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Sperm motility is a crucial factor in male fertility. Photobiomodulation (PBM) has been reported to increase sperm motility, but a consistent approach suitable for identifying standardizable protocols is lacking. We collected asthenozoospermic (n = 70) and normozoospermic (n = 20) semen. The asthenozoospermic samples were irradiated with an 810 nm diode laser, in continuous wave mode, at 0.25 W, 0.5 W, 1 W and 2 W for 60 s on a circular area of 1 cm2 through a novel handpiece with an innovative flat-top profile. Sperm motility was assessed immediately, after 30 and 60 min. A sample size calculator, unpaired t-test and one-way ANOVA with post-hoc Tukey HSD tests were used for statistics. One and 2 W were the most effective outputs in increasing progressive motility compared to control (p < 0.001). The maximum effect was immediately after 1 W-PBM (p < 0.001) and decreased after 60 min (p < 0.001). Time physiologically decreased vitality (p < 0.001), but less in the 1 W-PBM samples (p < 0.05). 1 W-PBM did not affect chromatin condensation. Asthenozoospermic samples displayed an impairment of 80% in oxygen consumption and ATP production and a slight inefficiency of oxidative phosphorylation compared to normozoospermic samples (p < 0.001). 1 W-PBM partially restored the functionality of aerobic metabolism (p < 0.001) by recovery of oxidative phosphorylation efficiency. PBM did not affect lactate dehydrogenase (glycolysis pathway). No irradiated samples increased accumulated malondialdehyde, a marker of lipidic peroxidation. In conclusion, PBM improves progressive motility in asthenozoospermia through increased mitochondrial energetic metabolism without harmful oxidative stress. [ABSTRACT FROM AUTHOR]

During the artificial breeding process of Chinese pearl oysters, the quality and vitality of sperm and eggs are pivotal in determining the success rate of fertilization. This study delves into the impact of externally added fructose, glucose, and ammoniated water on sperm vitality and lifespan under varying temperature conditions. The results indicate that the addition of exogenous energy sources and cell protectants, specifically monosaccharide energy substances, significantly improves sperm vitality. Moreover, under 4°C conditions, adding 2.8 mmol/L fructose, glucose, and 2 mmol/L ammoniated water to sperm samples significantly enhances sperm vitality and prolongs their lifespan. This provides strong evidence for the feasibility of short-term preservation and vitality enhancement strategies for Chinese pearl oyster sperm and also offers important scientific support for optimizing the artificial reproduction techniques of Chinese pearl oysters. [ABSTRACT FROM AUTHOR]

Background Cyclophosphamide (CP) is an anticancer drug; however, clinical utilization of CP is limited, resulting from its considerable toxicities. This research was performed to explore the protective effects of Chlorogenic acid (CGA) on reproductive damage induced by CP in mice. Methods Blood samples were collected for analysis of hormone content subsequently; semen samples were evaluated for quality, and testis samples were used for histopathological evaluation and analysis of oxidative stress biomarkers, protein and gene expression levels of steroid regulatory factors, and steroid synthase. Results The results noted that CGA increased serum testosterone (T), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) activity; increased SOD, GPx, and GSH oxidative stress levels in testis tissue; and decreased MDA content in testis tissue. Testicular cells in the CGA treatment group gradually returned to normal morphology, and CYP11A1 and CYP17A1 levels increased after CGA treatment. The mRNA levels of CYP11A1, CYP17A1, StAR, 3β-HSD, and 17β-HSD were significantly raised in the CGA dose group. In the test dose range, CGA can improve sperm quality, quantitative abnormality, and serum T synthesis disorder caused by CP. This mechanism may be correlated with the inhibition of oxidative stress and antioxidation levels. Conclusions Therefore, CGA has a protective impact on testicular injuries arising from CP in mice. [ABSTRACT FROM AUTHOR]

Abstract Sperm motility is a crucial factor in male fertility. Photobiomodulation (PBM) has been reported to increase sperm motility, but a consistent approach suitable for identifying standardizable protocols is lacking. We collected asthenozoospermic (n = 70) and normozoospermic (n = 20) semen. The asthenozoospermic samples were irradiated with an 810 nm diode laser, in continuous wave mode, at 0.25 W, 0.5 W, 1 W and 2 W for 60 s on a circular area of 1 cm2 through a novel handpiece with an innovative flat-top profile. Sperm motility was assessed immediately, after 30 and 60 min. A sample size calculator, unpaired t-test and one-way ANOVA with post-hoc Tukey HSD tests were used for statistics. One and 2 W were the most effective outputs in increasing progressive motility compared to control (p

Necrozoospermia is a poorly documented condition with a low incidence, and its definition and clinical significance are unclear. Herein, we provide a reference range for necrozoospermia and discuss its possible etiology and impact on male fertility and assisted reproductive outcomes. We extracted relevant information from 650 Chinese male partners of infertile couples and statistically analyzed sperm vitality. Necrozoospermia was present in 3.4% (22/650) of our study population, and the lower cut-off value for sperm vitality was 75.3%. We compared two methods for assessing sperm vitality (eosin-nigrosin head staining and hypo-osmotic swelling test [HOST]), for which the percentage in the eosin-nigrosin group (mean ± standard deviation [s.d.]: 77.5% ± 10.5%) was significantly higher than that in the HOST group (mean ± s.d.: 58.1% ± 6.7% [5–10 min after incubation] and 55.6% ± 8.2% [25–30 min after incubation]; both P < 0.001). The incidence of necrozoospermia increased with age (odds ratio [OR] = 1.116, 95% confidence interval [CI]: 1.048–1.189, P = 0.001), while the percentage of normal sperm morphology and DNA fragmentation index (DFI) were significantly associated with necrozoospermia, with ORs of 0.691 (95% CI: 0.511–0.935, P = 0.017) and 1.281 (95% CI: 1.180–1.390, P < 0.001), respectively. In the following 6 months, we recruited 166 patients in the nonnecrozoospermia group and 87 patients in the necrozoospermia group to compare intracytoplasmic sperm injection (ICSI) and pregnancy outcomes between the two groups. The necrozoospermia group had a significantly lower normal fertilization rate (74.7% vs 78.2%, P = 0.041; OR = 0.822; 95% CI: 0.682–0.992) than that in the nonnecrozoospermia group. This study presents substantial information on necrozoospermia to establish comprehensive and applicable reference values for sperm vitality for spontaneous conception and artificially assisted reproductive management.

