Sperm Morphological Defects Research Articles

Substitution of Egg Yolk with Arabic Gum in Extenders Used for Ram Semen Cold Storage or Cryopreservation

The current research was designed to compare the effects of substitution of egg yolk (EY) in ram extender with Arabic gum (AG) in different concentration in cooled or frozen semen collected by artificial vagina (AV) or electro-ejaculator (EE) in Noemi rams. Semen was collected 2 times per week for 6 weeks from 6 rams by EE method and after a rest period of 2 weeks, semen samples were collected by AV method. Tris extender either contained 15% EY (Control) or 3%, 5% or 7% AG (AG3, AG5 and AG7, respectively) were used as semen extenders. Spermatozoa were evaluated after cold storage (4 °C) or freezing/thawing in liquid nitrogen for sperm motility parameters: total motile spermatozoa (% TMS), rapid progressively motile sperm (% PRS), curvilinear velocity (VCL) μm/s, rectilinear velocity (VSL) μm/s, the average path velocity (VAP) μm/s, straightness index (% STR), linearity coefficient (% LIN), morphological defects and acrosome membrane integrity. Results showed that AV-collected sperm and diluted with AG or EY were more efficient in motility than EE. The values of VCL, VSL and VAP were higher in AG3 and AG5 and EY extenders for semen collected by AV compared to EE. After cold storage, AG3 gave better (P<0.01) results of LIN and STR either in spermatozoa collected by AV or EE. Significant differences in morphology defects were noted in semen collected by EE regardless of extenders used, however, AG supplementation enhance morphological defects in spermatozoa collected by AV (P<0.05). Post-freezing, Tris extender containing EY and AG5 in AV-collected spermatozoa had higher TMS, PRS, VCL, VSL and VAP compared to others. Sperm morphological defects were higher in AG7 extender in EE and EY extender in AV group. Plasma membrane integrity was significantly (P<0.05) higher in AG3 and AG5 extenders in AV-group compared to control and other groups. The acrosome membrane integrity in AV-semen groups was greater (P<0.05) than that of EE-semen group. It can be concluded that, Arabic gum is a good candidate to replace egg yolk in ram semen extender when added at a concentration of 3 or 5%. Although method of artificial vagina semen collection gave better results than electro-ejaculator. However, Arabic gum could be added to ram semen extender to enhance the electro-ejaculator-collected semen characteristics when added at a concentration of 5%

Oxidative Stress is Associated with Reduced Sperm Motility in Normal Semen.

Infertility is among the most serious medical problems worldwide. Male factors contribute to 40%–50% of all infertility cases, and approximately 7% of men worldwide are affected by infertility. Spermatozoa are extremely vulnerable to oxidative insult. Oxidative stress results in axonemal damage and increased midpiece sperm morphological defects, which lead to reduced sperm motility. The aim of the study is to evaluate the association between sperm motility and the levels of selected antioxidants, cytokines, and markers of oxidative damage in the seminal plasma.The study group included 107 healthy males, who were split into two subgroups based on the percentage of motile spermatozoa after 1 hr: low motility (LM, n = 51) and high motility (HM, n = 56).The glucose-6-phosphate dehydrogenase (G6PD) activity was 52% lower in the LM group compared to that in the HM group. The level of malondialdehyde (MDA) was 12% higher in the LM group compared to that in the HM group. Similarly, the median values of interleukin (IL)-1β, IL-10, IL-12, and tumor necrosis factor alpha (TNF-α) were higher in the LM group than those in the HM group. Results of the present study revealed that the percentage of motile spermatozoa after 1 hr correlated positively with the levels of IL-1β, IL-10, IL-12, and TNFα.The lower motility of spermatozoa in healthy men is associated with a decreased activity of G6PD and increased levels of cytokines, which may be related to increased oxidative stress in seminal plasma that manifests as an increased level of MDA.

