Context: Collecting, cooling, and cryopreserving semen is essential for accessing genetically superior stallions. However, preserving stallion sperm presents unique challenges compared with other species. Aims: This study aimed to investigate the effect of zinc (Zn) supplementation on the quality of equine sperm during cold storage. Methods: Various factors contributing to sperm quality were assessed at 24 and 48 h after cooling, and after subjecting the sperm samples to a heat-resistance test. In Experiment 1, four experimental groups were examined, each with different concentrations of Zn sulfate, as follows: 0, 1, 2, and 3 mM. Subsequently, Experiment 2 involved testing a wider range of concentrations (0, 0.1, 0.2, 0.4, 0.8, 1.6, and 3.2 mM), including evaluation of samples after incubation for 240 min at 37°C (heat-resistance test). Key results: The addition of different concentrations of Zn to the extender INRA96 did not yield substantial improvements in sperm-quality parameters for cooling stallion semen after 24 or 48 h. Moreover, no protective benefits were observed when samples underwent a heat-resistance test. Concentrations of Zn surpassing 3 mM had an adverse effect on sperm-quality parameters. Conclusions and implications: These findings have contributed to the understanding of Zn supplementation as a strategy for improving semen preservation in stallions. This study explores the impact of zinc supplementation to improve the quality of cooled equine semen, addressing a crucial aspect of genetic preservation in horses. The findings showed that adding various concentrations of zinc to the sperm does not significantly enhance sperm quality during cold storage and concentrations exceeding 3 mM had detrimental effects. These findings challenge the common belief of the protective role of zinc in sperm preservation, contributing valuable insights for the equine breeding industry. This article belongs to the Collection Equine Science Part II. [ABSTRACT FROM AUTHOR]

Background: Single sperm cryopreservation (SSC) is a specific technique especially used in individuals with small numbers of sperm who suffered from non-obstructive azoospermia (NOA). Testicular specimens possess poor motility and low population of viable spermatozoa. Therefore, sperm selection methods such as applying pentoxifylline (PTX) may improve motility in these cases. The main aim of this study was to evaluate the protective effects of PTX on testicular spermatozoa before and after performing SSC. Methods: Thirty testicular samples were obtained from men with azoospermia. This study was conducted in two phases. Phase 1 evaluated the effect of PTX for sperm selection before SSC. Twenty testicular samples were divided to two experimental groups: SSC without (I) and with PTX treatment (II). For PTX treatment spermatozoa were incubated with PTX at 37°C for 30 min and only motile spermatozoa were selected for SSC. In phase 2, ten testicular samples were cryopreserved with SSC and warming procedure was carried out in droplet with and without PTX. Motility and viability rates, morphology by motile sperm organelle morphology examination (MSOME), DNA fragmentation by sperm chromatin dispersion test (SCD) and mitochondrial membrane potential (MMP) were evaluated. Results: In phase 1, post warm motility rate was higher in PTX exposed group compared to the unexposed group (25.6 ± 8.13 vs. 0.85 ± 2.1) (p > 0.00). Recovery rate, viability and morphology were not significantly different between groups. DNA integrity and MMP were also similar between both groups. In phase 2 although motility increased in PTX group compared to without PTX group (29.30 ± 12.73 vs. 1.90 ± 2.64) (p > 0.00), the viability rate was not different (70.40 ± 12.12 vs. 65.30 ± 11.87). All above mentioned parameters were similar between the two SSC groups. Conclusions: Supplementation of testicular spermatozoa with PTX before cryopreservation increases motility and did not have adverse effects on viability, morphology, DNA integrity and MMP. PTX could be used as sperm selection method before single sperm cryopreservation, but PTX could not maintain motile the most of viable testicular sperms. [ABSTRACT FROM AUTHOR]

This article, titled "Unraveling the Molecular Mechanisms Underlying N-formyl-L-aspartate-mediated Sperm Chemotaxis," explores the role of N-formyl-l-aspartate (NFA) in guiding sperm towards the egg during in vitro fertilization (IVF). The study investigates the chemotactic signaling, receptor identification, and the effect of NFA on sperm capacitation and acrosome reaction. The results indicate that NFA induces chemotaxis in sperm through interaction with the β-2-adrenergic receptor (β-2-AR), leading to calcium mobilization and linear swimming towards the egg. This research provides insights into the molecular mechanisms of sperm chemotaxis and suggests the potential use of NFA as a chemoattractant to improve IVF success rates. [Extracted from the article]

