Difference In Sperm Motility Research Articles

Trehalose, Glycerol and Dimethyl Sulfoxide Supplementation for Cryopreservation Media of Honey Bee Drone Semen

The cryogenic storage of honey bee drone (Apis mellifera) semen is a rapidly developing practice, driven by the need to preserve bee colonies and the decline in gamete quality following cryopreservation. Advances in cryobiology, coupled with procedural refinements and the development of new methods for cryopreservation of male gametes at different temperatures, provide promising results for improving drone semen preservation. Innovative strategies involve the incorporation of additives within preservation media to enhance important spermatozoa parameters. Our study aimed to assess the effectiveness of cryoprotectants (CPAs), namely trehalose, glycerol, and dimethyl sulfoxide (DMSO), on the motion characteristics of honey bee drone sperm cells during cryopreservation and subsequent thawing. Using Sperm Class Analyzer (SCA) and Computer-Assisted Sperm Analysis (CASA), we evaluated the impact of these supplements on sperm motility and kinetic parameters. Our findings indicated significant differences in sperm motility (non-progressiveness and progressiveness) and kinetics (curvilinear velocity and linearity) across treatments with trehalose, DMSO, and glycerol. Trehalose supplementation retained significantly higher percentages of sperm motility and kinetic parameters post-cryopreservation compared to DMSO and glycerol. Nevertheless, despite the utilization of new protocols, media, and supplementation, the results obtained are often unsatisfactory. The research provides valuable information for enhancing the efficacy of cryopreservation media to optimize the successful outcome of honey bee drone spermatozoa storage.

Effect of seawater temperature and pH on the sperm motility of the European eel.

The current climate change situation could bring critical effects for marine species, especially those already considered endangered. Although some species can adapt fast to the environmental changes, it is necessary to get into the worst scenario and develop tools to anticipatedly assess the physiological effects of such environmental change. With this purpose, our study aims to determine the effect of a range of seawater temperatures and pHs on sperm motility in the European eel (Anguilla anguilla). Low seawater pH (6.5-7.4) decreased the eel sperm motility in comparison to the control (pH = 8.2). We also studied the combined effect of the pH of the artificial seminal plasma (the plasma where the sperm cells are suspended) with the pH of Artificial Sea Water (ASW, pH 7.8 or and 8.2). We did not find statistical differences in sperm motility and kinetic parameters caused by the artificial seminal plasma pH. However, seawater pH induced significantly higher values of total sperm motility, and the percentage of fast spermatozoa with a pH of 8.2 in comparison with a pH of 7.8. In contrast, the seawater temperature did not affect sperm motility parameters or sperm longevity. To study the effect of the interaction between seawater temperature and pH on sperm motility, two temperatures: 4 and 24°C, and two pHs 7.8 and 8.2, were tested. There were significant differences between temperature and pH in several kinetic parameters and, in general, the lowest values were observed in the samples activated at low temperature and low pH (4°C, pH 7.8). This work suggest that eel sperm motility and kinetics will not be affected by the expected changes in pH or temperature due to the climate change.

The Drosophila melanogaster Y-linked gene, WDY, is required for sperm to swim in the female reproductive tract

Unique patterns of inheritance and selection on Y chromosomes have led to the evolution of specialized gene functions. We report CRISPR mutants in Drosophila of the Y-linked gene, WDY, which is required for male fertility. We demonstrate that the sperm tails of WDY mutants beat approximately half as fast as those of wild-type and that mutant sperm do not propel themselves within the male ejaculatory duct or female reproductive tract. Therefore, although mature sperm are produced by WDY mutant males, and are transferred to females, those sperm fail to enter the female sperm storage organs. We report genotype-dependent and regional differences in sperm motility that appear to break the correlation between sperm tail beating and propulsion. Furthermore, we identify a significant change in hydrophobicity at a residue at a putative calcium-binding site in WDY orthologs at the split between the melanogaster and obscura species groups, when WDY first became Y-linked. This suggests that a major functional change in WDY coincided with its appearance on the Y chromosome. Finally, we show that mutants for another Y-linked gene, PRY, also show a sperm storage defect that may explain their subfertility. Overall, we provide direct evidence for the long-held presumption that protein-coding genes on the Drosophila Y regulate sperm motility.