Semen collection, sperm characteristics and ultrasonographic features of reproductive tissues in crab-eating fox (Cerdocyon thous)

Our objective was to condition adult male crab-eating fox (Cerdocyon thous) to allow semen collection by digital manipulation of the penis, assess sperm characteristics, and use ultrasonography to assess and measure testes and prostate and determine echotexture, as well as hemodynamics of testicular and spermatic cord arteries. Five captive C. thous (2–5 y, ∼7 kg) were used. After conditioning for digital sperm collection, 13 ejaculates were obtained from two males. Ejaculate characteristics (mean ± SD) were volume (393.2 ± 178.0 μL), concentration (463.7 ± 594.4 × 106 sperm/mL), total sperm number (217.4 ± 304.0 × 106 sperm), total motility (68.0 ± 6.1%), progressive motility (48.8 ± 5.1%) and rapid sperm (57.2 ± 6.8%). The most prevalent sperm morphological defects were abnormal acrosome, small abnormal head, and diadem defect. Under chemical restraint, testes and prostate were evaluated with B-mode and spectral Doppler ultrasonography. Based on ultrasonography, testicular volumes were (2.2 ± 0.7 and 2.1 ± 0.7 cm3 for left and right testes, respectively), whereas prostate dimensions were 2.3 ± 0.4 cm long and 1.5 ± 0.3 cm thick. Lastly, using triplex Doppler, peak systolic velocity (PSV) was higher in testicular versus intratesticular arteries. In conclusion, adult male C. thous were conditioned to allow sperm collection by digital manipulation without urine contamination, enabling evaluation of sperm characteristics. Furthermore, B-mode ultrasound facilitated evaluation of testes and prostate morphometry and echotexture, whereas Doppler was used to characterize testicular blood flow. This was proof of concept for semen collection and baseline data for future male reproductive soundness examinations and development of assisted reproductive techniques in this species.

Auricularia polytricha restores altered reproductive parameters in streptozotocin-induced diabetic Wistar rat

BackgroundLocal Nigerian men use Auricularia polytricha (wood ear mushroom) growing in farm lands and dead woods to manage sexual dysfunction and this practice has been going on for ages without supporting evidence from scientific experiments. This study was to investigate the ameliorative effect of ethanolic extract of Auricularia polytricha on male sex hormones (FSH, LH, and testosterone), sperm parameters (sperm count, sperm motility, and morphology), and histopathological examination of testis using Streptozotocin (STZ)-induced diabetic rats as a model.MethodsThe experiment included five groups of adult male Wistar rat, group A (normal control, treated with normal saline), group B (diabetic control treated with 65 mg/kg.bw of STZ), groups C, D, and E (treated with 250 mg/kg.bw, 500 mg/kg.bw, 1000 mg/kg.bw A. polytricha after inducing diabetics). At termination, homogenate was prepared for semen analysis using improved Neubauer’s hemocytometer, serum samples were used for hormonal assay with ELISA (enzyme-linked immune sorbent assay) kit and light microscope was used for histopathological examination.ResultsResults revealed significant (p < 0.05) reduction in sperm count and motility in group B when compared to normal control, values in group C were not statistically significant (p > 0.05) when compared to diabetic control. However, groups D and E recorded sperm count and sperm motility that were significantly (p < 0.05) increased when compared to the diabetic control (group B). Morphological defects in diabetic control rats were significantly (p < 0.05) higher when compared to normal control but there was no statistical significance in group C when compared to the diabetic control, meanwhile values of sperm morphological defects in groups D and E were significantly (p < 0.05) lower when compared to the diabetic control group. All gonadotropic hormones considered in this research were remarkably lower at p < 0.05 when compared to normal control. However, these values were significantly higher in groups D and E when compared to diabetic control. Structural alterations in cytoarchitecture of testes as a result of sustained hyperglycemia were reversed in a dose dependent manner following A. polytricha administration.ConclusionAlterations in sperm parameters, gonadotropic hormones, and histology of testes caused by diabetes were regulated and reversed following administration of graded doses of A. polytricha. This reversal may be attributed to essential phytochemical and therapeutic constituents in A. polytricha which can be found useful in prevention and treatment of diabetes induced testicular dysfunction.

Impact of sperm characteristics on time-lapse embryo morphokinetic parameters and clinical outcome of conventional in vitro fertilization.

