Lack of trusted diagnostic tools for undetermined male infertility.

Semen analysis is the cornerstone of evaluating male infertility, but it is imperfect and insufficient to diagnose male infertility. As a result, about 20% of infertile males have undetermined infertility, a term encompassing male infertility with an unknown underlying cause. Undetermined male infertility includes two categories: (i) idiopathic male infertility-infertile males with abnormal semen analyses with an unknown cause for that abnormality and (ii) unexplained male infertility-males with "normal" semen analyses who are unable to impregnate due to unknown causes. The treatment of males with undetermined infertility is limited due to a lack of understanding the frequency of general sperm defects (e.g., number, motility, shape, viability). Furthermore, there is a lack of trusted, quantitative, and predictive diagnostic tests that look inside the sperm to quantify defects such as DNA damage, RNA abnormalities, centriole dysfunction, or reactive oxygen species to discover the underlying cause. To better treat undetermined male infertility, further research is needed on the frequency of sperm defects and reliable diagnostic tools that assess intracellular sperm components must be developed. The purpose of this review is to uniquely create a paradigm of thought regarding categories of male infertility based on intracellular and extracellular features of semen and sperm, explore the prevalence of the various categories of male factor infertility, call attention to the lack of standardization and universal application of advanced sperm testing techniques beyond semen analysis, and clarify the limitations of standard semen analysis. We also call attention to the variability in definitions and consider the benefits towards undetermined male infertility if these gaps in research are filled.

SARS-CoV-2 infection, male fertility and sperm cryopreservation: a position statement of the Italian Society of Andrology and Sexual Medicine (SIAMS) (Società Italiana di Andrologia e Medicina della Sessualità).

PURPOSE: The recent pandemic of severe acute respiratory syndrome (SARS) due to coronavirus (CoV) 2 (SARS-CoV-2) has raised several concerns in reproductive medicine. The aim of this review is to summarize available evidence providing an official position statement of the Italian Society of Andrology and Sexual Medicine (SIAMS) METHODS: A comprehensive Pubmed, Web of Science, Embase, Medline and Cochrane library search was performed. Due to the limited evidence and the lack of studies, it was not possible to formulate recommendations according to the Oxford 2011 Levels of Evidence criteria. RESULTS: Several molecular characteristics of the SARS-CoV-2 can justify the presence of virus within the testis and possible alterations of spermatogenesis and endocrine function. Orchitis has been reported as a possible complication of SARS-CoV infection, but similar findings have not been reported for SARS-CoV-2. Alternatively, the orchitis could be the result of a vasculitis as COVID-19 has been associated with abnormalities in coagulation and the segmental vascularization of the testis could account for an orchitis-like syndrome. Finally, available data do not support the presence of SARS-CoV-2 in plasma seminal fluid of infected subjects. CONCLUSION: Data derived from other SARS-CoV infections suggest that in patients recovered from COVID-19, especially for those in reproductive age, andrological consultation and evaluation of gonadal function including semen analysis should be suggested. Studies in larger cohorts of currently infected subjects are warranted to confirm (or exclude) the presence of risks for male gametes that are destined either for cryopreservation in liquid nitrogen or for assisted reproduction techniques.

Sperm selection methods in the 21st century.

Natural sperm selection in humans is a rigorous process resulting in the highest quality sperm reaching, and having an opportunity to fertilize, the oocyte. Relative to other mammalian species, the human ejaculate consists of a heterogeneous pool of sperm, varying in characteristics such as shape, size, and motility. Semen preparation in assisted reproductive technologies (ART) has long been performed using either a simple swim-up method or density gradients. Both methodologies provide highly motile sperm populations; however neither replicates the complex selection processes seen in nature. A number of methods have now been developed to mimic some of the natural selection processes that exist in the female reproductive tract. These methods attempt to select a better individual, or population of, spermatozoa when compared to classical methods of preparation. Of the approaches already tested, platforms based upon sperm membrane markers, such as hyaluronan or annexin V, have been used to either select or deselect sperm with varied success. One technology that utilizes the size, motility, and other characteristics of sperm to improve both semen analysis and sperm selection is microfluidics. Here, we sought to review the efficacy of both available and emerging techniques that aim to improve the quality of the sperm pool available for use in ART.

CASA: tracking the past and plotting the future.

