Transmission electron microscopy reveals the presence of SARS-CoV-2 in human spermatozoa associated with an ETosis-like response.

BACKGROUND: Severe acute syndrome coronavirus 2 can invade a variety of tissues, including the testis. Even though this virus is scarcely found in human semen polymerase chain reaction tests, autopsy studies confirm the viral presence in all testicular cell types, including spermatozoa and spermatids. OBJECTIVE: To investigate whether the severe acute syndrome coronavirus 2 is present inside the spermatozoa of negative polymerase chain reaction-infected men up to 3 months after hospital discharge. MATERIALS AND METHODS: This cross-sectional study included 13 confirmed moderate-to-severe COVID-19 patients enrolled 30-90 days after the diagnosis. Semen samples were obtained and examined with real-time polymerase chain reaction for RNA detection and by transmission electron microscopy. RESULTS: In moderate-to-severe clinical scenarios, we identified the severe acute syndrome coronavirus 2 inside spermatozoa in nine of 13 patients up to 90 days after discharge from the hospital. Moreover, some DNA-based extracellular traps were reported in all studied specimens. DISCUSSION AND CONCLUSION: Although severe acute syndrome coronavirus 2 was not present in the infected men's semen, it was intracellularly present in the spermatozoa till 3 months after hospital discharge. The Electron microscopy (EM) findings also suggest that spermatozoa produce nuclear DNA-based extracellular traps, probably in a cell-free DNA-dependent manner, similar to those previously described in the systemic inflammatory response to COVID-19. In moderate-to-severe cases, the blood-testes barrier grants little defence against different pathogenic viruses, including the severe acute syndrome coronavirus 2. The virus could also use the epididymis as a post-testicular route to bind and fuse to the mature spermatozoon and possibly accomplish the reverse transcription of the single-stranded viral RNA into proviral DNA. These mechanisms can elicit extracellular cell-free DNA formation. The potential implications of our findings for assisted conception must be addressed, and the evolutionary history of DNA-based extracellular traps as preserved ammunition in animals' innate defence might improve our understanding of the severe acute syndrome coronavirus 2 pathophysiology in the testis and spermatozoa.

Serum lipid profile levels and semen quality: new insights and clinical perspectives for male infertility and men's health.

PURPOSE: Several clinical scenarios regulate the final ejaculated semen, which is pivotal to reproductive success. Sperm motility and plasma membrane fusogenic activity primarily rely on the peculiar sperm lipid composition, influenced by the patient's metabolism, genetics, nutritional, environmental status, and concomitant clinical entities such as varicocele. This study aimed to determine the relationship between serum lipid profile and testicular function (semen quality and testosterone levels). METHODS: This retrospective study uses medical charts of 278 infertile men who attended andrological care between 2000 and 2019. Seminal analysis data, lipid profile, and total serum testosterone were collected. A multiple linear regression analysis was performed to evaluate the influence of the lipid parameters on the seminal variables. Statistical analyses were carried out with p ≤ 0.05 considered statistically significant. RESULTS: Seminal creatine kinase activity (p = 0.024) is negatively related to HDL (p = 0.032) and triglycerides (p = 0.037), while total testosterone (p < 0.0001) and seminal volume (p = 0.046) appeared both to be negatively related to triglycerides (p = 0.030 and p = 0.033, respectively). CONCLUSION: Medical advice commonly advocated to prevent endothelial dysfunction and cardiovascular disease and improve HDL-cholesterol and triglyceride levels in dyslipidemic patients should also be given to infertile men. Physicians should give patients a thorough assessment, including the blood lipid profile, hormonal status, and routine seminal examinations. We propose a more comprehensive men´s health check-up for the infertile male population, not limited to a simple evaluation of basic sperm parameters.

Marijuana Is Associated With a Hormonal Imbalance Among Several Habits Related to Male Infertility: A Retrospective Study.