Safety of Plasma Rich in Growth Factors (PRGF) as additive to healthy human sperm samples: a pilot study.

The aim of our study was to assess if the addition of PRGF to healthy human sperm affects its motility and vitality. 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. 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. 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.

Performance evaluation of sperm concentration, motility, and morphological analysis for GSA-810 series of sperm quality analysis system.

The performance evaluation of each computer-assisted sperm analysis (CASA) system may provide a basis for the interpretation of clinical results and further improvement of the CASA system. The accuracy of the GSA-810 CASA system was evaluated by detecting latex bead quality control products. The precision of sperm concentration, morphology, and percentages of progressively motile sperm (PR) were evaluated by coefficient of variation (CV). Three samples with sperm concentration of about 100 × 106 /mL were diluted to evaluate the linear range. The detection values of latex beads were within the range of target values. The CVs of sperm concentration and PR were significantly and negatively correlated with sperm concentration (r = -0.561, p = 0.001) and PR value (r = -0.621, p < 0.001), respectively. The R2 values of the linear range of sperm concentration were ≥0.99. There was no significant difference in sperm motility and PR within 1-10 min at 36.5°C ± 0.5°C. The coincidence rates of sperm morphology and sperm head morphology for 36 semen samples analyzed by the GSA-810 system and manual method were 99.40% and 99.67%, respectively. The CVs of the percentage of sperm with abnormal morphology and percentage of sperm with abnormal head morphology were less than 5%. The GSA-810 system can accurately analyze normal semen samples, but the repeatability of the results is poor for oligozoospermia and asthenozoospermia samples. The future CASA system for analyzing sperm morphology should focus on recognizing the middle and tail segments of a spermatozoon.

Antioxidant Activity of Fermented Red Bean Extract on Sperm Quality of Mice Exposed to Cigarette Smoke

Smoking has a negative effect on fertilization as it decreases sperm quality. The current research aimed to investigate the effect of fermented red bean (Phaseolus vulgaris L.) extract on sperm motility, viability, and plasma membrane integrity of white mice (Mus musculus) exposed to cigarette smoke. The red beans were subjected to a 36-hour fermentation process using Rhizopus spp. Then, methanol was extracted by maceration method for 24 hours until maceration was obtained. In this study, 25 male white mice aged 3 months were randomly divided into 5 groups of 5 mice. Group C (negative control) was given 0.5 mL of carboxymethyl cellulose natrium (CMC Na) 1% solution orally without unfiltered cigarette smoke exposure, and group C+ (positive control) was given 0.5 mL of CMC Na 1% solution orally and unfiltered cigarette smoke exposure. Treatment groups T1, T2, and T3 were orally given fermented red bean extract at doses of 26 mg/kg Body weight (BW), 52 mg/kg BW, and 104 mg/kg BW, respectively, and then were exposed to unfiltered cigarette smoke. For 36 days, treatment groups (except the negative control) were subjected to the inhalation of smoke from an unfiltered cigarette containing a nicotine dose of 2.2 mg. The exposure period lasted for 20 minutes each day. Each group was put into a cigarette smoke-exposing box. The sperm motility (observing the forward movement of spermatozoa), the sperm viability (examining the color of the sperm head), and the sperm plasma membrane integrity (observing the tail shape using the hypoosmotic swelling test) were then evaluated. The findings indicated significant differences in sperm motility, viability, and plasma membrane integrity of each group with positive control. A dose of 104 mg/kg BW of fermented red bean extract had the best potential to maintain sperm motility (70%), viability (82.13%), and plasma membrane integrity (61.93%) of mice exposed to unfiltered cigarette smoke.