The human semen sample carries a wealth of information of varying degrees of accessibility ranging from the traditional visual measures of count and motility to those that need a more computational approach, such as tracking the flagellar waveform. Although computer-aided sperm analysis (CASA) options are becoming more widespread, the gold standard for clinical semen analysis requires trained laboratory staff. In this review we characterise the key attitudes towards the use of CASA and set out areas in which CASA should, and should not, be used and improved. We provide an overview of the current CASA landscape, discussing clinical uses as well as potential areas for the clinical translation of existing research technologies. Finally, we discuss where we see potential for the future of CASA, and how the integration of mathematical modelling and new technologies, such as automated flagellar tracking, may open new doors in clinical semen analysis.

Interpretation of semen analysis using WHO 1999 and WHO 2010 reference values: Abnormal becoming normal.

Reference values of WHO 1999 manual were used for the interpretation of semen analysis until 2010 when new reference values were introduced which have lower cut-off compared to WHO 1999. Therefore, several men who previously were diagnosed abnormal based on their semen analysis have now become normal using new reference values. This study was conducted on semen analyses of 661 men from Middle East region and Pakistan. All semen analyses were reviewed using WHO 1999 and WHO 2010 criteria. Results showed that based on new criteria, 19% of the population changed classification from abnormal to normal when all normal semen parameters were considered. When at least one or more abnormal semen parameters were considered, of the total 661, 44% (288) of the population changed its classification from abnormal to normal with shift from WHO 1999 to 2010 criteria. These findings show that using new cut-off values, many more men are considered normal, but using old criteria (WHO 1999), the same men would be classified as abnormal. This warrants further discussion over the investigations and management plans for patients whose semen analyses fall below WHO 1999 but above WHO 2010 cut-offs.

Decline in semen quality in men in northern Taiwan between 2001 and 2010.

Semen quality is a key indicator of the male reproductive ability. The decline in semen quality has been debated for several decades. However, data on decline in semen quality might be inconsistent given factors such as geographical area, ethnicity, environmental exposure and lifestyle changes. This study aimed to provide information on the semen quality of men in Northern Taiwanese. We recruited 7,187 male participants between 2001 and 2010 from a reproductive medical center. The age of the participants ranged from 26 to 57 years, with a mean age of 36.9 ± 4.77 years. Semen analysis was performed through standardized methods outlined in the World Health Organization laboratory manual. Increasing age (per year) was significantly and negatively associated with semen volume (1.006 ml), progressive sperm motility, rapid progressive sperm motility and sperm with normal morphology (reduction by 1.010%, 1.013% and 1.002% per year, respectively). In addition, sperm concentration, semen volume, number of sperms, progressive sperm motility, rapid progressive sperm motility, and sperm with normal morphology were significantly reduced annually by 1.013 × 106/ml, 1.015 ml, 1.028 × 106, 1.021 %, 1.017% and 1.016%, respectively). Age-period-cohort (APC) analyses revealed that age and cohort had effects on reduction of the progressive and rapid progressive sperm motility. Moreover, all the sperm parameter values were significantly reduced annually. Our findings provide useful information for clinical practice and public health investigations of male reproductive health.

Semen quality trends in French regions are consistent with a global change in environmental exposure.

A retrospective study carried out recently in a large sample of men, close to the general population, has reported a significant and strong decline in sperm concentration and morphology in the whole of France between 1989 and 2005. We studied these trends within each region of France. Data were obtained from the Fivnat database. The study sample comprised male partners of sterile women in whom both tubes were absent or blocked. They were located at the assisted reproductive technology center. A Bayesian spatio-temporal model with parametric time trends, adjusted for age, was used to model overall time trends for each region. The results show that sperm concentration decreased in almost all regions of France. Among them, Aquitaine showed the highest decrease and Midi-Pyrénées had the lowest average for the whole period. Regarding total motility, most regions showed a slight increase while Bourgogne showed a steep and significant decrease. While considering sperm morphology, there was a decrease in most of the regions. The decrease in Aquitaine and Midi-Pyrénées was stronger when compared with the overall trend. In conclusion, a decrease in sperm concentration and morphology, already shown at the French metropolitan territory level, was observed in most regions of France. This is consistent with a global change in environmental exposure, according to the endocrine disruptor hypothesis especially. Indeed, ubiquitary exposure to chemicals has been growing in the general population of France since the 1950s, and the results do not appear to support the lifestyle hypothesis. The highest decreases and lowest values are consistently observed in two proximate regions that are both highly agricultural and densely populated.

Environmental & lifestyle factors in deterioration of male reproductive health.