Sperm abnormalities may negatively affect embryo development. To determine the influence of sperm abnormalities (morphology, motility, DNA fragmentation) on embryo morphokinetic variables and clinical outcome of conventional IVF. Participants were 86 couples undergoing in vitro fertilization (IVF). Sperm morphology was evaluated according to the strict criteria proposed by Kruger/Tygerberg. CASA system was applied for sperm motility assessment. Sperm DNA fragmentation was assessed by the chromatin structure assay (SCSA). Morphokinetic parameters were determined in 223 embryos obtained from conventional IVF only and cultured in a single-step medium using time-lapse imaging technology. Time-lapse variables from the initial embryo development, such as time of pronuclei fading (tPNf) and time for two cells (t2), were those more strongly related with abnormalities of sperm motility, morphology, and DNA fragmentation. Sperm morphological abnormalities rather than sperm motility were more closely associated with embryo morphokinetics. Sperm head defects were mainly correlated with the last stages of embryonic development (t9 to tHB), sperm midpiece defects with intermediate cleaving embryos (t5-t9), and sperm tail defects with the initial stages of embryonic development (tPNa-t4). Excess residual cytoplasm was positively correlated with all embryo morphokinetic parameters except t2 and tM. Absence of acrosomes, pinheads, coiled tails, and multiple sperm morphological defects correlated negatively with time-lapse embryo morphokinetic variables. A large number of sperm-related variables, including frequency of specific morphological defects, morphological indexes, DNA fragmentation and motility, and time-lapse embryo variables, such as time intervals based mainly of 15 time points were recorded. There were strong associations between specific sperm defects of the head, midpiece, and tail with certain stages of embryonic development from observation of pronuclei to the hatched blastocyst. Coiled tail, cumulative head defects, and multiple abnormalities index (MAI) were associated both with embryo morphokinetics and the implantation success.

Knockout of mouse receptor accessory protein 6 leads to sperm function and morphology defects†.

Receptor accessory protein 6 (REEP6) is a member of the REEP/Ypt-interacting protein family that we recently identified as essential for normal endoplasmic reticulum homeostasis and protein trafficking in the retina of mice and humans. Interestingly, in addition to the loss of REEP6 in our knockout (KO) mouse model recapitulating the retinal degeneration of humans with REEP6 mutations causing retinitis pigmentosa (RP), we also found that male mice are sterile. Herein, we characterize the infertility caused by loss of Reep6. Expression of both Reep6 mRNA transcripts is present in the testis; however, isoform 1 becomes overexpressed during spermiogenesis. In vitro fertilization assays reveal that Reep6 KO spermatozoa are able to bind the zona pellucida but are only able to fertilize oocytes lacking the zona pellucida. Although spermatogenesis appears normal in KO mice, cauda epididymal spermatozoa have severe motility defects and variable morphological abnormalities, including bent or absent tails. Immunofluorescent staining reveals that REEP6 expression first appears in stage IV tubules within step 15 spermatids, and REEP6 localizes to the connecting piece, midpiece, and annulus of mature spermatozoa. These data reveal an important role for REEP6 in sperm motility and morphology and is the first reported function for a REEP protein in reproductive processes. Additionally, this work identifies a new gene potentially responsible for human infertility and has implications for patients with RP harboring mutations in REEP6.

Plasma estradiol-17β, cortisol, and insulin concentrations and serum biochemical parameters surrounding puberty in Japanese Black beef bulls with normal and abnormal semen

The associations of semen abnormalities with circulating hormones (estrogens, glucocorticoid, insulin) and common biochemical parameters are unclear in beef bulls. We compared plasma concentrations of estradiol-17β, cortisol, and insulin and serum biochemical parameters surrounding puberty in Japanese Black beef bulls (n = 96) with normal post-thaw or abnormal semen (fresh and frozen). Blood samples were collected monthly from 4 to 24 months of age (n = 50) for the assays of plasma estradiol, cortisol, and insulin and every 3 months from 6 to 21 months of age (n = 92) for the serum biochemical analyses. Semen was collected weekly from 12 months until at least 18 months of age. Fresh semen was evaluated for semen volume, sperm progressive motility, concentrations, and morphological defects. The normal fresh semen was frozen by a standard method and examined for post-thaw sperm motility and fertility, which were evaluated for rates of transferable embryos. Bulls were classified as having either normal fresh semen or abnormal fresh semen (when at least one of the above test items was abnormal for 6 months). The normal fresh semen was categorized as having either normal post-thaw semen or low fertility post-thaw semen. The abnormal fresh semen was categorized as having sperm morphological defects, low motility, or morphological defects plus low motility. Plasma cortisol concentrations in the abnormal fresh semen group were higher than those of the normal fresh semen group (p < 0.0001). Plasma estradiol-17β and insulin concentrations in the low-fertility post-thaw semen group were lower than those of the normal post-thaw semen group (p < 0.0001). Serum aspartate aminotransferase and magnesium concentrations were greater for the abnormal fresh semen group vs. the normal fresh semen group (p < 0.005). These results suggest that fresh semen abnormality in pubertal beef bulls might be associated with increased circulating aspartate aminotransferase, magnesium and cortisol. Low-fertility post-thaw semen could have been involved with the lower peripheral estradiol and insulin levels in beef bulls.