Comparison of Pregnancy Outcomes in Thyroid Antibody-Positive Women with Different Fertilization Methods

<i>Objective</i> To compare the pregnancy outcomes of thyroid antibody (ATA) positive women treated by different fertilization methods during in vitro fertilization-embryo transfer. Methods A total of 204 female patients with normal thyroid function and positive ATA in the reproductive Medicine Center of the First Hospital of Shanxi Medical University from January 2020 to January 2022 were selected. According to different fertilization methods, they were divided into IVF group and ICSI group, including 115 IVF cycles and 89 ICSI cycles. The general data, clinical indicators after ovulation induction, fertilization, embryo status, and pregnancy outcome of the two groups were compared. <i>Results</i> There were no significant differences in age, BMI, bE₂, FSH, FT₃, FT₄, AFC, Gn dosage, Gn stimulation time, E₂ on HCG day, P on HCG day, intrauterine thickness on HCG day, number of retrieved oocytes and average number of embryos transferred between the two groups (P>0.05). The fertilization rate of the ICSI group was higher than that of IVF group within the group, while the high-quality embryo rate was significantly lower than that of the IVF group, with statistical significance (P<0.05). There were significant differences in sperm motility and sperm density between the two groups (P<0.05), and there was no significant difference between sperm volume and normal morphology rate (P>0.05). There were no significant differences in embryo implantation rate, biochemical pregnancy rate, clinical pregnancy rate, early abortion rate and live birth rate between the two groups (P>0.05). Logistic regression analysis showed that the method of assisted pregnancy and semen quality were not independent factors affecting high-quality embryo (P>0.05). In IVF group, TGAb was negatively correlated with fertilization rate (r=-0.202, P<0.05); and in ICSI group, TPOAb was negatively correlated with high quality embryo rate (r=-0.349, P<0.05). <i>Conclusion</i> Thyroid autoimmunity (TAI) in infertile women undergoing assisted reproductive technology (ART), the semen quality should be considered as the priority criteria for ATA. ICSI should not be recommended as routine fertilization.

Safety Evaluation of bioactive Sub-Fraction of Lawsonia inermis Linn leaves in Male Wistar Rats

This study evaluated the safety of bioactive labelled sub-fraction of Lawsonia inermis in male Wistar rats. Twenty male rats ‎were used for this study of five rats per group. Three groups were administered methanol bioactive sub-fraction of Lawsonia ‎inermis at 2.5, 5 and 10 mg/kg for fourteen days while control received distilled water orally. Physiological weight increased in ‎all treatment groups. The group fed with 10 mg/kg had the highest weight gain compared to other treatment groups and the ‎control. Relative organ weight of the heart, liver, kidneys, testes and pancreas did not increase (p&gt;0.05) in all the treatment ‎groups except the pancreas in the group fed with 5 mg/kg that increased significantly (p&lt;0.05). The sperm count in rats fed at ‎dose rate of 10 mg/kg increased significantly (p&lt;0.01) compared to other treatment groups. There were no significant ‎differences in sperm motility, volume and live/dead ratio in the treatment groups compared to control. Haematological ‎parameters such as, PCV, RBC, WBC and haemoglobin concentration were not significantly different compared to control. ‎The AST increased significantly (p&lt;0.01) while urea and creatinine decreased significantly (p&lt;0.001) in group fed with 2.5 ‎mg/kg. 5 mg/kg showed significant reduction (p&lt;0.05) in total protein, urea and ALP while ALT increased significantly. Serum ‎chloride ion at the dose rate of 2.5 mg/kg decreased significantly (p&lt;0.05) when compared with other treatment groups. ‎Sodium ion (Na+) decreased significantly (p&lt;0.01), while Bicarbonates (HCO3-) increased significantly in the 5 mg/kg group. It ‎was concluded that the bioactive sub-fraction of Lawsonia. inermis was safe at the administered dosages.‎

Effects of preconception exposure to phthalates on mouse sperm capacitation parameters.