BACKGROUND & OBJECTIVES: Male reproductive function in the general population has been receiving attention in recent years due to reports of various reproductive and developmental defects, which might be associated with various lifestyle and environmental factors. This study was carried out to determine the role of various lifestyle and environmental factors in male reproduction and their possible association with declining semen quality, increased oxidative stress as well as sperm DNA damage. METHODS: Semen samples were obtained from 240 male partners of the couples consulting for infertility problem. Semen analysis was carried out using WHO criteria and subjects were categorized on the basis of self reported history of lifestyle as well as environmental exposure. The oxidative and antioxidant markers; lipid peroxidation (LPO), superoxide dismutase (SOD) and catalase (CAT) as well as DNA damage by acridine orange test (AO) were determined. RESULTS: The presence of abnormal semen parameters was significantly higher among the lifestyle and/or environmental exposed subjects as compared to the non-exposed population. Further, the levels of antioxidants were reduced and sperm DNA damage was more among the lifestyle and/or environmental exposed subjects, though the changes were not significant. INTERPRETATION & CONCLUSIONS: These findings indicated that various lifestyle factors such as tobacco smoking, chewing and alcohol use as well as exposure to toxic agents might be attributed to the risk of declining semen quality and increase in oxidative stress and sperm DNA damage.

Proteomic revolution to improve tools for evaluating male fertility in animals.

Artificial insemination has been used as a common breeding technique for the rapid dissemination of important genes to improve livestock quality. However, infertility or subfertility in the male leads to the disintegration of the breeding system and large economic losses. Therefore, the development of an accurate diagnostic protocol for male fertility is of critical importance. To this end, many basic laboratory assays have been developed on the basis of semen analysis. Although these assays may provide a preliminary estimate of male fertility, their accuracies are often unacceptably low. Therefore, it is vital to develop new semen analyses that are simple to use and accurate. Proteomic approaches will shed light on understanding sperm physiology and help in developing new diagnostic tools for male fertility. The aim of this study was to review the retrospective semen analyses and prospective proteomic studies of male fertility determination and usefulness of proteomic approaches in diagnosing male fertility potential in animal industry.

Semen analysis: update on clinical value, current needs and future perspectives.

At present, evaluation of male reproductive function consists primarily of routine semen analysis, a collection of conventional microscopic assessments ideally performed following the guidelines set by the World Health Organization. While providing some insight into testicular function, these long-performed tests are limited in the information that they impart; more specifically, they are unable to predict true fertility potential. As a consequence, there is a need for the appraisal and consideration of newer semen parameters that may be more indicative of reproductive success. Although various novel assays have been introduced that broaden the scope of information available to both researcher and clinician, the utility of these tests remains limited due to the lack of standardisation of protocols and the absence of clinically established, dependable reference ranges. As such, it is not surprising that most of these parameters and their associated methods remain recommended for 'research purposes only'. With the burgeoning 'omics' revolution, nanotechnology and the development of new analytical instruments, there is now an opportunity for the identification and measurement of previously unknown features that may prove to be more indicative of each sperm's true functional status and capability. Once optimised, simplified, clinically validated and made more readily accessible, these new approaches hold the promise of forming the fulcrum upon which andrological investigations can enter a new era.

[Male infertility: recent developments]. / Infertilität bei Männern : Aktuelle Entwicklungen.

Although male reproductive functions are impaired in about half of the infertile couples seeking offspring, even today the examination and treatment of the male partner continues to be neglected. Despite the lack of evidence for a "sperm crisis", so highly touted in the press, the public remains worried, while the fact that male fertility declines beyond the age of 40 years and is accompanied by increasing genetic risks for the offspring goes largely unnoticed. In addition to a thorough physical examination supplemented by imaging techniques such as ultrasonography of the scrotal organs, semen analysis according to WHO guidelines, hormone determinations, and cyto- and molecular genetic analyses form part of the routine investigation of the infertile male. Few disorders have become subjects of rational treatment, such as hypogonadotropic hypogonadism with gonadotropins or GnRH, treatment of sexually transmitted diseases by antibiotics, and microsurgical reconstruction of blocked seminal ducts. Early treatment of maldescended testes in boys or changing lifestyle (e.g., discontinuation of smoking) are important preventive measures. In the age of evidence-based medicine, most empirical treatments have been demonstrated to be ineffective. Thus, pregnancy rates from patients with varicocele who underwent long-practiced surgical or radiologic interventional therapy were not different from those of patients receiving counseling. At present, in cases of non-obstructive azoospermia or severe oligoasthenoteratozoospermia, intracytoplasmic sperm injection (ICSI) using single sperm derived from semen or extracted from testicular biopsy tissue (TESE) represents the most successful treatment modality, although it remains symptomatic and not curative.

Semen variation in a population of fertile donors: evaluation in a French centre over a 34-year period.

