Male Fertility · Male Endocrine and Genetic Factors
Molecular karyotyping of single sperm with nuclear vacuoles identifies more chromosomal abnormalities in patients with testiculopathy than fertile controls: implications for ICSI
Is there a difference between molecular karyotype of single sperm selected by high-magnification microscopy from infertile patients with testicular damage and from proven fertile controls?
Summary Answer
The molecular karyotype of single sperm from patients with testiculopathy had a significantly higher percentage of chromosomal alterations than fertile controls.
What Is Known Already
Infertile patients with testicular impairment have many sperm with aneuploidies and/or increased structural chromosome alterations. In these patients, sperm use by ICSI has poor outcome and raises concerns about the possible impact on pregnancy loss and transmission of genes abnormalities in offspring. High-magnification microscopy has been recently introduced to select morphologically better sperm aimed at improving ICSI outcome. However, there are no studies evaluating the molecular karyotype of sperm selected by this method.
Study Design, Size, Duration
Three consecutive infertile patients with oligozoospermia due to testicular damage and three age-matched proven fertile men attending a tertiary care center, were enrolled in the study from September to November 2014. Inclusion criteria of patients were age ≥30 ≤35 years, at least 2 years of infertility, oligozoospermia (sperm count below 10 million), reduced testicular volumes high FSH plasma levels and absence of altered karyotype, Y chromosome microdeletions, cystic fibrosis transmembrane conductance regulator gene mutations, sperm infections, cigarette smoking, varicocele, obesity.
Participants/Materials, Setting, Methods
Participants were evaluated for sperm parameters, sex hormones and testicular color-doppler ultrasound. From each semen sample, 20 sperm with large vacuoles (LVs), 20 with small vacuoles (SVs) and 20 with no vacuoles (NVs) were retrieved individually by a micromanipulator system. Each cell was further analyzed by whole genome amplification and array comparative genomic hybridization (aCGH).
Main Results and the Role of Chance
The aCGH allowed us to detect chromosomal aneuploidies, unbalanced translocations and complex abnormalities. Sperm selected from infertile patients showed a higher percentage of abnormal molecular karyotypes than controls (19.4 versus 7.7%, respectively, P < 0.001). In particular, sperm with LV and SV showed 38.3 and 20.0% abnormal karyotype in infertile men versus 18.3 and 5.0% in controls, respectively (both P < 0.01). Complex abnormalities were found only in the LV category. An abnormal karyotype was never found in NV sperm from both patients and controls.
Limitations, Reasons for Caution
The main limitation of this study is the low number of included subjects. Moreover, a time of writing we have no data regarding the ICSI outcome using LV, SV or NV sperm. This is the first study evaluating the molecular karyotype of single sperm selected by high-magnification microscopy and further confirmation of the data is needed.
Wider Implications of the Findings
Our data showed that sperm from infertile patients with testicular impairment have a higher percentage of abnormal molecular karyotypes than sperm from fertile controls. Therefore, if confirmed, our data suggest that the use of individually retrieved NV sperm may improve ICSI outcome in infertile men with testicular damage.
Mazzilli R et al., 2026·Journal of endocrinological investigation
Male factor could contribute, alone or in combination with female factor, to the inability to conceive in a couple in more than half of cases. The effect of male factor infertility (MFI) on embryological assisted reproductive technology (ART) outcomes remains an ongoing discussion.
to evaluate the impact of MFI on embryo aneuploidy rates, to better understand the role and possible indication for Preimplantation genetic testing for aneuploidies (PGT-A), considering the role of sperm characteristics, paternal age, sperm DNA fragmentation and sperm aneuploidies.
this narrative review included all available articles, published up to January 2026.
Available evidence, often based on heterogeneous and old-fashioned methodologies, suggests that MFI may impair embryonic development, particularly in terms of fertilization rate and blastulation rate, more than embryo euploidy; however, a comprehensive evaluation of MFI should be integrated into clinical practice in the context of couple infertility, to also optimize the efficiency and outcomes of ART. Promising results emerged from sperm DNA fragmentation as a tool to predict embryo euploidy, but further investigations involving larger sample sizes and improved standardization are required.