Phthalates have been linked to adverse male reproductive health, including poor sperm quality and embryo quality as well as a longer time to pregnancy (months of unprotected intercourse before conception occurs). The present study aimed to evaluate the effect of preconception exposure to two ubiquitous phthalate chemicals, di(2-ethylhexyl) phthalate (DEHP), di-n-butyl phthalate (DBP), and their mixture on sperm function, fertilization, and embryo development in mice. Adult male C57BL/6Jmice aged 8-9weeks were exposed to di(2-ethylhexyl) phthalate, di-n-butyl phthalate, or their mixture (di-n-butyl phthalate+di(2-ethylhexyl) phthalate) at 2.5mg/kg/day or vehicle for 40days (equivalent to one spermatogenic cycle) via surgically implanted osmotic pumps. Caudal epididymal spermatozoa were extracted and analyzed for motility using computer-assisted sperm analyses. Sperm phosphorylation of protein kinase A substrates and tyrosine phosphorylation, markers of early and late capacitation events, respectively, were analyzed by Western blots. In vitro fertilization was used to evaluate the sperm fertilizing capacity. While the study did not reveal any significant differences in sperm motility and fertilization potential, abnormal sperm morphology was observed in all phthalate exposures, particularly in the phthalate mixture group. In addition, the study revealed significant differences in sperm concentration between control and exposed groups. Moreover, protein phosphorylation of protein kinase A substrates was decreased in the di(2-ethylhexyl) phthalate and mixture exposure groups, while no significant changes in protein tyrosine phosphorylation were observed in any of the groups. Assessment of the reproductive functionality did not reveal significant effects on in vitro fertilization and early embryo development rates but showed wide variability in the phthalate mixture group. Our findings suggest that preconception phthalate exposure affects sperm numbers and phosphorylation of protein kinase A substrates involved in capacitation. Future research is warranted to examine the associations between phthalate exposure and capacitation in human spermatozoa.

TLR7/8 agonist (R848) inhibit bovine X sperm motility via PI3K/GSK3α/β and PI3K/NFκB pathways

Sex-control technology have great economic value and is one of the hot topics in livestock research. To produce more milk, dairy farmers prefer female offspring. X/Y sperm separation is an effective method for offspring sex control. Currently, the major commercial production method for sperm separation is flow cytometry sorting in cattle. However, flow cytometry requires expensive equipment and long sorting times. So, a simple and inexpensive method for producing a higher number of dairy cows is required. In this study, R848 activates toll-like receptor 7/8 (TLR7/8), thereby separating X from Y sperm. The results showed TLR7/8 is expressed in the tail of X sperm. Immunofluorescence (IF) of testes, epididymis, and ejaculate shows that the number of TLR7+/8+ sperm cells is up to 50 %. Furthermore, TLR7/8 agonist (R848) affects mitochondrial function through the PI3K/GSK3α/β/hexokinase and PI3K/NFκB/hexokinase signalling pathways, inhibiting X sperm motility, while the motility of Y-sperm remains unchanged. The difference in sperm motility causes Y sperm (with high motility) to move to the upper layer and X-sperm (with low motility) to the lower layer allowing the separation of X and Y sperm. Based on this study, we reveal a simple and effective method for enriched X/Y sperms from cattle.

The effect of individual selection on sperm motility in tropical abalone Haliotis squamata

High-quality broodstock is required to support successful cultivation activities of the tropical abalone. This study aimed to find out the structure of the sperm under the influence of individual selection. A total of 50 eggs per batch were measured in egg diameter, calculated by the degree of fertilization and hatching rate. Sperm collection is carried out from the natural spawning of male abalone from the wild (F0) and the selected offspring from individual Fillial-1, F2, and F3 selections. Samples of sperm were collected by artificial spawning and analyzed using the Sperm Class Analyzer® CASA System. The results showed that the wild abalone (F0) had the highest hatching rate but was not significantly different from other derivatives (P &gt; 0.05). The average spermatozoa was 2.39 million cells/m, with the highest spermatozoa (F0) of 7.110 million cells/ml. The most increased sperm motility and rapid velocity were in the wild (89.78%; 34.00%) and the lowest of F2 (22.00; 4.00%). A significant difference in sperm motility and rapid velocity were found between F0 and other treatment. The consequences of individual selection will aid in studying reproductive factors and sperm motility, which are most likely crucial in tropical abalone fertilization.