Marijuana is one of the most consumed drugs worldwide. There is increasing evidence of an association between marijuana and male infertility. This study intends to assess the repercussion of marijuana smoking and other habits (sedentary lifestyle, alcohol, and tobacco use) in the testicular function of infertile men seeking andrological evaluation. A retrospective study was performed using medical records data of men aged 18-59 years from 2009 to 2017. Complete semen analyses, sperm functional tests, SHBG, and hormonal levels, testosterone-to-estradiol ratio (T/E2), and testis volume were evaluated. Exclusion criteria included cryptorchidism, infertility caused by genetic or infectious diseases, and cancer. A multiple linear regression analysis was performed to investigate which habit could predict certain parameters using the software SPSS 23.0 (P < 0.05). In a sample of 153 men, semen parameters, testosterone levels, and testis volume were not significantly influenced. Marijuana use had the broader hormonal changes since it influences estradiol (P = 0.000; B = -11.616), prolactin (P = 0.000; B = 3.211), SHBG levels (P = 0.017; B = 7.489), and T/E2 (P = 0.004; B = 14.030). Sedentary lifestyle (P = 0.028; B = 1.279) and tobacco smoking (P = 0.031; B = -2.401) influenced the prolactin levels. Marijuana is associated with hormonal imbalance in this infertile cohort by lowering estradiol levels and inhibiting aromatase function.

Sperm motility in asthenozoospermic semen samples can be improved by incubation in a continuous single culture medium (CSCM®).

Standard protocols for clinical in vitro fertilization (IVF) laboratories recommend incubating semen at 37°C in 5% CO2 without strictly specifying which medium should be used or for how long. This study aimed to test the most common different incubation media used in Latin American andrology and micromanipulation laboratories and verify which, if any, is the most appropriate medium to improve asthenozoospermic semen samples' motility in the infertile male population. Ejaculates (136) collected from asthenozoospermic men were divided into two cohorts with similar characteristics (cohort 1; n = 28 and cohort 2; n = 108). Cohort 1 was used to evaluate the optimal incubation time with regard to unprepared asthenozoospermic sample sperm motility. After defining an optimal incubation period of 2 h, cohort 2 was used to evaluate which of the four media commonly used in IVF clinics (continuous single culture medium = CSCM®; SpermRinse medium = SR®; in vitro fertilization medium = G-IVF® and human tubal fluid medium = HTF®) was preferred for semen samples from asthenozoospermic patients. Overall, it was determined that a 2-h incubation in CSCM® medium led to the highest asthenozoospermic sperm motility. Thus, this simple, cost-effective, easily reproducible protocol could prove extremely useful for andrology laboratories working with IVF clinics dealing with asthenozoospermic semen specimens. This is particularly relevant since the incidence of the latter is on the rise as semen quality decreases around the globe.Abbreviations: ANOVA: Analysis of variance; ARTs: Assisted reproductive techniques; BWW: Biggers, Whitten, and Whittingham; CO2: Carbon dioxide; CPM: counted per minute; CSCM: Continuous Single Culture Medium; DAB: 3.3'- diaminobenzidine; DFI: DNA Fragmentation Index; DMSO: Dimethyl sulfoxide; G-IVF: In Vitro Fertilization Medium; GSH: Glutathione; GPx: glutathione peroxidase; HDS: High DNA Stainability; HSA: Human Serum Albumin; HTF: Human Tubal Fluid; HYP: Hyperactivity; ICSI: Intracytoplasmic sperm injection; IUI: Intrauterine insemination; IVF: in vitro fertilization; LIN: Linearity; ROS: Reactive Oxygen Species-level; SC: Sperm concentration; SCA: Sperm Computer Analysis; SCSA: Sperm Chromatin Structural Assay; SR: SpermRinse medium; SSS: Synthetic Serum Substitute; STR: Straightness; SOD: superoxide dismutase; TNE: Tris-Borate-EDTA; TSC: Total sperm count; VAP: Mean velocity; VCL: Curvilinear velocity; VSL: Linear velocity; WHO: World Health Organization; WOB: Wobble; spz: spermatozoa; AO: antioxidant.

Viral infections and implications for male reproductive health.