The present study evaluated the protective effect of ascorbic acid (ASCB) against gasoline fumes (PET) induced testicular oxidative stress, sperm toxicity, and testosterone imbalance in Wistar rats. Twenty-four (24) male albino rats (75 ± 16 g) were randomized into three experimental groups (N = 8). The control group: received normal saline, PET group: exposed to PET 6 h daily by inhalation in an exposure chamber and PET + 200 mg ASCB/kg body weight group: exposed to PET 6 h daily by inhalation and administered ASCB per os. Treatment of ASCB and PET exposure was done thrice and five times weekly for a period of 10 weeks respectively. ASCB co-treatment prevented PET-induced increases in the oxidative stress markers (glutathione, glutathione S -transferase, superoxide dismutase, catalase, hydrogen peroxide generation, nitric oxide, and lipid peroxidation) and serum testosterone concentration (p <.05). Sperm quality was low and those with damaged heads and tails increased alongside histological injuries in the PET-exposed rats, which were also minimized with ASCB administration. ASCB protected against PET-induced oxidative stress, sperm, and testis damage in rats. [ABSTRACT FROM AUTHOR]

Male infertility is a global health issue, with 40–50% attributed to sperm abnormalities. The subjectivity and irreproducibility of existing detection methods pose challenges to sperm assessment, making the design of automated semen analysis algorithms crucial for enhancing the reliability of sperm evaluations. This paper proposes a comprehensive sperm tracking algorithm (Sperm YOLOv8E-TrackEVD) that combines an enhanced YOLOv8 small object detection algorithm (SpermYOLOv8-E) with an improved DeepOCSORT tracking algorithm (SpermTrack-EVD) to detect human sperm in a microscopic field of view and track healthy sperm in a sample in a short period effectively. Firstly, we trained the improved YOLOv8 model on the VISEM-Tracking dataset for accurate sperm detection. To enhance the detection of small sperm objects, we introduced an attention mechanism, added a small object detection layer, and integrated the SPDConv and Detect_DyHead modules. Furthermore, we used a new distance metric method and chose IoU loss calculation. Ultimately, we achieved a 1.3% increase in precision, a 1.4% increase in recall rate, and a 2.0% improvement in mAP@0.5:0.95. We applied SpermYOLOv8-E combined with SpermTrack-EVD for sperm tracking. On the VISEM-Tracking dataset, we achieved 74.303% HOTA and 71.167% MOTA. These results show the effectiveness of the designed Sperm YOLOv8E-TrackEVD approach in sperm tracking scenarios. [ABSTRACT FROM AUTHOR]

The coral Acropora spp., known for its reef-building abilities, is a simultaneous hermaphroditic broadcast spawning species. Acropora spp. release gametes into seawater, activating sperm motility. This activation is mediated by adenylyl cyclase (AC) and protein kinase A (PKA). Notably, membrane-permeable cAMP (8-bromo-cAMP) promotes sperm motility activation of Acropora florida. While the signal transduction for PKA-dependent motility activation is highly conserved among animals, the downstream signaling of PKA remains unclear. In this study, we used mass spectrometry (MS) analyses to identify sperm proteins in the coral Acropora digitifera, as well as the serine/threonine residues of potential PKA substrates, and then, we investigated the conservation of these proteins from corals to vertebrates. We identified 148 sperm proteins of A. digitifera with typical PKA recognition motifs, namely RRXT and RRXS. We subsequently used ORTHOSCOPE to screen for orthologs encoding these 148 proteins from corals to vertebrates. Among the isolated orthologs, we identified positive selection in 48 protein-encoding genes from 18 Acropora spp. Subsequently, we compared the conservation rates of the PKA phosphorylation motif residues between the orthologs under positive and purifying selections. Notably, the serine residues of the orthologs under positive selection were more conserved. Therefore, adaptive evolution might have occurred in the orthologs of PKA substrate candidates from corals to vertebrates, accompanied by phosphorylation residue conservation. Collectively, our findings suggest that while PKA signal transduction, including substrates in sperm, may have been conserved, the substrates may have evolved to adapt to diverse fertilization conditions, such as synchronous broadcast spawning. [ABSTRACT FROM AUTHOR]