Quality of cauda epididymal sperm immediately after collection and after freezing-thawing from Amur leopard cats (Prionailurus bengalensis euptilurus) and a local population of the subspecies Tsushima leopard cats.

In this study, cauda epididymal sperm were collected from Amur leopard cats with various causes of death as well as Tsushima leopard cats that underwent castration surgery, and sperm quality was compared with that in domestic cats. A sufficient number of sperm similar to those in domestic cats could be collected from the cauda epididymis of Amur leopard cats. However, in old leopard cats, no or very few cauda epididymal sperm were recovered, although there were no differences in sperm motility and sperm abnormality. There were no significant differences in sperm quality immediately after collection and after freezing-thawing of cauda epididymal sperm compared with corresponding estimates in domestic cats.

The Quality of Fresh and Frozen Semen and its Correlation with Molecular Weight of Seminal Plasma Protein in Bali Cattle

This study evaluated the quality of fresh and frozen semen of Bali cattle and its correlation with the molecular weight (MW) of seminal plasma protein. This study collected semen from 10 bulls aged 5–10 years using an artificial vagina and evaluated the samples macroscopically and microscopically. Two batches of frozen semen obtained in 2020 and 2021 were also analyzed. The frozen semen samples were thawed at 37 °C for 30 seconds. The sperm motility, viability, intact plasma membrane (IPM), and sperm abnormalities were investigated. The concentration of the seminal plasma proteins was determined using the Bradford method, and the proteins were characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D-SDS-PAGE). Additionally, the gels were stained with Coomassie brilliant blue, and the MWs of the proteins were determined using MW markers. The sperm motility, viability, and abnormalities of fresh semen varied significantly among the bulls (p<0.05); however, sperm IPMs among the bulls were similar (p>0.05). No differences in sperm motility after freezing were found among the bulls. However, the sperm viability, abnormality, and IPM varied among the bulls. Meanwhile, the seminal plasma proteins contained bands with different MWs. No difference in the expression of protein bands between bulls. Linearity analysis showed that sperm motility (r= 0.281), viability (r= 0.189), abnormalities (r= 0.141), and IPM (r= 0.173) were positively correlated with the protein bands at each MW (p<0.05). The results conclude there was a positive correlation between the MW of the protein marker and the same protein expression levels in Bali bulls. Therefore, the band intensity of Bali cattle seminal plasma proteins can be used as a biomarker for selecting superior Bali bulls.

Molecular, behavioural and morphological comparisons of sperm adaptations in a fish with alternative reproductive tactics.

In species with alternative reproductive tactics, there is much empirical support that parasitically spawning males have larger testes and greater sperm numbers as an evolved response to a higher degree of sperm competition, but support for higher sperm performance (motility, longevity and speed) by such males is inconsistent. We used the sand goby (Pomatoschistus minutus) to test whether sperm performance differed between breeding-coloured males (small testes, large mucus-filled sperm-duct glands; build nests lined with sperm-containing mucus, provide care) and parasitic sneaker-morph males (no breeding colouration, large testes, rudimentary sperm-duct glands; no nest, no care). We compared motility (per cent motile sperm), velocity, longevity of sperm, gene expression of testes and sperm morphometrics between the two morphs. We also tested if sperm-duct gland contents affected sperm performance. We found a clear difference in gene expression of testes between the male morphs with 109 transcripts differentially expressed between the morphs. Notably, several mucin genes were upregulated in breeding-coloured males and two ATP-related genes were upregulated in sneaker-morph males. There was a partial evidence of higher sperm velocity in sneaker-morph males, but no difference in sperm motility. Presence of sperm-duct gland contents significantly increased sperm velocity, and nonsignificantly tended to increase sperm motility, but equally so for the two morphs. The sand goby has remarkably long-lived sperm, with only small or no decline in motility and velocity over time (5 min vs. 22 h), but again, this was equally true for both morphs. Sperm length (head, flagella, total and flagella-to-head ratio) did not differ between morphs and did not correlate with sperm velocity for either morph. Thus, other than a clear difference in testes gene expression, we found only modest differences between the two male morphs, confirming previous findings that increased sperm performance as an adaptation to sperm competition is not a primary target of evolution.