Simple Summary: This study explores how different concentrations of ammonium ions affect the sperm health of three important bivalve species: Noble Scallop Mimachlamys nobilis, Chinese Pearl Oyster Pinctada fucata martensii, and Small Rock Oyster Saccostrea mordax. Bivalves are crucial to aquatic ecosystems, and understanding the factors that impact their reproduction is essential. Our research found that higher levels of ammonium reduced sperm vitality and movement in Noble Scallop and Small Rock Oysters, especially at the highest concentration we tested. Interestingly, the Chinese Pearl Oyster showed improved sperm vitality and movement at moderate ammonium levels. Additionally, we observed the increased curling of sperm tails in Noble Scallops and Small Rock Oysters with higher ammonium levels, but this effect was not seen in Chinese Pearl Oysters. These findings highlight how varying environmental conditions can differently affect species and emphasize the importance of maintaining balanced ammonium levels in aquatic habitats. This knowledge is valuable for protecting the reproductive health of these bivalves, which play a key role in the health and sustainability of aquatic ecosystems. Ammonium (NH4+) plays a crucial role in the reproductive processes of key biotic groups in aquatic ecosystems—bivalves. This study aims to elucidate the effects of three different ammonium ion concentrations on sperm vitality, swimming kinematics, and morphology of Mimachlamys nobilis, Pinctada fucata martensii, and Saccostrea mordax. The results indicate that the sperm vitality and motility rates of M.nobilis and S. mordax are inversely proportional to the ammonium concentration, especially in the treatment group with an ammonium concentration of 3 mmol/L, where the decrease in sperm vitality and motility is most significant. In contrast, the sperm of P. fucata martensii reacted differently to increasing ammonium concentrations. After the addition of 2 mmol/L of ammonium, the sperm vitality and motility of P. fucata martensii reached a peak, showing a significant stimulatory effect. Additionally, as the ammonium concentration increased, the curling of the sperm flagella in M.nobilis and S. mordax increased. However, sperm flagella curling in P. fucata martensii showed no change compared to the control group. This study provides insights into the effects of ammonium concentrations on the sperm vitality and motility of three marine bivalve species and highlights the importance of sperm flagella curling as a factor affecting sperm. [ABSTRACT FROM AUTHOR]

Oxidative processes give rise to reactive oxygen species/oxidants which are detrimental to living cells and underlie many pathologic conditions. On the other hand, many plant species possess antioxidant properties and phytochemicals which are closely linked to their therapeutic potentials. Solanum macrocarpon is a tropical plant which is traditionally claimed to possess therapeutic properties. Although, antioxidant potentials have been reported for different Solanum species and antioxidants have been reported to improve sperm morphology and motility; this has not been fully elucidated in S. macrocarpon. Following a comparative phytochemical and in vitro antioxidant screening of S. macrocarpon fruit, leaf and root ethanol extracts, this study evaluated the effect of S. macrocarpon leaf ethanol extract (SMLE) on sperm parameters in vitro and antioxidant enzymes in vivo. The effect of SMLE on sperm motility and vitality was examined in vitro at concentrations of 2.5, 5 and 10 mg/ml at 1, 5 and 10 min' intervals. Furthermore, thirty male albino rats (10–12 weeks old), separated at random into six groups (A–F, n = 5), and administered SMLE for 42 days, were used to investigate the antioxidant potency (AP) of SMLE in vivo. The phytochemical screening of different parts of S. macrocarpon revealed that SMLE had the highest (p < 0.05) composition (mg/100 g) of tannins, flavonoids, steroids, terpenoids and saponin compared to fruit and root extracts. Furthermore, SMLE had the highest (p < 0.05) DPPH-scavenging antioxidant potency, which was similar to that of the control (ascorbic acid) at 200 µg/ml concentration, compared to the fruit and root extracts. SMLE significantly (p < 0.05) exhibited spermicidal potency in vitro in terms of reduced sperm motility and vitality but showed no significant effect on lipid peroxidation/antioxidant enzymes in vivo. The findings suggest that the SMLE possesses in vitro spermicidal potential despite its potent antioxidant activity. [ABSTRACT FROM AUTHOR]

Several studies have demonstrated interesting results considering the implication of three growth factors (GFs), namely nerve growth factor (NGF), erythropoietin (EPO), and the insulin-like growth factor-I (IGF-1) in the physiology of male reproductive functions. This review provides insights into the effects of NGF, EPO, and IGF-1 on the male reproductive system, emphasizing mainly their effects on sperm motility and vitality. In the male reproductive system, the expression pattern of the NGF system varies according to the species and testicular development, playing a crucial role in morphogenesis and spermatogenesis. In humans, it seems that NGF positively affects sperm motility parameters and NGF supplementation in cryopreservation media improves post-thaw sperm motility. In animals, EPO is found in various male reproductive tissues, and in humans, the protein is present in seminal plasma and testicular germ cells. EPO receptors have been discovered in the plasma membrane of human spermatozoa, suggesting potential roles in sperm motility and vitality. In humans, IGF-1 is expressed mainly in Sertoli cells and is present in seminal plasma, contributing to cell development and the maturation of spermatozoa. IGF-1 seems to modulate sperm motility, and treatment with IGF-1 has a positive effect on sperm motility and vitality. Furthermore, lower levels of NGF or IGF-1 in seminal plasma are associated with infertility. Understanding the mechanisms of actions of these GFs in the male reproductive system may improve the outcome of sperm processing techniques. [ABSTRACT FROM AUTHOR]