Purpose: We aimed to evaluate whether and to what extent an association exists between male aging and worsening of semen parameters and to determine whether a threshold age can be identified above which the decline in semen quality becomes statistically significant. Methods: 2612 men (age: 16–56 years) attending an andrology outpatient clinic for semen analysis and clinical evaluation were studied. Semen analyses were performed according to the ongoing WHO-recommended procedures. Total motile count (TMC) and total progressive motile count (TPMC) were calculated by multiplying total sperm number by total motility and progressive motility, respectively. Results: Significant negative correlations were found between age and total motility (r = − 0.131, p < 0.0001), progressive motility (r = − 0.112, p < 0.0001), TPMC (r = − 0.042, p = 0.037), and normal sperm morphology (r = − 0.053, p = 0.007). All these associations persisted in multivariate regression models adjusted for abstinence time, smoking, history of male accessory gland infections, varicocele and the year in which semen analysis was performed. When comparisons were performed among quartiles of increasing age, the fourth quartile, corresponding to the age group > 40 years, was associated with a significant decrease in total and progressive motility. An earlier decline in the TPMC and percentage of normal forms was also observed. Conclusion: Advancing male age exhibits an independent association with a decrease in the percentage of motile and morphologically normal spermatozoa, with greater evidence from the age of > 40 years. Further studies are warranted to elucidate the mechanisms and clinical reflections of these associations. [ABSTRACT FROM AUTHOR]

In this study, we investigated the role of a newly identified homozygous variant (c.1245 + 6T > C) in the CFAP61 gene in the development of multiple morphologically abnormal flagella (MMAF) in an infertile patient. Using exome sequencing, we identified this variant, which led to exon 12 skipping and the production of a truncated CFAP61 protein. Transmission electron microscopy analysis of the patient's spermatozoa revealed various flagellar abnormalities, including defective nuclear chromatin condensation, axoneme disorganization, and mitochondria embedded in residual cytoplasmic droplets. Despite a fertilization rate of 83.3% through ICSI, there was no successful pregnancy due to poor embryo quality. Our findings suggest a link between the identified CFAP61 variant and MMAF, indicating potential disruption in radial spokes' assembly or function crucial for normal ciliary motility. Furthermore, nearly half of the observed sperm heads displayed chromatin condensation defects, possibly contributing to the low blastulation rate. This case underscores the significance of genetic counseling and testing, particularly for couples dealing with infertility and MMAF. Early identification of such genetic variants can guide appropriate interventions and improve reproductive outcomes. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the efficacy of magnetic-activated cell sorting (MACS) or testicular sperm aspiration (TESA) to improve reproductive outcomes in cases with elevated sperm DNA fragmentation undergoing assisted reproduction. Methods: This randomized controlled trial included couples with failed IVF cycles and sperm DNA fragmentation > 30%. Sperm DNA fragmentation was assessed using the sperm chromatin structure assay (SCSA) method. Participants were randomly assigned to either the MACS or TESA group. Testicular sperm retrieval was performed for the TESA group, while MACS involved sperm selection using magnetic beads. Extended blastocyst culture, freeze all policy of blastocysts by vitrification, and frozen embryo transfer were undertaken as per clinic's standard operating protocols. Blastocyst formation rate, implantation rate, miscarriage rate, multiple pregnancy rate, and live birth rate were analyzed and compared between MACS and TESA groups. Results: There were no significant differences in female age, male age, or sperm DNA fragmentation index (DFI) between the MACS and TESA groups. The blastocyst conversion rate was slightly higher in the TESA group (39%) compared to the MACS group (32%). However, the MACS group had a higher implantation rate (50%) than the TESA group (35%). Miscarriage rates, multiple pregnancy rates, and live birth rates did not show statistically significant differences between the groups. A chi-squared test was conducted to compare categorical variables, and t-tests were done to compare continuous variables. Conclusion: In cases with raised sperm DNA fragmentation, sperm selection by MACS or TESA seems to offer comparable reproductive outcomes. There seems no superiority of one intervention over the other in cases with raised sperm DNA fragmentation undergoing assisted reproduction. Both interventions seem to be beneficial for couples seeking assisted reproduction with raised sperm DNA fragmentation. [ABSTRACT FROM AUTHOR]