P-072 Collecting semen samples at home for fertility assessment has a positive effect on sperm quality

Study questionDoes site of semen collection (home/clinic) affect semen parameters?Summary answerSemen collection is superior when performed at home as compared with at a clinic in terms of sample volume, sperm concentration, and total sperm count.What is known alreadyIn fertility clinics the standard approach to semen collection for diagnostic analysis and use in ART involves a private room close to the andrological/embryological laboratory. In fact, WHO recommends that semen samples should be collected at clinic to avoid fluctuations in temperature and to control the time between collection and processing. On the other hand, today - due to the COVID-19 pandemic - semen collection at home is widely advised to reduce the time of stay of patients at clinic. There are still no firm conclusions whether collecting semen at home has any influence on sperm quality and reproductive competence.Study design, size, durationThis retrospective longitudinal cohort study performed at a tertiary level public fertility center included 8634 semen samples from 5880 men undergoing fertility assessment from 2015 to 2021: 5530 samples were collected at clinic from 3773 men, and 3104 at home from 2107 men. For a subgroup analysis comparing clinic to home collection within the same patient, we included 1260 samples from 428 men. Possible effect of seasonal variation on semen parameters was also investigated.Participants/materials, setting, methodsThe impact of sample collection site on endpoints (semen volume, concentration, motility) was evaluated using a generalized linear model for repeated measures in which the following covariates were included: age, BMI, days of sexual abstinence, smoke habit. Paired comparisons between home- and clinic-collected samples within individuals were made by paired t-test or Wilcoxon signed rank test, as appropriate. The effect of seasonality on sperm parameters was investigated plotting the time series of differences between samples.Main results and the role of chanceSamples collected at home had significantly higher semen volume (p = 0.016), sperm concentration (p < 0.0001), and total sperm count (p < 0.0001) respect to samples collected at clinic. The abstinence period, known to potentially influence these parameters, was similar for all patients. There was no difference in sperm motility. Paired comparisons of semen characteristics in the 428 patients with home-collected (n = 583) and clinic-collected (n = 677) samples confirmed the aforementioned results. The semen sample collections were distributed among all months over 5 years: regarding seasonal variations in semen quality, we did not find any evidence of variations of sample parameters by month.Limitations, reasons for cautionSome weaknesses should be considered: i) only men undergoing fertility check-up were assessed and it remains to be validated whether our conclusions are applicable to men attempting pregnancy by ART; ii) we did not measure type and degree of psychological stress that may have affected the men in our study.Wider implications of the findingsOur observation of decrease in semen quality of clinic-collected samples may be the result of an acute psychological stress experienced by patients, as suggested by previous reports. Thus, when possible, semen collection at home should be encourage at least for men who undergo fertility assessment.Trial registration numbernot applicable

Polymorphisms in the HSF2, LRRC6, MEIG1 and PTIP genes correlate with sperm motility in idiopathic infertility.

The aim of this study was to investigate the association of 24 functionally important single nucleotide polymorphisms (SNPs) with male infertility. In this cross-sectional study, we genotyped 24 functionally important single nucleotide polymorphisms in 24 infertility candidate genes in 500 oligo-/astheno-/oligoastheno-/normo-zoospermic infertile men with idiopathic infertility. Sequenom iPlex gold assay was used for genotyping. Sperm count and motility were compared between prevalent genotypes at each test locus. We did not observe any significant difference in the average sperm count between the alternate genotypes for the loci in the KLK3, LRRC6, MEIG1, HSF2, ESR2 and PTIP genes. However, we observed a significant difference in sperm motility between the alternate genotypes for the loci in the LRRC6, MEIG1, HSF2 and PTIP genes. Polymorphisms in the LRRC6 (rs200321595), MEIG1 (rs150031795), HSF2 (rs143986686) and PTIP (rs61752013) genes show association with sperm motility.