Ammonium (NH4+) plays a crucial role in the reproductive processes of key biotic groups in aquatic ecosystems—bivalves. This study aims to elucidate the effects of three different ammonium ion concentrations on sperm vitality, swimming kinematics, and morphology of Mimachlamys nobilis, Pinctada fucata martensii, and Saccostrea mordax. The results indicate that the sperm vitality and motility rates of M.nobilis and S. mordax are inversely proportional to the ammonium concentration, especially in the treatment group with an ammonium concentration of 3 mmol/L, where the decrease in sperm vitality and motility is most significant. In contrast, the sperm of P. fucata martensii reacted differently to increasing ammonium concentrations. After the addition of 2 mmol/L of ammonium, the sperm vitality and motility of P. fucata martensii reached a peak, showing a significant stimulatory effect. Additionally, as the ammonium concentration increased, the curling of the sperm flagella in M.nobilis and S. mordax increased. However, sperm flagella curling in P. fucata martensii showed no change compared to the control group. This study provides insights into the effects of ammonium concentrations on the sperm vitality and motility of three marine bivalve species and highlights the importance of sperm flagella curling as a factor affecting sperm.

The main aim of this randomized controlled pilot trial was to evaluate the effects of hydrogen-rich water on spermiogram parameters in normospermic and oligospermic men. As many as 12 healthy young men (age 29.1 ± 5.9 years; n = 6 normospermic; n = 6 oligospermic) volunteered. Participants were allocated in a double-blind manner to receive 1 L of hydrogen-rich water per day or 1 L of tap water fortified with magnesium for 8 weeks. Following hydrogen-rich water supplementation, sperm concentration and morphology tended to increase by 12.4 million per milliliter (95% CI; -31.8 to 56.6), and live cells increased by 3.8% (95% CI; -12.5 to 20.1), respectively (P = 0.30). A significant difference between the two interventions was found in sperm vitality (P = 0.03), with hydrogen-rich water being superior to the placebo in terms of an increase in the number of live sperm cells. Subgroup analyses revealed no significant differences between interventions in sperm viability outcomes in oligospermic men (P > 0.05). However, sperm count increased by over 20 million in two of the three oligospermic men (66.7%) receiving hydrogen-rich water, while no oligospermic men in the placebo group increased sperm concentration to euspermic levels (P < 0.01). These preliminary findings suggest hydrogen-rich water is an effective intervention for improving some aspects of male subfertility; further large-sample trials are warranted to corroborate our results. [ABSTRACT FROM AUTHOR]

Epicatechin (EPC) is a flavonoid belonging to the family of catechins; it has been described as a powerful scavenger of a wide spectrum of reactive oxygen species (ROS) and a modulator of ex vivo sperm vitality. In this study, we assessed the potential protective abilities of EPC on cryopreserved bovine spermatozoa. We focused on conventional quality parameters, as well as the oxidative profile of spermatozoa alongside capacitation patterns, and expression profiles of proteins involved in the process of capacitation. Semen samples were cryopreserved in the presence of 25, 50 or 100 μmol/L EPC and compared to native semen (negative control) as well as ejaculates frozen in the absence of EPC (positive control). A dose-dependent improvement of conventional sperm quality parameters was observed following EPC administration, particularly in case of the sperm motility, membrane, acrosome and DNA integrity in comparison to the positive control. Experimental groups exposed to all EPC doses presented with a significantly lower proportion of capacitated spermatozoa as opposed to the positive control. While no significant effects of EPC were observed in cases of superoxide production, a significant decrease in the levels of hydrogen peroxide and hydroxyl radical were recorded particularly in the experimental groups supplemented with 50 and 100 μmol/L EPC. Western blot analysis revealed that supplementation of particularly 100 μmol/L EPC to the semen extender prevented the loss of the cation channel of sperm (CatSper) isoforms 1 and 2, sodium bicarbonate cotransporter (NBC) and protein kinase A (PKA), which play important roles in the process of sperm capacitation. In summary, we may hypothesize that EPC is particularly effective in the stabilization of the sperm membrane during the freeze–thaw process through its ability to quench ROS involved in damage to the membrane lipids and to prevent the loss of membrane channels crucial to initiate the process of sperm capacitation. These attributes of EPC provide an additional layer of protection to spermatozoa exposed to low temperatures, which may be translated into a higher post-thaw structural integrity and functional activity of male gametes. [ABSTRACT FROM AUTHOR]

Two isoforms of the gonadotropin‐releasing hormone (GnRH), GnRH‐I and GnRH‐II, are expressed in mammals, and the presence of one or more GnRH‐like peptides has been demonstrated in the male reproductive tract. GnRH and its receptors (GnRHR) are present in human and non‐human primate testis, prostate, epididymis, seminal vesicle, spermatozoa and seminal human plasma. GnRH‐II is site‐specific and acts directly in an inhibitory or stimulatory fashion. Previous studies speculated that GnRH‐II could disrupt specific sperm processes, such as sperm motility or capacitation and could be utilized as an effective contraceptive agent. Our study aimed to investigate the in‐vitro effects of GnRH‐I and GnRH‐II on Vervet monkey sperm function. Electro‐ejaculated semen samples from 10 Vervet monkeys (Chlorocebus aethiops) were used to select motile sperm populations. Sperm aliquots were incubated with GnRH‐I and GnRH‐II at different concentrations for 1 h, where after sperm motility and kinematic parameters were assessed using the automated Sperm Class Analyser. Additional sperm aliquots were incubated with two 10‐amino acid control peptides, a non‐related peptide and an inactive peptide to exclude the possible influence on sperm motility from other peptides with a structure similar to GnRH. Additionally, a GnRHR‐I antagonist (GnRHR‐A), Cetrorelix, was tested to establish its antagonistic capability on GnRH. The effect of selected concentrations of GnRH‐I and GnRH‐II on sperm vitality and acrosome intactness was also evaluated after 10‐ and 60 min exposure. Analysis of the percentage total sperm motility revealed that different concentrations for GnRH‐I and GnRH‐II inhibited sperm motility significantly. While sperm progressiveness was also notably affected and a trend of decreased sperm kinematics were evident, no effect was found on sperm vitality or acrosome intactness. The non‐related and inactive peptides had no impact on sperm motility. The GnRHR‐A demonstrated no effect on sperm motility and effectively blocked the inhibitory outcome on the motility of both GnRH isoforms. While GnRH‐I or GnRH‐II at low‐dose concentrations resulted in in‐vitro inhibition of sperm motility, it appears to have no adverse effects on other sperm functional parameters evaluated. These collective observations possibly indicate an essential role for GnRH in the in‐vivo process of sperm selection in the female reproductive tract. [ABSTRACT FROM AUTHOR]