Can maintenance intake reduce production costs without compromising the reproductive performance in bucks?

Excessive intake of nutrients can result in decreases in the reproductive efficiency of goat bucks and increase production costs in goat e nterprises. In this study, the aim was to compare the reproductive features of young bucks fed diets for maintenance of growth or were fed ad libitum. Ten bucks were used (n = 5/group), with a completely randomized design. Blood and semen were collected from each animal every 2 weeks for nine collections. Semen was frozen and later analyzed using flow cytometry to compare reactive oxygen species, plasma membrane integrity, acrosome integrity, and membrane lipid peroxidation. Body condition score, sexual behavior, testicular biometry and histomorphometry of the testis, as well as oxidative enzymes and carbonylated proteins, were also evaluated. Animals fed ad libitum had a greater body condition score (3.32 + 0.08 × 2.88 + 0.05), sperm volume (0.68 + 0.04 × 0.52 + 0.03 mL), and testis circumference (25.2 + 0.21 × 24.5 + 0.24 cm) and volume (121 + 5.83 × 107 + 4.38 cm³, left testicle; 116 + 5.26 × 100 + 3.75 cm³, right testicle). Ejaculates of the bucks fed diets to maintain growth rate had a larger number of minor and total sperm morphology defects. There were no differences in testosterone concentrations and sexual behavior. In conclusion, there can be feeding for growth rate maintenance instead of ad libitum feeding, for the reduction in production costs of young bucks without compromising semen quality and reproductive functions.

Genotoxicity and sperm defects induced by 5-FU in male mice and the possible protective role of Pentas lanceolata-iridoids

5-Fluorouracil (5-FU) is a widely used antineoplastic drug. In this work, a comprehensive study was performed to detect the extent of chromosomal damage and morphological sperm defects induced by 5-FU in male mice and the possible protective role of the iridoids-rich fraction of Pentas lanceolata leaves (IFPL).Six main groups were examined in micronucleus and chromosomal assays: I- control negative, II- control positive (i.p. treated with single dose of 75 mg/kg 5-FU), III- control plant (orally administrated IFPL, 300 mg/kg, 5 consecutive days), and IV-VI- treated with IFPL (100, 200 and 300 mg/kg, 5 consecutive days) plus 5-FU (i.p. treated at the last day). Samples were taken 24 h post treatment. The study of morphological sperm anomalies, single and repeated treatments were examined and samples were taken after 35 days from the 1st treatment.In bone marrow, 5-FU induced a significant increase in the micro-nucleated polychromatic erythrocytes, chromosome anomalies (CAs) and also cytotoxic effects. A significant percentage of CAs was recorded in spermatocytes after 5-FU treatment reached 22.80 ± 1.32 vs 4.20 ± 0.37 for control (mainly X–Y univalent, 90%). IFPL was recorded to be non-mutagenic in all tests examined. In addition, it alleviated the previous defects in a dose-dependent manner. A significant and dramatic increase in the percentage of morphological sperm defects was recorded after single and repeated treatments with 5-FU reached 13.24 ± 0.24, 30.42 ± 0.32 respectively vs 2.56 ± 0.14 for control. Amorphous head-sperm and sperm with coiled tail were the most pronounced types of abnormalities. Significant protection was detected with the highest tested dose of IFPL.In conclusion: 5-FU demonstrated to be a genotoxic agent. Its genotoxicity in germ cells is serious and may lead to reproductive toxicity, infertility or heritable defects. The results also demonstrated the biosafety of IFPL and its possible protective role in combined treatment with 5-FU.

Dependence of sperm structural and functional integrity on testicular calcineurin isoform PPP3R2 expression.