Semen characteristics and second successful artificial insemination of Asian elephant (Elephas maximus) in Thailand.

Background and Aim:As the number of wild Asian elephants (Elephas maximus) continues to decline, maintaining healthy populations under human care is vital. Male fertility assessment is essential for understanding the reproductive status, which can help to uncover underlying problems and improve the rate of pregnancy success. The objectives of this study in Asian elephants were as follows: (1) To investigate the semen characteristics; (2) to compare the relative seminal vesicle size and semen characteristics; (3) to compare the semen characteristics between good-motile (>60% progressive motility) and poor-motile (<60% progressive motility) ejaculates; and (4) to investigate the pregnancy success rate after artificial insemination (AI) with combined chilled and frozen semen.Materials and Methods:In total, 153 ejaculates were collected by manual rectal stimulation from 25 bulls. The volume, pH, sperm concentration, progressive motility, viability, morphology, and membrane integrity were investigated in each ejaculate. Assessment of accessory sex glands was conducted using transrectal ultrasonography to compare the relative seminal vesicle size and semen characteristics, and the bulls were divided into two groups according to the size of the ampulla (<7 or ≥7 cm2). For the comparison of good and poor-motile ejaculates and semen characteristics, the samples were divided into two groups: Good-motile (>60% progressive motility) and poor-motile (<60% progressive motility) ejaculates. Semen ejaculates for AI were collected from three bulls. The estrous cycles of four females were monitored using an enzyme immunoassay. Seven AI attempts were conducted using frozen and/or chilled semen by endoscopic visualization. AI was repeated 1 day before the luteinizing hormone surge, on the day of the surge, and 1 day after the surge. Pregnancy was confirmed by monitoring the serum progesterone profile and the abdomen and mammary glands changes.Results:From 153 ejaculates, the mean±standard error values of progressive motility, semen volume, sperm concentration, pH, and viability were 40.18%±2.28%, 40.94±3.86 mL, 1,205.58±62.26×106 sperm/mL, 7.50±0.10, and 56.17%±1.96%, respectively. Comparing ampulla size and semen characteristics revealed that the bulls with ampullae of ≥7 cm2 yielded significantly larger volume ejaculates. However, there were no significant differences in sperm motility and concentration. The comparison of semen characteristics between good- and poor-motile ejaculates revealed that the former had significantly higher pH, viability, normal acrosomes, intact membranes, and normal head and tail morphology but often had a significantly lower volume and sperm concentration. From seven AI attempts in four females, one female had a confirmed pregnancy (14.3% pregnancy rate), and delivered a healthy live female baby weighing 128 kg at 21 months and 12 days of gestation. The baby is now 3 years old and in a healthy condition, with normally developing growth and behavior.Conclusion:The semen characteristics of Asian elephants can be used as the baseline reference for further applications. The ampullae size indicates semen quantity but not quality. Our success in producing an elephant calf from AI using frozen and chilled semen demonstrated that AI can be used as an alternative approach for the breeding management of Asian elephants. However, the semen of Asian elephants is of poor quality, especially in terms of membrane integrity; thus, the improvement in semen quality through intensive and careful management of elephant health and fertility remains a challenge for the future. Furthermore, a sperm bank should be established to develop sperm cryopreservation, which will be invaluable for improving the genetic diversity of the Asian elephant.

Interferon γ, IL-17, and IL-1β impair sperm motility and viability and induce sperm apoptosis.