Purpose: Female genital candidiasis is a common gynecological complaint. Studies demonstrated its negative impact on sperm motility. In the current study we aimed at determining the effect of vaginally extracted Candida albicans on different semen parameters. Study Design: Vaginal swabs were obtained from 25 females suffering from vaginitis and semen samples from 18 normozoospermia healthy males. Semen samples were inoculated with different concentrations of vaginal isolates of Candida albicans (0 (control), 20000, 60000 and 100000 cfus/0.5ml) for two incubation periods (5 and 15 hours). Semen was tested for sperm motility, vitality, morphology, and fertilizing capacity as well as seminal fructose, seminal level of reactive oxygen species (ROS) and pH. Results: Significant concentration and duration dependent decrease in sperm motility, vitality, normality and sperm fertilizing capacity was observed following incubation with candida. The drop in the percent of normal forms was within the acceptable range for normal semen. Level of ROS decreased significantly only at high candida concentration (100000 cfus/0.5 ml) and duration of incubation (15 hours). Seminal fructose was mainly affected by candida concentration while pH decreased only at low incubation time. However, the observed change in pH was still within the normal semen pH range. Relation between patient’s results and types of graft used showed no statistically significant differences between them. Conclusion: Vaginally extracted Candida albicans may have negative impact on different semen parameters affecting mainly sperm motility, vitality as well as sperm fertilizing capacity. To what extent this effect may influence couple fertility needs further studies. [ABSTRACT FROM AUTHOR]

The ricefield eel (Monopterus albus) is inherently timid and highly sensitive to stress. Our previous studies have shown that low-temperature weather could significantly affect the sperm vitality of ricefield eels. This study aims to investigate the regulatory mechanism of low-temperature effects on testicular function and sperm vitality in ricefield eels. The ricefield eels were initially reared at low (10 °C) and normal (25 °C) temperatures for 24 h. Low temperatures were found to induce the expression of pituitary pro-opiomelanocortin (POMC) and testes insulin-like growth factor-binding protein 1 (IGFBP1) mRNA expression, suggesting that the reduction in sperm vitality could be attributed to the activation of the stress axis. Moreover, the results indicated a significant decrease in sperm occupancy and count in the testes, along with a reduced percentage of motile sperm. Subsequent transcriptome analysis showed substantial inhibition of reproductive hormone genes (gnrh1, lh, and fsh) in the brain and pituitary, and downregulation of meiosis-related genes (dmc1, rec8, and sycp3) in the testes. These findings suggest that low temperatures might disrupt testicular development and spermatogenesis by inhibiting the reproductive axis. Metabolomics analysis then demonstrated a significant reduction in the levels of metabolites related to glycolysis, fatty acid metabolism, and the tricarboxylic acid (TCA) cycle in the testes after low-temperature treatment. Interestingly, the expression of zona pellucida sperm-binding proteins 3 and 4 (ZP3 and ZP4), which may affect sperm vitality and spermatogenesis, was significantly induced by low temperatures in the testes. In conclusion, these findings suggested that low temperatures might affect testicular function and sperm vitality by simultaneously activating the stress axis and inhibiting the reproductive axis and energy metabolism in the testes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

A basic semen investigation has established principles that are necessary for ascertaining reliable and internationally comparable results. Although these principles have been present in the WHO manual since its inception, the baseline issue across most published studies and practice in reproductive medicine (in which the male is considered) is repetitive failure to adhere to these principles, thereby leading to relevant comparable data and accuracy. To address this failure, the sixth edition of the WHO manual includes revised basic methods, and a complementary formal standard of the International Standards Organization (ISO23162:2021) for basic semen examination has been published. Perhaps the most significant change in the sixth edition is the reintroduction of the four-category distinction of sperm motility, which causes additional work for laboratories in changing reporting parameters but is clinically important. Another essential change is the widened focus from mainly a prognostic tool for medically assisted reproduction to additionally raising awareness of semen examination as a measure of male reproductive functions and general male health. [ABSTRACT FROM AUTHOR]

Objective: The aim of our study was to assess if the addition of PRGF to healthy human sperm affects its motility and vitality., Methods: This was a prospective study, with 44 sperm donors on whom sperm analysis was performed. Nine mL of blood was collected and PRGF was obtained using PRGF-Endoret® technology. The influence of different dilutions of PRGF (5%, 10%, 20%, 40%) applied to 15 sperm donors was compared, and sperm motility was assessed after 30 minutes. In the second part of the study, 29 sperm donors were studied to analyze the influence of 20% dilution of PRGF at 15, 30 and 45 minutes in fresh and thawed sperm samples. Motility was assessed after the addition of PRGF and after analysis each aliquot was frozen. After thawing, concentration and motility were assessed at the same time periods., Results: There were no differences in sperm motility in fresh samples between dilutions of PRGF when assessed 30 minutes after administration, nor between them, nor when compared to the control group immediately prior to treatment. No trend was observed between motility and PRGF dilution in linear regression analysis. There were no significant differences in thawed samples., Conclusions: The administration of 20% PRGF dilution had no effect on sperm motility compared to samples without PRGF. In addition, there was no change in sperm vitality when comparing samples with and without PRGF. More studies focusing on subnormal sperm samples, analyzing different PRGF concentrations and increasing the number of study variables are needed.