Although it has been suspected that there is a decrease in semen quality over time, the results reported to date remain debatable because of methodological issues. The aim of the study reported here was to investigate the evolution of semen quality over time in a population of 1114 fertile candidates for sperm donation at CECOS, Tours, between 1976 and 2009. We investigated semen volume, sperm concentration, progressive motility, vitality, percentage of normal forms and multiple abnormalities index of the first ejaculate in this population. We did not find a decline in semen volume, whereas we observed a significant decrease in total sperm count (from 443.2 million in 1976 to 300.2 million in 2009), motility (from 64% in 1976 to 49% in 2009) and vitality (from 88% to 80%). Moreover, a significant decline in the percentage of normal forms was noted between 1976 and 1997 (from 67% to 26%) with a steady rise in the multiple abnormalities index between 1998 and 2009 (from 1.19 to 1.65). This study involving a population of fertile men from a restricted area revealed various degrees of decline in semen parameters over a period of 34 years. These findings will have to be compared with findings in other geographical areas.

What advances may the future bring to the diagnosis, treatment, and care of male sexual and reproductive health?

Over the past 40 years, since the publication of the original WHO Laboratory Manual for the Examination and Processing of Human Semen, the laboratory methods used to evaluate semen markedly changed and benefited from improved precision and accuracy, as well as the development of new tests and improved, standardized methodologies. Herein, we present the impact of the changes put forth in the sixth edition together with our views of evolving technologies that may change the methods used for the routine semen analysis, up-and-coming areas for the development of new procedures, and diagnostic approaches that will help to extend the often-descriptive interpretations of several commonly performed semen tests that promise to provide etiologies for the abnormal semen parameters observed. As we look toward the publication of the seventh edition of the manual in approximately 10 years, we describe potential advances that could markedly impact the field of andrology in the future.

Evolution of the WHO "Semen" processing manual from the first (1980) to the sixth edition (2021).

As stated clearly in all editions of the WHO Laboratory Manual for the Examination and Processing of Human Semen, the goal of the manual is to meet the growing needs for the standardization of semen analysis procedures. With constant advances in andrology and reproductive medicine and the advent of sophisticated assisted reproductive technologies for the treatment of infertility, the manual has been continuously updated to meet the need for new, evidence-based, validated tests to not only measure semen and sperm variables but also to provide a functional assessment of spermatozoa. The sixth edition of the WHO manual, launched in 2021, can be freely downloaded from the WHO website, with the hope of gaining wide acceptance and utilization as the essential source of the latest, evidence-based information for laboratory procedures required for the assessment of male reproductive function and health.

The past, present, and future of the semen analysis.

This month's Views and Reviews provides an added perspective to the World Health Organization laboratory manual for the examination and processing of human semen, which was recently published in the 6th edition. The first artice provides a historical context of the prior editions of the World Health Organization manuals and modifications adopted over the years. The next piece then provides additional perspectives on the methodologies used for the performance of semen analysis. The third article then examines some of the new semen analytic technologies and enhancements that have become more common over the years. Finally, the last article proposed where male reproductive testing will head in the coming years with emerging research and technologies.

Association between urinary bisphenol A concentrations and semen quality: A meta-analytic study.

Although preclinical research has revealed disrupting effects on male reproductive functions of bisphenol A (BPA), as yet clinical studies have led to inconsistent results. The present meta­analysis aims to establish the existence and the extent of the association between BPA exposure and semen quality. A thorough search of PubMed, Scopus and Web of Science databases was carried out. Only studies reporting data from multivariable linear regression analyses (ß-coefficients with 95% CI), assessing the association between urinary levels of BPA and standard semen parameters were included. Nine studies provided information about an overall sample of 2,399 men. Only the negative association between urinary BPA levels and sperm motility reached statistical significance (pooled ß-coefficient = -0.82; 95% CI: -1.51 to -0.12, p = 0.02; Pfor heterogeneity = 0.1, I2 = 42.9%). Yet, such a significance was lost after data adjustment for publication bias, as well as at the sensitivity analysis, when each of the two studies that contributed most to the overall estimate was excluded. In conclusion, the overall estimates of data produced by clinical studies point to a clinically negligible, if any, association between urinary BPA concentrations and semen quality. Further studies in workers at high risk of occupational exposure are warranted to corroborate the herein revealed weak correlation with a worse sperm motility.

ESHRE special interest group for andrology basic semen analysis course: a continued focus on accuracy, quality, efficiency and clinical relevance.