After leaving the testis, mammalian sperm undergo a sequential maturation process in the epididymis followed by capacitation during their movement through the female reproductive tract. These phenotypic changes are associated with modification of protein phosphorylation and membrane remodeling, which is requisite for sperm to acquire forward motility and induce fertilization. However, the molecular mechanisms underlying sperm maturation and capacitation are still not fully understood. Herein, we show that PPP3R2, a testis-specific regulatory subunit of protein phosphatase 3 (an isoform of calcineurin in the testis), is essential for sperm maturation and capacitation. Knockout of Ppp3r2 in mice leads to male sterility due to sperm motility impairment and morphological defects. One very noteworthy change includes increases in sperm membrane stiffness. Moreover, PPP3R2 regulates sperm maturation and capacitation via (i) modulation of membrane diffusion barrier function at the annulus and (ii) facilitation of cholesterol efflux during sperm capacitation. Taken together, PPP3R2 plays a critical role in modulating cholesterol efflux and mediating the dynamic control of membrane remodeling during sperm maturation and capacitation.

26 Baobab oil supplemented extender preserves post-thaw bull sperm quality parameters

The processes of semen cryopreservation and thawing affect sperm membrane integrity and motility and increases morphological defects as well as DNA damage. The most influential cause of this is oxidative stress. When endogenous antioxidant capacity of seminal plasma is reduced during the freeze-thawing process, plant extracts exhibiting strong antioxidant activity can be used as supplements for compensation. Baobab oil has gained interest because it is rich in powerful antioxidants, which could protect sperm cells from oxidative damage during cryopreservation. Our study aimed to assess the effects of baobab oil on post-thaw sperm quality parameters in an egg-yolk-based extender. Thirty semen ejaculates were collected from 15 Nguni bulls using an electro ejaculator. Semen samples were randomly allocated to control (no baobab oil), 20μL (1%), 50μL (2.5%), and 100μL (5%) baobab oil per millilitre extender. Following dilution, semen samples were loaded into 0.25-mL semen straws, equilibrated for 4h at 5°C, and transferred into a controlled rate programmable freezer. The frozen semen straws were stored in a liquid nitrogen tank (−196°C) until thawing. Semen straws were thawed (37°C/60 minutes) after 1 week of cryopreservation and analysed for (1) sperm motility using a computer-aided sperm analyser, (2) morphological defects and viability using eosin-nigrosin stain, (3) membrane integrity by hypo-osmotic swelling test, and (4) DNA integrity by terminal deoxynucleotidyl transferase dUTP nick end labelling assay. Data was analysed using analysis of variance. Treatment means were compared in relation to the control group by Dunnett's test. We found that supplementing semen extender with baobab oil at 1% significantly (P&amp;lt;0.05) preserved sperm DNA integrity (88.3±3.7) and membrane integrity (74.0±4.2) when compared with the control group (71.7±3.7 and 55.8±4.4, respectively). Baobab oil supplementation either at 1% (5.9±0.5), 2.5% (7.2±0.5), or 5% (6.0±0.5) significantly reduced sperm morphological defects compared with control (9.5±0.5). Total motility (1% (72.7%), 2.5% (72.7%), 5% (71.9%), control (59.3%)) and viability (1% (79.1%), 2.5% (79.8%), 5% (77.8%), control (67.6%)) were also improved by supplementation; however, the difference was not significant. In conclusion, it was demonstrated that supplementing bull semen extender with 1% baobab oil protects sperm from morphological defects, maintains membrane integrity, as well as preserves sperm DNA. All the baobab oil supplementation levels preserved post-thaw bull-sperm quality parameters.

Mutations in TTC29, Encoding an Evolutionarily Conserved Axonemal Protein, Result in Asthenozoospermia and Male Infertility