Lately, there is a systematic research consensus that reveals adverse effects of aspirin on semen quality characteristics; however, such consensus is lacking further confirmation by human studies. Therefore, here, we asked whether sperm motility and vitality are affected in the presence of aspirin at 0.1 and 1 mM in the ejaculated semen, and whether such effect may be due to an alteration in seminal calcium ions or seminal nitric oxide production. Forty‐three semen samples from different normozoospermic men were recruited in this study. Sperm motility was measured by Makler counting chamber, and sperm vitality was measured by Eosin test. Calcium chelating effect of aspirin and seminal nitric oxide production was measured spectrophotometrically. Aspirin at both tested concentrations significantly (p <.05) reduced progressive grade‐a motility and vitality of spermatozoa. Additionally, aspirin was found to have significant ability (p <.05) to bind seminal calcium ions, but insignificantly reduced the amount of seminal nitric oxide. In conclusion, sperm motility and vitality were reduced in the presence of aspirin at 0.1 and 1 mM in semen. Such reduction may be attributable to the ability of aspirin to chelate seminal calcium ions, but not to an alteration in the amount of nitric oxide produced. [ABSTRACT FROM AUTHOR]

Viability assessment is an important part of semen analysis, and various live/dead staining protocols have been used in semen of avian species. Results of live/dead count differed between dyes, staining protocols and bird species, impeding comparability between studies and requiring species‐specific comparisons of viability stains. In raptor semen, similar comparisons are absent. Thus, the aim of the present study was to compare eight conventional viability stains. Eosin blue 2% [EB], eosin blue 2% with the addition of 3% sodium citrate [EB2], eosin blue–nigrosin 5% [EBN5], eosin yellow–nigrosin 5% [EYN5], eosin yellow–nigrosin 10% [EYN10], eosin blue–aniline blue [EBA], eosin yellow–aniline blue [EYA] and bromophenol blue–nigrosin [BBN] were evaluated in comparison with the fluorescence stain SYBR® Green–propidium iodide [SYBR‐PI] in spermatozoa of falcons. The comparison was performed using conventional light microscopy which is applicable in breeding centres, veterinary practices and field studies. Additionally, live/dead stains were correlated to motility values of the same samples to validate sperm viability. Light microscopy using EB and using SYBR‐PI enabled an effective and clear differentiation between alive and dead spermatozoa of falcons. Motility values correlated significantly and strongly with EB only (r =.629; p <.001), but not with any other stain used in the study. Therefore, our results suggest EB as the most suitable stain for viability assessment in the semen of large falcons. [ABSTRACT FROM AUTHOR]

Background: Infertility affects 7%‐12% of men, and its etiology is unknown in half of cases. To fill this gap, use of the male genital tract color‐Doppler ultrasound (MGT‐CDUS) has progressively expanded. However, MGT‐CDUS still suffers from lack of standardization. Hence, the European Academy of Andrology (EAA) has promoted a multicenter study ("EAA ultrasound study") to assess MGT‐CDUS characteristics of healthy, fertile men to obtain normative parameters. Objectives: To report (a) the development and methodology of the "EAA ultrasound study," (b) the clinical characteristics of the cohort of healthy, fertile men, and (c) the correlations of both fertility history and seminal features with clinical parameters. Methods: A cohort of 248 healthy, fertile men (35.3 ± 5.9 years) was studied. All subjects were asked to undergo, within the same day, clinical, biochemical, and seminal evaluation and MGT‐CDUS before and after ejaculation. Results: The clinical, seminal, and biochemical characteristics of the cohort have been reported here. The seminal characteristics were consistent with those reported by the WHO (2010) for the 50th and 5th centiles for fertile men. Normozoospermia was observed in 79.6% of men, while normal sperm vitality was present in almost the entire sample. Time to pregnancy (TTP) was 3.0[1.0‐6.0] months. TTP was negatively correlated with sperm vitality (Adj.r =−.310, P =.011), but not with other seminal, clinical, or biochemical parameters. Sperm vitality and normal morphology were positively associated with fT3 and fT4 levels, respectively (Adj.r =.244, P <.05 and Adj.r =.232, P =.002). Sperm concentration and total count were negatively associated with FSH levels and positively, along with progressive motility, with mean testis volume (TV). Mean TV was 20.4 ± 4.0 mL, and the lower reference values for right and left testes were 15.0 and 14.0 mL. Mean TV was negatively associated with gonadotropin levels and pulse pressure. Varicocoele was found in 33% of men. Conclusions: The cohort studied confirms the WHO data for all semen parameters and represents a reference with which to assess MGT‐CDUS normative parameters. [ABSTRACT FROM AUTHOR]

How many spermatozoa and slides need to be counted to make a reliable assessment of sperm vitality? Currently, various authorities recommend assessing human sperm vitality on counts of at least 200 cells, but on one or two slides. This was an observational study on duplicate eosin–nigrosin stained sperm vitality slides made from 58 ejaculates. Assessments were made using counts of 2 × 100 and 1 × 200 cells per slide, all performed by the same trained expert observer. Although assessments tend to show fewer and smaller outlier values when based on counts of 200 spermatozoa than 100, and on 2 × 200 than 1 × 200, counting 200 spermatozoa from one slide provides a result with sufficient accuracy for the clinical purpose of ascertaining whether the immotile spermatozoa in an ejaculate showing low sperm motility are alive or dead. While the increased accuracy of results derived from counts of 2 × 200 cells might be important in research studies where sperm vitality is the specific end-point of interest, counting at least 200 spermatozoa from one smear is sufficiently accurate for the clinical purpose of establishing whether the immotile spermatozoa seen in ejaculates with low sperm motility are alive or not. Consequently, the extra workload of performing replicate counts and the associated calculations does not increase the clinical value of the result, and hence is unnecessary in routine semen analysis. Careful laboratory technique as well as proper staff training and competence are essential. The conclusions might not be applicable to staining methods other than the recommended one-step eosin–nigrosin technique. [ABSTRACT FROM AUTHOR]