Objective: We aim to investigate the effect of extreme emotional and psychological trauma following Hamas terrorist attack on October 7th, 2023, on IVF cycle outcome. Patients and methods: The study population consisted of all couples undergoing 2 consecutive IVF attempts with ovum pick-up in our institute, before and during the week of October 8th to 12th, 2023. Embryological/ laboratory variables of the IVF cycles were assessed and compared between the patients' IVF cycle before and those that underwent OPU during the spoken week. Results: Twenty-three couples were eligible for analysis. There were no differences between the cycles in the length of ovarian stimulation, total dose of gonadotropin used, and the peak estradiol and progesterone levels. Furthermore, while no differences were observed in the number of oocytes and mature oocytes retrieved or fertilization rate, the mean number of top-quality embryos per OPU (1.1 ± 1.7 vs. 2.2 ± 2.9; p < 0.02) and ratio of top-quality embryos per number of fertilized oocytes (0.5 ± 0.3 vs. 0.7 ± 0.2; p < 0.01) were significantly lower during the spoken week. Semen total motile count was significantly reduced during the spoken week. Conclusions: In the present study, we are witness to the effect of acute emotional and psychological trauma on IVF outcome, as reflected by its detrimental effect on sperm and embryo quality. [ABSTRACT FROM AUTHOR]

Background: The methodology of surgically extracted sperm cells in fertility treatments remains debated, mainly due to the lack of data evaluating its predictive value on treatment outcomes. Aim: To gain insight into the effectiveness of testicular fine‐needle aspiration (TEFNA) in a cohort of infertile men with absolute non‐obstructive azoospermia and to examine whether the number of retrieved sperm cells affects the fertilisation rate. Materials and Methods: A total of 89 infertile men, aged 26–47, meticulously diagnosed with non‐obstructive azoospermia, participated in the study. All participants underwent TEFNA. The primary outcome measure was the TEFNA success rate in retrieving mature sperm. The secondary outcome measures included fertilisation rate, clinical pregnancy rates, and live births associated with the retrieved sperm. Results: Sperm cells were successfully retrieved from 40 out of 89 patients (45%) with no significant postoperative complications. Retrieval of up to ten sperm cells occurred in 11 procedures (25%); ten procedures (22.7%) resulted in producing dozens of sperm cells, and 100s to 1000s of sperm cells were obtained from the remainder of 23 procedures (52.3%). Patients whose TEFNA resulted in only a few sperm cells had a much lower fertilisation rate (16.6%) than the other two groups (40.1% and 47.2%, respectively, P = 0.003). Conclusions: The utilisation of TEFNA for sperm extraction in men with non‐obstructive azoospermia is a simple, fast‐learning, effective, and safe treatment option. In cases where sperm retrieval was successful, the fertilisation rate was strongly related to the number of sperm cells obtained. [ABSTRACT FROM AUTHOR]

Glutathione peroxisomal‐5 (Gpx5) promotes the elimination of H2O2 or organic hydrogen peroxide, and plays an important role in the physiological process of resistance to oxidative stress (OS). To directly and better understand the protection of Gpx5 against OS in epididymal cells and sperm, we studied its mechanism of antioxidant protection from multiple aspects. To more directly investigate the role of Gpx5 in combating oxidative damage, we started with epididymal tissue morphology and Gpx5 expression profiles in combination with the mouse epididymal epithelial cell line PC1 (proximal caput 1) expressing recombinant Gpx5. The Gpx5 is highly expressed in adult male epididymal caput, and its protein signal can be detected in the sperm of the whole epididymis. Gpx5 has been shown to alleviate OS damage induced by 3‐Nitropropionic Acid (3‐NPA), including enhancing antioxidant activity, reducing mitochondrial damage, and suppressing cell apoptosis. Gpx5 reduces OS damage in PC1 and maintains the well‐functioning extracellular vesicles (EVs) secreted by PC1, and the additional epididymal EVs play a role in the response of sperm to OS damage, including reducing plasma membrane oxidation and death, and increasing sperm motility and sperm‐egg binding ability. Our study suggests that GPX5 plays an important role as an antioxidant in the antioxidant processes of epididymal cells and sperm, including plasma membrane oxidation, mitochondrial oxidation, apoptosis, sperm motility, and sperm‐egg binding ability. [ABSTRACT FROM AUTHOR]