In humans, structural or functional defects of the sperm flagellum induce asthenozoospermia, which accounts for the main sperm defect encountered in infertile men. Herein we focused on morphological abnormalities of the sperm flagellum (MMAF), a phenotype also termed "short tails," which constitutes one of the most severe sperm morphological defects resulting in asthenozoospermia. In previous work based on whole-exome sequencing of a cohort of 167 MMAF-affected individuals, we identified bi-allelic loss-of-function mutations in more than 30% of the tested subjects. In this study, we further analyzed this cohort and identified five individuals with homozygous truncating variants in TTC29, a gene preferentially and highly expressed in the testis, and encoding a tetratricopeptide repeat-containing protein related to the intraflagellar transport (IFT). One individual carried a frameshift variant, another one carried a homozygous stop-gain variant, and three carried the same splicing variant affecting a consensus donor site. The deleterious effect of this last variant was confirmed on the corresponding transcript and protein product. In addition, we produced and analyzed TTC29 loss-of-function models in the flagellated protist T.brucei and in M.musculus. Both models confirmed the importance of TTC29 for flagellar beating. We showed that in T.brucei the TPR structural motifs, highly conserved between the studied orthologs, are critical for TTC29 axonemal localization and flagellar beating. Overall our work demonstrates that TTC29 is a conserved axonemal protein required for flagellar structure and beating and that TTC29 mutations are a cause of male sterility due to MMAF.

Developmental effects on hypothalamic, hypophyseal, testicular and steroidogenic patterns of sertraline-exposed male rats by accumulated doses from juvenile to puberty

Since the enduring exposure to selective serotonin reuptake inhibitors medications from juvenile period to puberty poses a growing concern, the aim is an attempt to evaluate the reproductive aspects of sertraline-treated postnatal male rats. Total 80 male rats were orally given 1.2 mg/kg bw/day from postnatal day 28 to puberty (balano-preputial separation). Necropsy takes place at 56th, 84th, 126th postnatal day (SGII, SGIII, and SGIV, respectively), along with the control group (SGI). Final body weight, weight gain, and weights of liver, kidneys, testes and epididymis were significantly decreased in the SGIII and SGIV groups compared to the controls. Levels of LH, FSH, and testosterone and 17β-HSD concentrations were significantly decreased in all groups. Male rats in SGIV group displayed a significant decline in sperm counts, motility and viability and increase in sperm morphological defects compared to control. The cumulative dose of 1.2 mg/kg of sertraline at the 126th postnatal days produced a significant depression in male virility (mating and fertility indices) compared to the control group. In addition, the cumulative treatment significantly increased the number of fetal resorptions in outcomes of female rats copulated by males in the SGIV group and decrease in both the number of implant sites and the viable fetuses. It is concluded that the sertraline-mediated reproductive deficits could entirely dependent on the robust spreading of the serotonergic receptors in the Leydig, Sertoli and germ cells, testis, epididymis, and vas deferens and simultaneously on the developmental-mediated timing of reproductive processes during the postnatal period to puberty.

Novel association between apoptotic sperm biomarkers with seminal biochemical parameters and acetylcholinesterase activity in patients with teratozoospermia.

We aimed to determine whether the dysfunction of physiological apoptosis and specific seminal biochemical parameters could be associated with male infertility and sperm morphological defects. Ejaculated sperm samples from sixty patients with isolated teratozoospermia and thirty fertile donors were analyzed. The proportion of both viable and dead spermatozoa expressing activated caspases was detected by fluorescence microscopy through the use of different specific carboxyfluorescein-labeled caspase inhibitors FLICA. The different stages of apoptosis in human were qualitatively and quantitatively determined by using the AO/EB fluorescent staining method. The levels of the seminal biochemical parameters (acetylcholinesterase (AChE), lactate dehydrogenase (LDH), creatine phosphokinase (CK), iron (Fe), calcium (Ca), and phosphorus (P)) were evaluated spectrophotometrically. Patients with teratozoospermia showed significantly higher proportions of dead and live spermatozoa with activated caspases and spermatozoa in the late stage of apoptosis when compared to controls. Among the different studied biochemical seminal parameters, the rates of acetylcholinesterase activity, creatine phosphokinase, iron, and calcium were significantly increased in the patient group. However, the rate of phosphorus was significantly decreased. Interestingly, significant relationships were found between the studied biochemical and apoptotic biomarkers and the rates of atypical sperm forms with the incidences of head, mid-piece, and tail abnormalities. Furthermore, positive correlations were found between P, AChE, Fe, CK, and LDH with apoptotic markers. These results emphasize the impact of apoptosis in the pathophysiology of teratozoospermia and suggest that seminal biochemical disturbance may arise such damage.