ESHRE has been running courses for basic semen analysis since 1994 and course material has been updated regularly in response to new findings and publications. Following publication of the 5th edition of the WHO laboratory manual, entitled WHO Laboratory Manual for the Examination and Processing of Human Semen (WHO5), the Subcommittee for training of the ESHRE Special Interest Group for Andrology evaluated potential amendments to its course. In respect of the updated ESHRE course, there are eight particular areas of discourse that are reviewed (i) maintaining the four-class differential motility count allowing distinction between rapid and slow progressive sperm for assisted reproduction technology. (ii) Maintaining the four-category assessment for sperm morphology with calculation of the teratozoospermic index. (iii) Continuing to advocate the use of three categories of results: 'normal', 'borderline' and 'abnormal' with respect to the clinical interpretation of the data. (iv) Presenting clear and unequivocal methods for performing assessments e.g. morphology. (v) Correcting the inconsistencies in WHO5, some of which are actually erroneous. (vi) Reducing the requirements for substantial extra work for what are unestablished improvements in accuracy and/or precision in the final results. (vii) Presentation of logical methods of sperm preparation. (viii) Discussion of the suddenly changed limits between fertile and subfertile men.

Is there a temporal trend in semen quality in Belgian candidate sperm donors and in sperm donors' fertility potential from 1995 onwards?

BACKGROUND: A recent meta-regression analysis reported a temporal trend in sperm count showing a significant decline in sperm count between 1973 and 2011. This decline is thought to affect fecundity. Moreover, semen quality is considered of key interest to public health given its association with all-cause male morbidity/mortality. The issue requires ongoing investigation due to geographical variation in semen quality and methodological errors in semen analysis. OBJECTIVE: To study whether there is a temporal trend in semen quality in Belgian candidate sperm donors and in sperm donors' fertility potential. MATERIALS AND METHODS: Retrospective analysis of samples provided by 439 candidate donors and pregnancy outcome in acceptors over a period of 23 years. RESULTS: A total of 807 specimens from 439 candidate donors were examined from January 1995 to December 2017 (Table S1). Sub-analyses performed with regard to TSC from 2010 onwards (weighing) revealed a significant negative trend (R2 =-0.033; ß=-0.18; CI: -0.16 to 0.07; p < 0.05). We found a statistically significant association between year of donation and morphology (R2 = 0.036; ß= -0.19; CI: -0.26 to -0.08; p < 0.0001). The mean (±SD) clinical pregnancy rate per effective donor recruited (n = 104), defined as the number of women with a clinical pregnancy, per number of women who initiated treatment with a donor's spermatozoa, was 68.5 (± 24.9) %. This measure did not show a significant change in function of year of donation. DISCUSSION: Candidate sperm donors represent a select group of men; as such, these results are not to be interpreted as representative for the general population. CONCLUSION: The study did not show a significant change in sperm concentration or fertility potential in sperm donors over a period of 23 years. However, a negative trend was found for TSC from 2010 onwards. Also, the results show a significant decrease in ideal morphology over time.

Machine-learning algorithm incorporating capacitated sperm intracellular pH predicts conventional in vitro fertilization success in normospermic patients.

OBJECTIVE: To measure human sperm intracellular pH (pHi) and develop a machine-learning algorithm to predict successful conventional in vitro fertilization (IVF) in normospermic patients. DESIGN: Spermatozoa from 76 IVF patients were capacitated in vitro. Flow cytometry was used to measure sperm pHi, and computer-assisted semen analysis was used to measure hyperactivated motility. A gradient-boosted machine-learning algorithm was trained on clinical data and sperm pHi and membrane potential from 58 patients to predict successful conventional IVF, defined as a fertilization ratio (number of fertilized oocytes [2 pronuclei]/number of mature oocytes) greater than 0.66. The algorithm was validated on an independent set of data from 18 patients. SETTING: Academic medical center. PATIENT(S): Normospermic men undergoing IVF. Patients were excluded if they used frozen sperm, had known male factor infertility, or used intracytoplasmic sperm injection only. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Successful conventional IVF. RESULT(S): Sperm pHi positively correlated with hyperactivated motility and with conventional IVF ratio (n = 76) but not with intracytoplasmic sperm injection fertilization ratio (n = 38). In receiver operating curve analysis of data from the test set (n = 58), the machine-learning algorithm predicted successful conventional IVF with a mean accuracy of 0.72 (n = 18), a mean area under the curve of 0.81, a mean sensitivity of 0.65, and a mean specificity of 0.80. CONCLUSION(S): Sperm pHi correlates with conventional fertilization outcomes in normospermic patients undergoing IVF. A machine-learning algorithm can use clinical parameters and markers of capacitation to accurately predict successful fertilization in normospermic men undergoing conventional IVF.