Rocca MS et al., 2026·Journal of translational medicine·Free to read
Standard semen analysis has little prognostic value in distinguishing fertile from infertile men. Furthermore, in men with semen parameters within the reference ranges, it cannot distinguish fertile men from infertile men, i.e. partners of couples with idiopathic infertility. Oxidative stress, mitochondrial dysfunction, sperm telomere shortening, and DNA fragmentation have been proposed as contributors to impaired male fertility. However, these biomarkers have not been evaluated as a whole in subjects with normal semen parameters, partners of fertile and infertile couples.
To characterise a multidimensional panel of sperm biomarkers-including sperm telomere length (STL), mitochondrial DNA copy number (mtDNAcn), reactive oxygen species (ROS), lipid peroxidation (LP), sperm chromatin dispersion (SCD), and respiratory control ratio (RCR)-and to identify whether these parameters might discriminate, in men with normal semen parameters, infertile men from fertile controls.
A total of 150 men with semen parameters within the normal ranges were enrolled: 47 male partners of couples with idiopathic infertility and 103 fertile controls. STL and mtDNAcn were quantified by qPCR; ROS were assessed using OxiSperm®II with semiquantitative imaging; LP was measured spectrophotometrically in seminal plasma; SCD was used to determine DNA fragmentation; and mitochondrial function was evaluated by oxygen consumption and RCR. Correlations between biomarkers and semen parameters were analysed using Pearson or Spearman coefficients, and intergroup comparisons were adjusted for age.
Semen parameters did not differ significantly between male partners of fertile and infertile couples. compared to men of fertile couples, men of infertile couples exhibited significantly shorter STL (0.57 ± 0.59 vs 1.21 ± 1.13, p_adj = 0.001) and higher oxidative stress, with both ROS (8300.9 ± 4214.8 vs 6555.3 ± 3394.3, p_adj = 0.027) and LP (88.33 ± 55.50 vs 40.29 ± 46.98, p_adj < 0.001) markedly elevated. mtDNAcn, SCD, and RCR showed no difference between the groups. Across the entire cohort, STL correlated negatively with ROS and LP and positively with RCR. ROS correlated negatively with total sperm count, motility and RCR, and positively with LP. LP displayed the strongest pattern of associations, correlating negatively with concentration, total count, motility, STL and RCR, and positively with age and volume.
Among men with normal semen analysis, partners of couples with idiopathic infertility exhibit a distinct sperm molecular profile characterised by telomere shortening and oxidative imbalance. STL, ROS and LP emerged as age-independent biomarkers associated with infertility status and showed promising discriminatory ability in this cohort, supporting their integration as second-level tests to complement routine semen analysis in cases of normozoospermia.
Ponce MR et al., 2026·Andrology·Free full text on PubMed Central
Transgender individuals assigned male at birth (AMAB) may choose to preserve their fertility prior to starting gender-affirming hormone therapy (GAHT). However, limited data exist regarding the baseline reproductive and hormonal characteristics of this population before and after GAHT.
To characterize semen quality and hormonal profiles in transgender AMAB individuals prior to GAHT and compare findings with cisgender men and with transgender individuals who discontinued GAHT for at least 3 months.
This retrospective study included transgender AMAB individuals from two tertiary andrology centers who underwent sperm cryopreservation before GAHT initiation (treatment-naïve group). Clinical evaluation included anthropometric measures, testicular volume assessment, varicocele detection, and sex hormone measures. Semen parameters were analyzed according to WHO criteria and compared with those of cisgender men and previously treated transgender individuals.
Thirty-two treatment-naïve transgender AMAB individuals were included. Hormonal parameters were generally within reference ranges. Only 46.9% of treatment-naïve individuals met WHO criteria for normozoospermia, compared with 75.5% of cisgender controls. Compared with nine previously treated individuals, estradiol levels were higher and seminal volume lower. A higher frequency of seminal abnormalities was observed, although not statistically significant.
A substantial proportion of treatment-naïve transgender AMAB individuals exhibited impaired semen parameters prior to GAHT initiation. Prior GAHT exposure showed a trend toward poorer semen quality. These findings support early fertility counseling and preservation in transgender individuals prior to GAHT initiation.
Besides gonadal involvement (hypogonadism, male factor infertility, and testicular hypotrophy), patients with Klinefelter syndrome (KS) may suffer from several extra-gonadic complications, including neoplastic events.