In this study, we sought to investigate the effect of lifestyle and demographic factors on classic and functional semen parameters. Three hundred and twenty-eight subjects who underwent semen analysis were recruited. Routine SA, sperm vitality, acrosome reaction (AR) assay and sperm DNA fragmentation index (DFI) were analyzed. Demographic and lifestyle information, including (1) BMI, (2) current smoking and alcohol drinking frequency, (3) sleep habits, (4) daily fluid intake, (5) weekly meat intake, (6) sports frequency, (7) trouser cell phone use, (8) age, and (9) abstinence time, were collected. Generalized additive models were used to analyze the possible non-linear association. The results showed that total sperm count (TSC) was significantly associated with age (P = 0.001), abstinence time (P = 0.001) and daily coffee intake (P = 0.044). Semen volume was significantly associated with age (P < 0.001) and daily coffee intake (P < 0.001). Sperm concentration was significantly associated with abstinence time (P = 0.011) and average sleep duration (P = 0.010). Sperm motility was significantly associated with age (P = 0.002) and daily juice intake (P = 0.001). Total motile sperm count was significantly associated with age (P = 0.003) and abstinence time (P = 0.009). DFI was significantly associated with age (P = 0.002), irregular sleeping habit (P = 0.008) and abstinence time (P = 0.032). The percentage of AR sperm was significantly associated with daily juice intake (P = 0.013). In conclusion, DFI and TSC were the most sensitive semen parameters for demographic and lifestyle features, whereas age had more influence on semen parameters than other demographic and lifestyle features. Abbreviations: BMI: body mass index; SA: semen analysis; AR: acrosome reaction; DFI: DNA fragmentation index; GAM: generalized additive model; TSC: total sperm count; TMC: total motile sperm count; IUI: intrauterine insemination; SCSA: sperm chromatin structure assay; SD: standard deviation; IQR: interquartile range; CBAVD: congenital bilateral absence of vas deferens; NEQAS: national external quality assessment service; HTF: human tubal fluid; HSA: human serum albumin. [ABSTRACT FROM AUTHOR]

Many male patients diagnosed with cancer are within their reproductive years. These men are advised to freeze their spermatozoa prior to the start of cancer treatment. Very often, sperm banking facilities may not be readily available and patients may be required to travel to distant sperm bank centres. Our objective was to design and standardise a remote home shipping sperm kit that allows patients to collect a semen sample at home and ship it overnight to a sperm bank. A total of 21 semen samples and two transport media (refrigeration media and human tubal fluid) and five different combinations of ice packs were tested for maintaining desired shipping temperature. Ten semen samples were assessed for pre- and post-shipment changes in sperm motility, membrane integrity, total motile spermatozoa and recovery of motile spermatozoa. Even though motility, membrane integrity and total motile spermatozoa declined both in samples examined under simulated shipped conditions and in overnight-shipped samples, the observed motility and total motile spermatozoa were adequate for use with assisted reproductive techniques. Using refrigeration media, cooling sleeve and ice packs, adequate sperm motility can be maintained utilising NextGen ® kit and these spermatozoa can be used for procreation utilising ART techniques such as intracytoplasmic sperm injection. [ABSTRACT FROM AUTHOR]

The effect of some pecies of bacteria and sperm/bacteria ratio on sperm vitality has been studied. Four species of bacteria were used in this study: Stapylococcus epidermidis, Streptococcus faecalis, Enterobacter aerogenes were obtained from semen culture of infertile men and E. coli was obtained from prostaltic fluid culture from men with prostat and urinary system disturbances. Five semen samples fulfilling the WHO criteri (1992) were used in this study. After preparation by Percoll gradient-column method, sperm were inoculated in a microplate with Stapylococcus epidermidis, Streptococcus faecalis, Enterobacter aerogenes and E. coli under the sperm/bacteria ratio 1:10 and 1:1000. Sperm vitality was observed immediately, 3 and 6 hours after inoculation. At the second experiment, the detrimental influence of bacteria on sperm was prevented by adding penicillin. Result of this study indicated that Stapylococcus epidermidis, Streptococcus faecalis, and Enterobacter aerogenes were not affected on the sperm vitality. The effect of E. coli on sperm vitality occurred at the ratio of sperm/bacteria 1:10 after 3 and 6 hours incubation and at the ratio of sperm/bacteria 1:1000 occurred after 6 hours incubation. It might be concluded that the negative influence of bacteria on sperm vitality in vitro, is dependent in species of bacteria, bacteria concentration, and time of incubation. The most detrimental effect on sperm vitality was shown by E. coli at the ratio of sperm/bacteria 1:10 after 6 hours incubation. The detrimental effect was not prevented by the addition of penicillin.

Background/aim: About 10%-15% of couples around the world sufer from infertility. Male infertility is responsible directly or indirectly in ~60% of cases. A deiciency in semen is the most common cause of male infertility. Materials and methods: he study included 180 male subjects aged 18-50 years with 26 fertile and 154 infertile. he infertile subjects were further subdivided according to the WHO guidelines of semen analysis (2010) into diferent clinical groups. Sperm DNA damage was estimated using a neutral comet assay. Plasma gonadotropin and testosterone levels were measured using a chemiluminescence assay. Results: he results of the study revealed no signiicant diferences in semen volume, pH, and liquefaction time between the fertile and all infertile groups. However, sperm concentration, sperm vitality, and sperm motility were signiicantly lower in all infertile groups as compared to the fertile males. he morphological forms of the sperm and its DNA fragmentation varied signiicantly between the fertile and infertile males. Reproductive hormone levels were observed to be signiicantly lower in the infertile than in the fertile males. Conclusion: Sperm DNA fragmentation was higher in all of the infertile subjects as compared to the fertile ones. Reproductive hormone levels varied signiicantly between the infertile patients and the fertile ones. [ABSTRACT FROM AUTHOR]