Background and Aim: The management of incubation conditions impacts embryonic development, hatchability, and posthatch performance. This study aimed to examine the effects of thermal manipulation (TM) during embryonic development on roosters' thermotolerance, antioxidant activity, immunity, and semen quality under heat-stress conditions. Materials and Methods: 1200 fertile eggs were distributed evenly between two groups, each containing three replicates (200 eggs/replicate). The first group (G1) was held in the commercial setter with a consistent temperature of 37.5°C and 55% relative humidity (RH) through the 18-day incubation period, acting as a control, while the second group (G2) experienced these conditions until only the 11th day. The eggs were incubated at 39.5°C with 60% RH for 4 h each day from the 12th to the 18th day. From the 19th to 22nd incubation days, both groups maintained a consistent temperature of 37.2°C with a RH of 70%. Two hundred hatched male chicks per treatment group were moved into a closed-system house. All roosters were exposed to a 6-h daily heat challenge with a temperature of 35°C and a humidity of 70% between their 36th and 40th weeks of age. Results: Roosters of G2 exposed to thermal challenge showed improvements (p ≤ 0.05) in multiple blood biochemical, antioxidant, and immunity markers, including total protein, globulin, aspartate aminotransferase, alanine aminotransferase, triiodothyronine, thyroxine, corticosterone, testosterone, total antioxidant capacity, malondialdehyde, immunoglobulin G, immunoglobulin M, and immunoglobulin A levels. Improved semen quality characteristics, including ejaculate volume, sperm concentration, motility, livability, and quality factor, as well as enhanced thermoregulation in post-hatch cocks, were also achieved (p ≤ 0.05). Conclusion: To boost antioxidant activity, immunity, thermotolerance, and semen parameters in roosters under heat-stress conditions, TM application during egg incubation, specifically at 12-18 days, is recommended. [ABSTRACT FROM AUTHOR]

Objective: Recently, oral antioxidants in combined forms have been used to treat men with idiopathic infertility. This study aimed to evaluate the effects of treatment with vitamin C, vitamin E, selenium, zinc, arginine, L-carnitine, and coenzyme Q10 on sperm quality parameters, DNA integrity, reproductive hormones, and pregnancy rates in men with infertility and idiopathic oligoasthenoteratozoospermia (OAT). Methods: A prospective study was conducted on 420 men with infertility and idiopathic OAT who took an oral supplement of antioxidant SP-Power tablets twice daily for 6 months. Semen quality, reproductive hormones, and the DNA fragmentation index (DFI) were evaluated at baseline and at 3 and 6 months after supplementation, using the World Health Organization 2021 guidelines. Results: No significant difference was observed in volume or the percentage of typical morphology during treatment. A significant improvement in sperm concentration was observed after supplementation (8.67±1.41, 12.17±1.91, and 19.01±0.86 at baseline, 3, and 6 months respectively, p<0.01). The total motility, progressive motility, and total motile sperm count also increased significantly (p<0.01), whereas the DFI decreased after 6 months. There was an increase in normal FSH levels and testosterone levels after 6 months of supplementation of antioxidant SP-Power, but these differences were not statistically significant (p=not significant and p=0.06, respectively). Conclusion: Supplementation with SP-Power tablets improved sperm quality parameters, sperm DFI, some reproductive hormones, and pregnancy rates in men with infertility and idiopathic OAT, which could be attributed to the supplement's synergistic antioxidant action. Further studies are needed to determine the effects of supplementation on oxidative stress markers. [ABSTRACT FROM AUTHOR]

Vincristine (VCR) is a microtubule‐destabilizing chemotherapeutic agent commonly administered for the treatment of cancers in patients, which can induce severe side effects including neurotoxicity. In context of the effects on female fertility, ovarian toxicity has been found in patients and mice model after VCR exposure. However, the influence of VCR exposure on oocyte quality has not been elucidated. We established VCR exposure in vitro and in vivo model. The results indicated in vitro VCR exposure contributed to failure of oocyte maturation through inducing defects in spindle assembly, activation of SAC, oxidative stress, mitochondrial dysfunction, and early apoptosis, which were confirmed by using in vivo exposure model. Moreover, in vivo VCR exposure caused aneuploidy, reduced oocyte‐sperm binding ability, and the number of cortical granules in mouse oocyte cortex. Taken together, this study demonstrated that VCR could cause meiotic arrest and poor quality of mouse oocyte. [ABSTRACT FROM AUTHOR]