The aim of this review is to summarize all major clinical evidence dealing with the association between KS and neoplastic diseases and provide practical suggestions for the management of patients with KS regarding neoplastic risk.
This narrative review was conducted through a comprehensive search of the PubMed database, using combinations of the following keywords: "Klinefelter syndrome," "cancer," "neoplasm," "breast cancer," "germ cell tumor," "lymphoma," and "testosterone replacement therapy".
KS is associated with a higher risk of breast cancer and mediastinal germ cell tumors and an apparent higher risk of hematological malignancies, in particular non-Hodgkin lymphoma and leukemia. Evidence on testicular cancer is limited, with no clear increase in risk attributable to KS, while prostate cancer has a lower risk and lower mortality rate.
Given the complexity of the syndrome and the limited available evidence, regular clinical follow-up with targeted investigations when clinically indicated may facilitate early diagnosis and management of associated comorbidities.
Related research
Male Endocrine and Genetic Factors · Genetic Causes of Male Infertility
Foresta C et al., 2005·The Journal of clinical endocrinology and metabolism
Recent reports suggest that children born after intracytoplasmic sperm injection performed for male factor infertility are at increased risk of congenital malformations and chromosome aberrations. To explain these observations, we hypothesized that infertile men may be more likely than fertile men to have genetic abnormalities. We studied 750 severely oligozoospermic men (sperm count <5 million/ml) who were candidates for intracytoplasmic sperm injection, and 303 fertile men. We analyzed the peripheral blood karyotype, the Y chromosome long arm for detection of microdeletions in the azoospermia factors, and mutations in the cystic fibrosis gene and the androgen receptor gene. We also analyzed sperm for chromosome aneuploidies among the 421 men who subsequently entered the in vitro fertilization program. A total of 104 genetic abnormalities were diagnosed, corresponding to a frequency of 13.9% (104 of 750). Chromosomal aberrations were present in 5.6% (42 of 750) of infertile men and 0.3% of controls (one of 295), and they were in most cases alterations of the sex chromosomes. Y chromosome long-arm microdeletions were detected in 6.0% (45 of 750) of infertile men and most frequently included the azoospermia factor c, whereas no cases were found in controls (zero of 210). Mutations in the cystic fibrosis gene were diagnosed in 1.2% (nine of 750) of infertile men and 1.0% of controls (three of 303), and mutations in the androgen receptor gene were found in 1.1% (eight of 750) of infertile men and none of the 188 controls. Sperm sex chromosome aneuploidies were increased in men with karyotype anomalies and Y chromosome microdeletions as well as in subjects without constitutional genetic abnormalities. This study shows that the frequency of genetic alterations is increased among men with severe spermatogenic impairment. Genetic tests and genetic counseling should therefore be considered in oligozoospermic men who are candidates for intracytoplasmic sperm injection.
Male Endocrine and Genetic Factors · Genetic Causes of Male Infertility
Ferlin A et al., 2007·The Journal of clinical endocrinology and metabolism
An explosive growth in Y chromosome long arm (Yq) microdeletion testing demand for male infertility occurred in the past few years. However, despite the progresses in the biology of this chromosome, a number of molecular and clinical concerns are not supported by definitive data.
The objective was to provide information on the type and prevalence of microdeletions in infertile males, indication for testing, genotype-phenotype correlation, sperm aneuploidies, and genetic counseling.
We performed a prospective study from January 1996 to December 2005 in an academic clinic.
We studied 3073 consecutive infertile men, of which 625 were affected by nonobstructive azoospermia and 1372 were affected by severe oligozoospermia. Ninety-nine patients with microdeletions are described here.
Yq microdeletions, seminal analysis, reproductive hormones, testicular cytology/histology, and sperm sex chromosomes aneuploidies were used as outcome measures.
The prevalence of microdeletions was 3.2% in unselected infertile men, 8.3% in men with nonobstructive azoospermia, and 5.5% in men with severe oligozoospermia. Only 2 of 99 deletions were found in men with more than 2 million sperm/ml. No clinical data are useful to identify a priori patients with higher risk of Yq microdeletions. Most deletions are of the AZFc-b2/b4 subtype and are associated with variable spermatogenic phenotype, with sperm present in 72% of the cases. Complete AZFa and AZFb (P5/Proximal P1) deletions are associated with Sertoli cell-only syndrome and alterations in spermatocyte maturation, respectively, whereas partial deletions in these regions are associated with milder phenotype and frequent presence of sperm. Men with AZFc-b2/b4 deletions produce a higher percentage of sperm with nullisomy for the sex chromosomes and XY-disomy.