BACKGROUND Clusterin, a heterodimeric glycoprotein found at several sites in the human male reproductive tract, could be a marker of morphologically abnormal spermatozoa, while TUNEL positivity indicates DNA fragmentation. Metabolic disorders such as diabetes mellitus and obesity may compromise sperm quality and fertility of men; however, little evidence specifically links hypertension with the impairment of male reproductive function. METHODS By flow cytometric, immunofluorescence (TUNEL assay and clusterin immunolabeling) and immunohistochemical (peroxidase-streptavidin method) analyses, we have compared both clusterin- and TUNEL labeling in ejaculated spermatozoa from healthy normotensive donors and hypertensive subjects with the purpose to reveal possible differences between the two conditions. RESULTS Data analysis from the normotensive (n = 25) and hypertensive subjects (n = 25) demonstrate a significant correlation between high levels of clusterin immunolabeling and the presence of sperm DNA damage, which is often associated with abnormal morphology. In the normotensive subjects, a low percentage (15.3 ± 4.5) of spermatozoa positive for high levels of clusterin was detected; however, this percentage significantly increased (30.9 ± 13.0) (P < 0.01) in hypertensive subjects. Standard semen evaluations does not reveal any significant differences between the two groups of subjects, except for a reduced forward motility and lower sperm vitality in the hypertensive subjects. CONCLUSIONS This pilot study strongly suggests a relationship between hypertension and markers indicative of poor sperm quality. In hypertensive subjects, high levels of clusterin immunolabeling identified a consistent fraction of ejaculated spermatozoa carrying both DNA fragmentation and strong morphological alterations, which was not correlated with age or with sperm cell mortality. The alternative possibility that sperm damage observed is due to adverse effects of anti-hypertensive drugs does not find support in the literature nor in the drug data sheets. The relationship observed between hypertension and human semen represents a novel and possibly relevant information to be considered in the study of male fertility. [ABSTRACT FROM AUTHOR]

The effect of sample collection site on semen parameters in ten men aged between 22 and 24 years was investigated. Sperm was collected at two sites: in a university hospital restroom for general use and in a one-person hospital room. Samples were collected from the same individual twice, with an interval of two weeks between collections. Semen parameters for the two sites were compared. Samples were collected after a minimum of three days and not longer than seven days of sexual abstinence. Sperm concentration did not differ significantly between the university hospital restroom location (86.8 ± 25.4 × 106/ml; mean ± standard deviation) and the private hospital room (97.1 ± 72.0 × 106/ml). There was no difference in the total motile sperm count or daily sperm production between the collection sites. These results suggest that the collection site has little effect on semen parameters. [ABSTRACT FROM AUTHOR]

Individual variation in semen parameters was investigated in healthy young volunteers. Semen samples were collected approximately once a month over a one-year period for a total of 93 samples (5 to 10 samples per subject) from 12 volunteers in their twenties. Semen analysis was carried out according to the WHO Manual. The amount of variation in each semen variable was calculated for each subject by dividing the maximum value by the minimum value. The results showed that the semen volume varied by 1.9&ThickSpace;±&ThickSpace;0.8 fold (1.3 to 4.2 fold), the sperm concentration by 4.8&ThickSpace;±&ThickSpace;4.3 fold (1.5 to 17.2 fold), the percentage of sperm with forward progression by 2.8&ThickSpace;±&ThickSpace;1.4 fold (1.6 to 6.4 fold), the percentage of sperm with rapid linear progression by 3.4&ThickSpace;±&ThickSpace;2.6 fold (1.7 to 10.9 fold), the percentage of sperm with normal morphology by 1.9&ThickSpace;±&ThickSpace;0.4 fold (1.3 to 2.4 fold), and the percentage of live sperm by 1.5&ThickSpace;±&ThickSpace;0.4 fold (1.1 to 2.6 fold). A between-group comparison showed significant differences in all of the variables except the percentage of sperm with normal morphology. These results suggest multiple and considerable semen analyses are needed when evaluating semen parameters. [ABSTRACT FROM AUTHOR]

Proficiency testing samples for antisperm antibodies (ASAB), sperm count, morphology and vitality were mailed to participating laboratories. The majority participating utilized Immunobead ASAB procedures (81 versus 14% mixed antiglobulin reaction and 5% `other'), and there was 95.6 ± 1.2% agreement on the presence or absence of ASAB. The majority of laboratories utilized manual (79%) versus computer assisted semen analysis (CASA; 15%) methods. Approximately 64% used the haemocytometer and 26% used the Makler counting chambers for manual counts. Coefficients of variation (CV) in sperm counts ranged from 24 to 138%, with CASA displaying lower overall CV (53 ± 8%) than manual methods (80 ± 9%). A wide variation in the reports of percent normal morphology was noted (CVs calculated from arc sin transformed means ranged from 15 to 93%). Participants using American Society of Clinical Pathologists (ASCP) criteria reported sperm morphology values that were clustered in the `normal' range (11 out of 12 samples), while those using strict criteria were clustered in the `abnormal' range (10 out of 12 samples). Good agreement was observed in sperm vitality (overall mean CV = 18%). These data highlight the urgent need for improvement in overall quality of andrology testing and indicate that practical proficiency testing programmes can be made available on a large scale. [ABSTRACT FROM PUBLISHER]

The first two editions of the World Health Organization laboratory manual described the determination of live spermatozoa by a dye exclusion method as a sperm "viability" test, whereas subsequent editions classified it as a "vitality" test, without providing an explanation for the reclassification. Additionally, the hypo-osmotic swelling (HOS) test, which assesses the functional integrity of the human sperm membrane, was placed in the same category as the dye exclusion test. Although the two terms might seem synonymous, the term "vitality" merely means "alive," whereas "viability" assesses qualities or physiological functions of a living entity. After comparing the morphological, physiological, and clinical findings obtained from dye exclusion testing vs. the HOS test, we conclude that the HOS test should be classified as a viability test, not merely as a vitality test., (Copyright © 2021 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