Effect of breed, age, season and region on sperm morphology in 11,387 bulls submitted to breeding soundness evaluation in Australia

This study reports the distribution of sperm morphology defects by breed, age, season and region of 11,387 bulls in 500 herds in Australia and near Pacific Islands during annual BBSE. Bull location was divided into 4 broad climatic regions based upon temperature, vegetation and climatic risk. Taking into account the impact of age, season, region, and breed there were differences between breeds in both percent morphologically normal sperm and in some individual categories of sperm abnormality (P < 0.001). Independent of breed, season and region, proximal droplets were significantly increased in bulls less than 20 months of age. This is the first study to comprehensively collect data from this wide geographical area and compare sperm morphology profiles among the Bos indicus and Bos taurus breeds. The findings of this study will act as a guide for veterinary practitioners and cattle breeders in the proportion of bulls that can be expected to pass the PNS test, by breed, age and region, based on a robust data set.

Investigation of deep learning based detection of sperm morphological defects

Investigation of deep learning based detection of sperm morphological defects

Certain morphological characteristics of the spermatozoon used for intracytoplasmic sperm injection (ICSI) are associated with a higher likelihood of developmental arrest and embryo degeneration

Around 25% of in-vitro cultured human embryos arrest their development and degenerate during the first 5 days. The functional and morphological characteristics of the spermatozoa, selected for the intracytoplasmic sperm injection (ICSI) could account for the subsequent embryo development failure. The objective of the present study was to assess the relationship between the morphological characteristics of the spermatozoon used for ICSI and the increased risk of developmental arrest and embryo degeneration. Prospective observational study. A total of 83 spermatozoa from 68 men used in ICSI procedures were studied. Inclusion criteria were: 1) patient age < 39 years, 2) patient body mass index (BMI) < 25 kg/m2, 3) excellent quality oocytes. To find out which sperm morphological features could be responsible for the embryo degeneration the morphology of each individual spermatozoon used for ICSI was recorded. The morphology of all individual spermatozoa was evaluated using high magnification (×6100) immediately before the injection. Seven sperm morphological parameters and the presence of sixteen head, neck and tail abnormalities were evaluated. All 83 embryos were cultured in-vitro in Global one-step medium. The frequencies of occurrence of sperm morphological defects were compared between the resulted viable and degenerated embryos. Data was analyzed using the IBM SPSS software, v.21.0. Among 83 ICSI performed, 63 embryos (75.9%) had successful development and 20 embryos (24.1%) arrest their development before day 5. The incidence of pyriform head, thick midpiece and cytoplasmic residues among the spermatozoa that resulted in embryos with developmental arrest were significantly higher in comparison to spermatozoa that led to successfully developed embryos (20% vs. 13%, 40% vs. 21% and 20% vs. 6%, respectively, p<0.05, Chi-square test). In addition, the spermatozoa resulted in embryos with successful development had significantly shorter heads and tails in comparison with those that resulted in embryo development arrest (4.46 vs. 4.63 μm and 41.42 vs. 43.65 μm, p<0.05, Student t-test, respectively). When receiver operating curve (ROC) analysis was performed to find the optimum cut-off value to predict embryo development sperm head length had the best AUC (AUC = 0.66, CI 95%: 0.51–0.76). The optimum cut-off value for head length was 4.25 μm (sensitivity 89.5%, specificity 60.3%). The recorded spermatozoa tail length had lower AUC (0.63, CI 95%: 0.49–0.77) and the optimal cut-off value of 42.5 μm (sensitivity 63.2%, specificity 42.5%). Specific sperm morphological characteristics, such as head length and tail length and the presence of certain sperm abnormalities (pyriform head, thick midpiece and residual cytoplasmic droplets) are associated with an increased risk of developmental arrest and embryo degeneration. Selection of human spermatozoa with head length < 4.25 μm, tail length < 42.5 μm and without pyriform head, thick midpiece and cytoplasmic residues significantly increase the chance for successful embryo development.