B chromosomes are intriguing "selfish" genetic elements, many of which exhibit higher-than-Mendelian transmission. This perspective highlights a group of B chromosomes known as Paternal Sex Ratio chromosomes (PSRs), which are found in several insects with haplo-diploid reproduction. PSRs harshly alter the organism's reproduction to facilitate their own inheritance. A manifestation of this effect is the conversion of female destined individuals into males. Key to this conversion is the mysterious ability of PSRs to cause elimination of the sperm-inherited half of the genome during zygote formation. Here we discuss how PSRs were discovered, what is known about how they alter paternal chromatin dynamics to cause sex conversion, and how PSR-induced genome elimination is different from other forms of programmed genome elimination in different insects. PSRs also stand out because their DNA sequence compositions differ in remarkable ways from their insect's essential chromosomes, a characteristic suggestive of interspecies origins. Broadly, we also highlight poorly understood aspects of PSR dynamics that need to be investigated. [ABSTRACT FROM AUTHOR]

Chinese sleeper is an invasive species from South Asia inhabiting Eastern and Central Europe. So far, no detailed analysis of the reproductive cycle of Chinese sleepers has been conducted. The aim of the study was to describe the reproductive cycle of Chinese sleepers in the Wilanówka River, Vistula tributary (Poland, Central Europe). Fish morphometric data, age, gonadal maturity, and absolute and relative fecundity of females were analysed. A standard paraffin technique and six-stage scales were used to assess the gonad development and maturation in both sexes. The age of the studied fish ranged from 0+ to 6+.The female-to-male ratio was 1:1. Females and males mature at age 1+.The smallest female and male reaching maturity were 3.8 and 3.7 cm long, respectively. An extended spawning season from the beginning of April to mid-September was determined based on gonadal maturity analysis. The mean absolute fecundity was 2,971 (180–19,656) oocytes. The male reproductive system comprises paired testis (spermatogenic region, blackish color) and paired testicular gland (secretory region, whitish color). In both regions, seasonal changes were observed. Male gonads during winter had completed spermatogenesis, and the lobules were filled with spermatozoa. Very fast spermatogenesis and overlapping of reproductive cycles have been observed in Chinese sleeper males. Almost all year round testis contains spermatozoa. In the new habitat examined, this species was characterized by earlier maturation and longer reproduction season, compared to the native habitat and other native species of the examined area. Moreover, high female fecundity, multiple spawning, and lower investment in the development of gonads in males allow maintaining vitality and protecting the nest, contribute to the species competitiveness and successful colonization of new areas. [ABSTRACT FROM AUTHOR]

Epigenetic modifications can alter the function of genes. The epigenetics changes are caused by environmental effects, which lead to chemical modifications of the DNA or the chromatin. The mechanisms involve the influence of small interfering siRNAs on gene silencing. Epigenetic changes normally last only during the life‐time of an individual and are erased in embryos and eggs for a naive progeny. The genomes are reprogrammed and the chemical modifications removed to restart the next generation. However, there are mechanisms that allow the genome to escape from such a clearing effect so that modifications can be transmitted to one or more subsequent generations. In the germline of animal cells small RNAs, including piRNAs, have evolved which guarantee a higher degree of fidelity for transmission of genetic information, guarding especially against the detrimental effect caused by transposon activity. piRNA is essential for transposon silencing for survival of a species and protection of subsequent generations. Inactivation of piRNA results in abundant transposon activity and sperm infertility. The effect in humans has been described but is less distinct. Some stress‐induced epigenetic changes are transitory in mice and can be reversed by a change of environment or lifestyle. [ABSTRACT FROM AUTHOR]