Viral infections have haunted humankind since times immemorial. Overpopulation, globalization, and extensive deforestation have created an ideal environment for a viral spread with unknown and multiple shedding routes. Many viruses can infect the male reproductive tract, with potential adverse consequences to male reproductive health, including infertility and cancer. Moreover, some genital tract viral infections can be sexually transmitted, potentially impacting the resulting offspring's health. We have summarized the evidence concerning the presence and adverse effects of the relevant viruses on the reproductive tract (mumps virus, human immunodeficiency virus, herpes virus, human papillomavirus, hepatitis B and C viruses, Ebola virus, Zika virus, influenza virus, and coronaviruses), their routes of infection, target organs and cells, prevalence and pattern of virus shedding in semen, as well as diagnosis/testing and treatment strategies. The pathophysiological understanding in the male genital tract is essential to assess its clinical impact on male reproductive health and guide future research.

Serum vitamin D content is associated with semen parameters and serum testosterone levels in men.

The present study aimed to evaluate the influence of serum vitamin D levels on semen quality and testosterone levels. This is a cross-sectional study conducted at Androscience, Science and Innovation Center in Andrology and High-Complex Clinical and Andrology Laboratory in Sao Paulo, Brazil, with 508 male patients, aged 18-60 years, from 2007 to 2017. Seminal parameters and serum sexual hormones were correlated with serum vitamin D concentrations in 260 men selected by strict selection criteria. Patients were divided into normozoospermic group (NZG, n = 124) and a group with seminal abnormalities (SAG, n = 136). Evaluation included complete physical examination, past medical history, habits and lifestyle factors, two complete seminal analysis with sperm functional tests, serum levels of 25-hydroxy-vitamin D3(25(OH)VD3), total and free testosterone, luteinizing hormone (LH), follicle-stimulating hormone (FSH), sex hormone-binding globulin (SHBG), total cholesterol, homeostatic model assessment of insulin resistance (HOMA-IR) index, and karyotype. The mean concentration of 25(OH)VD3was significantly lower in the SAG (P < 0.001) and positively correlated with all baseline seminal parameters and total testosterone levels. In addition, serum vitamin D3concentration was found to be positively correlated with sperm concentration (ß= 2.103; P < 0.001), total number of spermatozoa with progressive motility (ß = 2.069; P = 0.003), total number of motile spermatozoa (ß = 2.571; P = 0.015), and strict morphology (ß = 0.056; P = 0.006), regardless of other variables. This is the first comparative study to address the issue of serum vitamin D3content between normozoospermic patients and those with sperm abnormalities. It clearly demonstrates a direct and positive relationship between serum vitamin D level and overall semen quality, male reproductive potential, and testosterone levels.

Melatonin and Caffeine Supplementation Used, Respectively, as Protective and Stimulating Agents in the Cryopreservation of Human Sperm Improves Survival, Viability, and Motility after Thawing compared to Traditional TEST-Yolk Buffer.

Cryopreservation processes can damage spermatozoa and impair structural and functional cell characteristics. Plasma, nuclear membranes, and cellular organelles can suffer from the freeze and thaw process. This study evaluates the protective and stimulant effect of melatonin and caffeine supplementation on the functional characteristics of human spermatozoa before and after freezing. Thirty seminal samples from normozoospermic men aged 19-45 years old collected between October 2012 and May 2017 were included. Semen samples were supplemented with either 2 mM melatonin (MEL) prior to cryopreservation, 2 mM caffeine (CAF) in postthaw, or CAF and MEL (CM) in precryopreservation and postthaw, respectively. Kinetics and seminal parameters, mitochondrial activity, DNA fragmentation, and reactive oxygen species (ROS) levels were analyzed before and after cryopreservation. A significant reduction in sperm concentration, total and progressive motility, sperm kinetics, and mitochondrial activity, as well as a significant increase in DNA fragmentation and ROS production in postthaw samples compared to fresh samples, was identified. After administration of a caffeine and/or melatonin supplement, there was a significant increase in progressive motility in the CAF (p = 0.005) and CM (p = 0.048) groups, as well as mitochondrial activity in the CM group (p < 0.05). Cryopreservation has negative effects on overall sperm quality and increases ROS production. A combination of caffeine and melatonin in prefreeze and postthaw sperm samples has proven to be a very effective and simple way to improve semen quality. This will be particularly useful for initial low-quality semen samples, those which suffer the most from the freezing/thawing process.