C 60 fullerene (C 60) is a nano-pollutant that can damage the respiratory system. Eugenol exhibits significant anti-inflammatory and antioxidant properties. We aimed to investigate the time course of C 60 emulsion-induced pulmonary and spermatic harms, as well as the effect of eugenol on C 60 emulsion toxicity. The first group of mice (protocol 1) received intratracheally C 60 emulsion (1.0 mg/kg BW) or vehicle and were tested at 12, 24, 72 and 96 h (F groups) thereafter. The second group of mice (protocol 2) received intratracheally C 60 emulsion or vehicle, 1 h later were gavaged with eugenol (150 mg/kg) or vehicle, and experiments were done 24 h after instillation. Lung mechanics, morphology, redox markers, cytokines and epididymal spermatozoa were analyzed. Protocol 1: Tissue damping (G) and elastance (H) were significantly higher in F24 than in others groups, except for H in F72. Morphological and inflammatory parameters were worst at 24 h and subsequently declined until 96 h, whereas redox and spermatic parameters worsened over the whole period. Eugenol eliminated the increase in G , H , cellularity, and cytokines, attenuated oxidative stress induced by C60 exposure, but had no effect on sperm. Hence, exposure to C 60 emulsion deteriorated lung morphofunctional, redox and inflammatory characteristics and increased the risk of infertility. Furthermore, eugenol avoided those changes, but did not prevent sperm damage. Image 1 • C60 emulsion impaired lung morphofunctional and inflammatory status (peak at 24 h). • A single dose of C60 emulsion generated sperm toxicity. • Eugenol avoided lung morphofunctional, redox and inflammatory impairment. [ABSTRACT FROM AUTHOR]

In order to address the large percentage of unexplained male infertility in humans, more detailed investigations using sperm functional tests are needed to identify possible causes for compromised fertility. Since many environmental and lifestyle factors might be contributing to infertility, future studies aiming to elucidate the effect of such factors on male fertility will need the use of appropriate research models. The current study aimed to assess the effects of two heavy metals, namely copper sulphate, and cadmium chloride, on non-human primate (NHP) sperm function in order to establish the possibility of using these primate species as models for reproductive studies. Our combined results indicated that the functionality of NHP spermatozoa is inhibited by the two heavy metals investigated. After in vitro exposure, detrimental effects, and significant lowered values ( p < 0.05) were obtained for sperm motility, viability and vitality, acrosome intactness, and hyperactivation. These metals, at the tested higher concentrations, therefore, have the ability to impair sperm quality thereby affecting sperm fertilizing capability in both humans and NHPs.

C 60 fullerene (C 60 ) is a nano-pollutant that can damage the respiratory system. Eugenol exhibits significant anti-inflammatory and antioxidant properties. We aimed to investigate the time course of C 60 emulsion-induced pulmonary and spermatic harms, as well as the effect of eugenol on C 60 emulsion toxicity. The first group of mice (protocol 1) received intratracheally C 60 emulsion (1.0 mg/kg BW) or vehicle and were tested at 12, 24, 72 and 96 h (F groups) thereafter. The second group of mice (protocol 2) received intratracheally C 60 emulsion or vehicle, 1 h later were gavaged with eugenol (150 mg/kg) or vehicle, and experiments were done 24 h after instillation. Lung mechanics, morphology, redox markers, cytokines and epididymal spermatozoa were analyzed. Protocol 1: Tissue damping (G) and elastance (H) were significantly higher in F24 than in others groups, except for H in F72. Morphological and inflammatory parameters were worst at 24 h and subsequently declined until 96 h, whereas redox and spermatic parameters worsened over the whole period. Eugenol eliminated the increase in G, H, cellularity, and cytokines, attenuated oxidative stress induced by C60 exposure, but had no effect on sperm. Hence, exposure to C 60 emulsion deteriorated lung morphofunctional, redox and inflammatory characteristics and increased the risk of infertility. Furthermore, eugenol avoided those changes, but did not prevent sperm damage., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

One known environmental risk factor impacting on human reproduction is heavy metal pollution. Although some metals (e.g., Cu, Se and Zn) have protective effects on the male reproductive system in low doses, heavy metals can accumulate to toxic levels and result in poor semen quality and decreased sperm function. We investigated the effect of CuSO4 and CdCl2 (10, 50, 100 and 250 µg/ml or 500 µg/ml) on human sperm motility and vitality by using computer‐aided sperm analysis (CASA) and two cytotoxicity assays (WST‐1 and XTT). Several sperm motility parameters were significantly reduced after 5 hr of exposure to the highest concentrations of CuSO4 (250 µg/ml) and CdCl2 (500 µg/ml). The WST‐1 assay also revealed significantly lower absorbance values for 50, 100 and 250 µg/ml CuSO4 and for 500 µg/ml CdCl2; however, no significant effect was seen with XTT. The calculated average IC50 value was 50.31±  4.34 µg/ml for CuSO4 and 392.32  ±76.79 µg/ml for CdCl2. The effects of these metals were confirmed with MgCl2, a positive control. This study provides threshold concentrations for the harmful effect of CuSO4 and CdCl2 on human spermatozoa and recommends the use of WST‐1 as vitality assay in future in vitro studies. [ABSTRACT FROM AUTHOR]

Many male patients diagnosed with cancer are within their reproductive years. These men are advised to freeze their spermatozoa prior to the start of cancer treatment. Very often, sperm banking facilities may not be readily available and patients may be required to travel to distant sperm bank centres. Our objective was to design and standardise a remote home shipping sperm kit that allows patients to collect a semen sample at home and ship it overnight to a sperm bank. A total of 21 semen samples and two transport media (refrigeration media and human tubal fluid) and five different combinations of ice packs were tested for maintaining desired shipping temperature. Ten semen samples were assessed for pre- and post-shipment changes in sperm motility, membrane integrity, total motile spermatozoa and recovery of motile spermatozoa. Even though motility, membrane integrity and total motile spermatozoa declined both in samples examined under simulated shipped conditions and in overnight-shipped samples, the observed motility and total motile spermatozoa were adequate for use with assisted reproductive techniques. Using refrigeration media, cooling sleeve and ice packs, adequate sperm motility can be maintained utilising NextGen(®) kit and these spermatozoa can be used for procreation utilising ART techniques such as intracytoplasmic sperm injection., (© 2015 Blackwell Verlag GmbH.)