Urogenital inflammation is a known cause of male infertility. Increased levels of inflammatory cytokines, leukocyte counts and oxidative stress are highly detrimental for sperm quality thus compromising male fertility. Although cytokines affect sperm by recruiting and activating leukocytes consequently inducing tissue inflammation and oxidative stress, scarce to absent data have been reported about the putative direct effects of inflammatory cytokines on spermatozoa. Herein, we analyzed whether IFNγ, IL-17A, IL-1β, and IL-8 can alter human sperm motility and viability per se. Fractions of viable and motile spermatozoa from normospermic healthy donors were in vitro incubated with recombinant human IFNγ, IL-17A, IL-1β or IL-8 and sperm ROS production, motility, viability and apoptosis were analyzed. Sperm exposed to different concentrations of IFNγ, IL-17A and IL-1β, or a combination of them, for either 1 or 3h showed significantly increased levels of mitochondrial ROS production and reduced motility and viability with respect to sperm incubated with vehicle. Moreover, the exposure to IFNγ, IL-17A and IL-1β resulted in significantly higher levels of early and/or late apoptotic and/or necrotic spermatozoa. Interestingly, no significant differences in sperm motility, viability and apoptosis were observed in sperm incubated with the concentrations of IL-8 analyzed, for either 1 or 3h, with respect to sperm incubated with vehicle. In conclusion, our results indicate that IFNγ, IL-17A and IL-1β per se impair sperm motility and decreases viability by triggering increased mitochondrial ROS production and inducing sperm apoptosis. Our results suggest that screening inflammatory cytokines in semen would be an additional helpful tool for the diagnostic workup of male infertility.

Ultra-low temperature cryopreservation and −80 °C storage of sperm from normal-male and pseudo-male Siniperca chuatsi

The purpose of this study was to evaluate the effects of ultra-low temperature cryopreservation and − 80 °C storage of sperm from normal-male and pseudo-male mandarin fish (Siniperca chuatsi). In this study, sperm motility, plasma membrane integrity, mitochondrial membrane integrity, DNA integrity, fertilization, hatching rate and offspring growth were evaluated. After thawing, there was no significant difference in sperm motility and motility parameters of stripped sperm and testicular sperm of normal-males and pseudo-males. The motility of frozen sperm thawed in a 37 °C water bath was the highest, and there was no significant difference in the motility of frozen sperm of different tubes (n = 30) after thawing. The motility of frozen sperm of normal-males and pseudo-males were more than 60%, and the DNA fragmentation were less than 10%. The integrity of mitochondria and plasma membrane were higher than 84% and 75% respectively. There were no significant differences in plasma membrane, mitochondrial membrane integrity and DNA fragmentation between fresh sperm of normal-males and pseudo-males, nor between frozen sperm. The fertilization and hatching rate of normal-male frozen sperm were 65% and 85%, and values of pseudo-male frozen sperm were 57% and 83%, respectively. There was no significant difference in the body weight and length of offspring of frozen sperm between the normal-males and pseudo-males. The sperm motility, motility parameters and mitochondrial membrane integrity of the frozen sperm of normal-males and pseudo-males did not significantly decrease after −80 °C storage for 1, 3 and 5 days. The fertilization and hatching rate of normal-male and pseudo-male frozen sperm were still higher than 50% and 75%. Frozen sperm can successfully reproduce offspring after 5 days of −80 °C storage. The results indicated that testicular sperm of normal-male and pseudo-male S. chuatsi were successfully cryopreserved by ultra-low temperature cryopreservation, and the research of −80 °C storage showed that frozen sperm can be successfully stored for a short time at −80 °C which can provide a possibility for the dry ice transportation of frozen sperm.

Characteristics of diluted-stored and post-thawed semen of Hutsul stallions.

The use of frozen semen lowers the risk of disease transmission, eliminates geographical limitations and supports the implementation of genetic resource protection programs. However, due to the very rare use of frozen semen from Hutsul stallions, their genetic material is not secured in sperm banks, and very little information is available about their semen, including its suitability for cryopreservation, and sperm survival rates after thawing. The aim of this study was to analyse basic parameters such as sperm motility, vitality and morphology in diluted-stored and post-thawed Hutsul semen, using a CASA system. There were no differences in sperm motility (P = 0.3372) or morphology between the groups, although the progressive motility was higher in thawed semen (P = 0.0151), while the sperm vitality was higher in diluted-stored semen (P = 0.00517). This study demonstrates that semen from Hutsul horses is suitable for cryopreservation, thus supporting the creation of a sperm bank as a genetic reserve for representatives of this breed.