Simple Summary: The choice of the optimal sperm preservation method is an important consideration in animal breeding. Stored semen can be used for reproductive purposes to introduce new genotypes and prevent inbreeding, which poses a considerable problem in cervid farms. The aim of this study was to evaluate and compare the effect of storage time and storage method (liquid state/epididymides) on the motility, morphology, and antioxidant status of European red deer sperm stored at 5 °C for up to six days (D0-D6). Sperm samples were assessed for motility, viability, morphology, activity of antioxidant enzymes (superoxide dismutase, SOD; glutathione peroxidase, GPx; catalase, CAT), and lipid peroxidation (based on malondialdehyde, MDA, content). Significant differences between storage variants were noted on D2 in sperm morphology; on D4 in the percentage of progressively motile sperm, MDA content, and SOD and GPx activity; and on D6 in the percentage of motile and viable spermatozoa. Sperm motility, viability, and antioxidant status are more effectively preserved during liquid storage than epididymal storage. Morphological and functional abnormalities of sperm were observed earlier during epididymal storage, which suggests that spermatozoa can be stored for shorter periods of time in the epididymides than in a liquid state. The aim of this study was to evaluate the motility, morphology, and antioxidant status of European red deer sperm stored in a liquid state (variant I) and in the epididymides (variant II). Spermatozoa were harvested post-mortem from the cauda epididymis. Sperm samples in both variants were stored for up to six days (D6) at 5 °C. Spermatozoa were assessed for motility, viability, morphology, activity of antioxidant enzymes (superoxide dismutase, SOD; glutathione peroxidase, GPx; catalase, CAT), and lipid peroxidation (malondialdehyde, MDA, content). Sperm samples were analyzed on storage days 0, 2, 4, and 6 (D0-D6). Storage time and storage method significantly (p ≤ 0.05) influenced the examined variables. On D2, a decrease in motility and acrosomal integrity was observed in both storage variants, whereas a decrease in viability and an increase in MDA content were noted in spermatozoa stored in the epididymides. On D4, higher values of SOD and GPx activity and MDA content were noted in variant I than in variant II. Catalase activity was very low. GPx is the key enzyme that participates in the reduction of hydrogen peroxide in sperm cells. Spermatozoa stored in a liquid state were characterized by higher motility and viability, improved morphology and antioxidant status than those stored in the epididymides; therefore, liquid storage is more recommended for short-term preservation of epididymal spermatozoa. [ABSTRACT FROM AUTHOR]

Simple Summary: Simple Summary: There are very few reports regarding Assisted Reproductive Technologies in South American camelids, including llamas (Lama glama). The sperm quality and in vitro fertilization rates from fresh and cryopreserved sperm collected from vas deferens using different preservation extenders were investigated in llama males as a model for its application in other South American camelids. The obtained results from our research showed that the preservation extender was a determining factor in significantly improving the in vitro fertilization rates when using fresh and cryopreserved sperm samples obtained from the vas deferens in llamas (Lama glama). The advances in Assisted Reproductive Technologies (ARTs) applied in South American camelid species are still scarce. The aim of this study was to compare the effects of three semen extenders, before and after the cryopreservation of spermatozoa obtained from the vas deferens, on sperm quality parameters and in vitro fertilization rates of llama (Lama glama) oocytes. Mature fertile llama males (Lama glama; n = 6; age: 48–60 mo.; BCS: ~2.7) were included in the study. Sperm samples were collected from each male using the surgical technique of the vas deferens deviation. Then, the sperm samples were pooled and diluted with the Tris-EY, Andromed®, or BioxCell® extender in order to subsequently carry out the sperm cryopreservation process. The sperm quality assessment related to each extender was performed before and after cryopreservation with regard to sperm morphological abnormalities, acrosome integrity, sperm viability, membrane permeability, and sperm motility traits. Moreover, in vitro fertilization (IVF) procedures were carried out to evaluate the in vitro fertility of the cryopreserved sperm samples using each extender. Overall, significant differences were observed before and after cryopreservation regarding acrosome integrity, sperm viability, membrane permeability, and sperm motility traits among the extenders used, where Tris-EY and Andromed® were better than BioxCell® (p < 0.05); however, no differences were observed regarding the sperm morphological abnormalities among extenders (p > 0.05). Moreover, multiple differences were observed with regard to the velocity and linearity kinematic parameters obtained by computerized analysis before and after the cryopreservation process, irrespective of the extender used (p < 0.05). Finally, differences were observed regarding the in vitro fertilization rates among the different extender-derived samples (p < 0.05). In conclusion, the sperm quality using Tris-EY and Andromed® was better before and after cryopreservation compared to that using BioxCell®. Although the number of fertilized oocytes obtained after the IVF process between Tris-EY and Andromed® was similar, Andromed®-derived samples showed the best sperm quality results before and after cryopreservation. This indicates that the cryopreservation extender is a determining factor in significantly improving in vitro fertilization rates when using sperm samples obtained from vas deferens in llama (Lama glama) males. [ABSTRACT FROM AUTHOR]