Dependence of the frequency of sperm defects and dimensions on sperm motility in ejaculates of Polish Landrace boars

An attempt was made to determine the dependence of the frequency of sperm defects and dimensions on sperm motility in ejaculates of Polish Landrace boars. The study was conducted on 393 ejaculates collected from 33 Polish Landrace boars. Ejaculates were grouped according to the percentage of sperm with progressive motility, distinguishing ejaculates in which the percentage of motile sperm was 70% and 80%. In each ejaculate, the frequency of morphological changes in the sperm was determined and morphometric measurements of the sperm were made. Ejaculates with a higher proportion of sperm with progressive motility were found to contain more sperm. The ejaculate volume and sperm concentration in the ejaculate were not found to be directly associated with sperm motility. The frequency of primary defects was linked to sperm motility. Ejaculates with higher sperm motility contained fewer sperm with primary defects. The frequency of minor morphological changes, however, shows no significant dependence on sperm motility in the ejaculate. The primary morphological sperm defects most often found in ejaculates are a proximal droplet and the Dag defect. Both of these morphological forms are more common in ejaculates with lower sperm motility. The most common secondary sperm defects include sperm with a simple bent tail, sperm with a free normal head, and sperm with a distal droplet. These defects were not found to depend on sperm motility in the ejaculate. Sperm cells in ejaculates with greater sperm motility had slightly larger dimensions than sperm in ejaculates with lower sperm motility. Ejaculates with higher sperm motility are preferable for use in practice, not only because more insemination portions can be prepared from them, but also due to the lower frequency of primary defects.

Multigenerational exposure to uranium changes morphometric parameters and global DNA methylation in rat sperm

There is increasing evidence that environmental exposures early in fetal development influence phenotype and give rise to disease risk in the next generations. We previously found that lifelong exposure to uranium, an environmental contaminant, induced subtle testicular and hormonal defects; however, its impact on the reproductive system of multiple subsequent generations was unexplored. Herein, rats were exposed to a supra-environmental and non-nephrotoxic concentration of natural uranium (U, 40 mg·L−1 of drinking water) from postnatal life to adulthood (F0), during fetal life (F1), and only as the germ cells from the F1 generation (F2). General parameters (reproductive indices, epididymal weight) and sperm morphology were assessed in the three generations. In order to identify the epigenetic effects of U, we analyzed also the global DNA methylation profile and described for the first time the mRNA expression levels of markers involved in the (de)methylation system in rat epididymal spermatozoa. Our results showed that the F1 generation had a reduced pregnancy rate. Despite the sperm number being unmodified, sperm morphology was affected in the F0, F1 and F2 generations. Morphometric analysis for ten parameters was detailed for each generation. No common parameter was detected between the three generations, but the head and the middle-piece were always modified in the abnormal sperms. In the F1 U-exposed generation, the total number of abnormal sperm was significantly higher than in the F0 and F2 generations, suggesting that fetal exposure to uranium was more deleterious. This effect could be associated with the pregnancy rate to produce the F2 generation. Interestingly, global DNA methylation analysis showed also hypomethylation in the sperm DNA of the last F2 generation. In conclusion, our study demonstrates that uranium can induce morphological sperm defects and changes in the DNA methylation level after multigenerational exposure. The epigenetic transgenerational inheritance of U-induced reproductive defects should be assessed in further experiments.

Genetic relationships among sperm quality traits of Duroc boars collected during the summer season

The goals of the study were to estimate the heritability and repeatability of boar sperm quality traits during the summer season, estimate phenotypic and genetic correlations between sperm quality traits, and assess the effect of these traits on total number born (TNB). Semen samples were collected and individually processed from Duroc boars (n = 363) from May through October 2017. Single sire litter information was available for 121 of the boars. Heritability, repeatability, genetic correlations, and phenotypic correlations were estimated for and among the following traits: total percentage of motile cells (MOT), total percentage of progressively motile cells (PROG), percentage of cells with a distal droplet (DIST), percentage of cells with a proximal droplet (PROX), percentage of cells with a bent tail (BENT), percentage of cells with a distal midpiece reflex (DMR), mean sperm head elongation (ELON), and total sperm cells per ejaculate (TOTSP). Heritability estimates ranged from 0.08 to 0.24 and repeatability estimates ranged from 0.21 to 0.62. The phenotypic and genetic correlations between sperm motility traits and morphological defects indicated a negative relationship. Sperm morphological defects had positive phenotypic and genetic correlations with each other. Total sperm per ejaculate showed positive genetic correlations with the motility traits. Positive phenotypic and genetic correlations existed between ELON and the motility traits. Total sperm motility (P < 0.001) positively affected TNB while DIST (P < 0.001), PROX (P < 0.001), BENT (P = 0.05) and DMR (P = 0.002) negatively affected TNB. Results indicate genetic selection could enhance sperm quality during the summer season in boars.