This extensive clinical research expands the knowledge on genotype-phenotype relationships and confirms that the identification of Yq microdeletions has significant diagnostic and prognostic value, adding useful information for genetic counseling in these patients.
Male Endocrine and Genetic Factors · Genetic Causes of Male Infertility
Selice R et al., 2010·Journal of endocrinological investigation
Klinefelter syndrome (KS) (47,XXY) is the most common sex chromosomal disorder, and it is a frequent form of male hypogonadism and infertility. Although the majority of these patients are azoospermic, they might have severe oligozoospermia or residual single-residual foci with spermatogenesis in the testis.
We report our experience on sperm retrieval in the ejaculate and testis, and evaluate the frequency of chromosome abnormalities in sperm of KS.
Eighty-four 47,XXY KS were evaluated with seminal analysis, body hair distribution, reproductive hormones, ultrasonographic scanning of the testis and prostate, bilateral testicular sperm extraction (TESE), sperm or testicular cells sex chromosomes aneuploidies.
Out of 84 patients, 7 (7/84; 8.3%) had sperm in the ejaculate. Out of the 77 azoospermic patients, 24 underwent TESE and 9 (9/24; 37.5%) had successful sperm recovery. The comparison of reproductive hormones, age and testicular volume did not show significant differences between patients with and without successful sperm recovery in semen or TESE . Patients without successful sperm recovery in semen analysis or TESE had signs of hypoandrogenism more evident than patients with successful sperm recovery. Patients with KS produced a higher number of sperm aneuploidy with respect to normozoospermic fertile controls and non-genetic severely oligozoospermic men.
Men with KS are not always sterile. In some of these patients sperm can be found in semen or in the testis, but the proportion of sperm aneuploidy is high. Signs of hypoandrogenism seem to be associated with low sperm recovery rate.
Male Endocrine and Genetic Factors · Genetic Causes of Male Infertility
Ferlin A et al., 2005·Cytogenetic and genome research
The most common type of karyotype abnormality detected in infertile subjects is represented by Klinefelter's syndrome, and the most frequent non-chromosomal alteration is represented by Y chromosome long arm microdeletions. Here we report our experience and a review of the literature on sperm sex chromosome aneuploidies in these two conditions. Non mosaic 47,XXY Klinefelter patients (12 subjects) show a significantly lower percentage of normal Y-bearing sperm and slightly higher percentage of normal X-bearing sperm. Consistent with the hypothesis that 47,XXY germ cells may undergo and complete meiosis, aneuploidy rate for XX- and XY-disomies is also increased with respect to controls, whereas the percentage of YY-disomies is normal. Aneuploidy rates in men with mosaic 47,XXY/46,XY (11 subjects) are lower than those observed in men with non-mosaic Klinefelter's syndrome, and only the frequency of XY-disomic sperm is significantly higher with respect to controls. Although the great majority of children born by intracytoplasmic sperm injection from Klinefelter subjects are chromosomally normal, the risk of producing offspring with chromosome aneuploidies is significant. Men with Y chromosome microdeletions (14 subjects) showed a reduction of normal Y-bearing sperm, and an increase in nullisomic and XY-disomic sperm, suggesting an instability of the deleted Y chromosome causing its loss in germ cells, and meiotic alterations leading to XY non-disjunction. Intracytoplasmic injection of sperm from Y-deleted men will therefore transmit the deletion to male children, and therefore the spermatogenic impairment, but raises also concerns of generating 45,X and 47,XXY embryos.
Male Fertility › Male Endocrine and Genetic Factors › Genetic Causes of Male Infertility · Assisted Reproduction › Add-Ons and Adjuncts › Adjunct Evidence Base · Genetics and Immunology › Reproductive Genetics › Chromosomal Abnormalities
Barbara Sartini, Ilaria Cosci, Alessandro Bertoldo, Elena Speltra
Barb Sartini, B Sartini, I Cosci, A Bertoldo, E Speltra
PMID 26405261 26405261 DOI 10.1093/humrep/dev202 10.1093/humrep/dev202 Garolla et al. 2015, Garolla 2015
00240q980