To investigate whether PIEZO–intracytoplasmic sperm injection (PIEZO-ICSI) increases the fertilization rate, decreases the degeneration rate, and increases the utilization rate per oocyte injected compared with conventional intracytoplasmic sperm injection (ICSI). Sibling oocyte split multicenter trial. Fertility clinics. Women with a diagnosis of infertility who used ICSI as their method of insemination and had ≥6 mature oocytes for injection. Participants had their mature oocyte cohort divided, where half were injected using conventional ICSI and the other half were injected using PIEZO-ICSI. For patients with an uneven oocyte number, the extra oocyte was injected using conventional ICSI. The injection technique used first was also randomized to ensure that there was no bias due to order of injection. The primary outcome measure was the fertilization rate after injection. A total of 108 patients underwent a sibling split use of conventional ICSI and PIEZO-ICSI. The fertilization rate was 71.6% in PIEZO-ICSI, which significantly increased compared with that in conventional ICSI 65.6%. In addition, the oocyte degeneration rate decreased in PIEZO-ICSI compared with that in conventional ICSI (6.3% vs. 12.1% respectively), and the blastocyst quality increased, as measured by the number of grade A and B quality blastocysts present on day 5 of development (33.3% vs. 27.5%). No significant differences in the aneuploidy or utilization rate, clinical pregnancy, or live birth outcome after single embryo transfer were noted between the two injection techniques. This trial supports the possibility that PIEZO-ICSI increases the fertilization rates, decreases the oocyte degeneration rates, and increases the blastocyst quality compared with conventional ICSI; however, it does not appear to influence the clinical pregnancy or live birth rate per transfer. Australian and New Zealand Clinical Trial Registry ACTRN12620000407998. [ABSTRACT FROM AUTHOR]

Elasmobranchs have an ancestral reproductive system, which offers insights into vertebrate reproductive evolution. Despite their unchanged design over 400 million years, they evolved complex mechanisms ensuring reproductive success. However, human activities induced a significant decline in elasmobranch populations worldwide. In the Mediterranean basin, the smooth-hound shark (Mustelus mustelus) is one of the species that are considered vulnerable to human activities. Conservation efforts necessitate a thorough understanding of its reproductive strategy. This study focused on mature male specimens of smooth-hound sharks that were captured in the Adriatic area and successively analyzed to provide, for the first time, a histologically detailed description of testicular development in the species. Seven phases of the spermatogenesis process were identified, along with the macromolecular characterization of cells obtained using Fourier-transform infrared imaging. Histological analysis showed structural and cellular features similar to those documented in the spermatocysts of other elasmobranchs. The examination of the evolution and migration of both germinative and Sertoli cells at each phase revealed their close connection. Furthermore, different expression levels of lipids, proteins, and phosphates (DNA) at each spermatogenesis stage were observed. This research provided new information on spermatogenesis in the common smooth-hound shark, which is crucial for conservation efforts against population decline and anthropogenic pressures. [ABSTRACT FROM AUTHOR]

Reproductive failure in dogs is often due to unknown causes, and correct diagnosis and treatment are not always achieved. This condition is associated with various congenital and acquired etiologies that develop inflammatory processes, causing an increase in the number of leukocytes within the female reproductive tract (FRT). An encounter between polymorphonuclear neutrophils (PMNs) and infectious agents or inflammation in the FRT could trigger neutrophil extracellular traps (NETs), which are associated with significantly decreased motility and damage to sperm functional parameters in other species, including humans. This study describes the interaction between canine PMNs and spermatozoa and characterizes the release of NETs, in addition to evaluating the consequences of these structures on canine sperm function. To identify and visualize NETs, May–Grünwald Giemsa staining and immunofluorescence for neutrophil elastase (NE) were performed on canine semen samples and sperm/PMN co-cultures. Sperm viability was assessed using SYBR/PI and acrosome integrity was assessed using PNA-FITC/PI by flow cytometry. The results demonstrate NETs release in native semen samples and PMN/sperm co-cultures. In addition, NETs negatively affect canine sperm function parameters. This is the first report on the ability of NETs to efficiently entrap canine spermatozoa, and to provide additional data on the adverse effects of NETs on male gametes. Therefore, NETs formation should be considered in future studies of canine reproductive failure, as these extracellular fibers and NET-derived pro-inflammatory capacities will impede proper oocyte fertilization and embryo implantation. These data will serve as a basis to explain certain reproductive failures of dogs and provide new information about triggers and molecules involved in adverse effects of NETosis for domestic pet animals. [ABSTRACT FROM AUTHOR]