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Study Question: Can genome-wide genotyping data be analysed using a hypothesis-driven approach to enhance the understanding of the genetic basis of severe spermatogenic failure (SPGF) in male infertility?, Summary Answer: Our findings revealed a significant association between SPGF and the SHOC1 gene and identified three novel genes ( PCSK4 , AP3B1 , and DLK1 ) along with 32 potentially pathogenic rare variants in 30 genes that contribute to this condition., What Is Known Already: SPGF is a major cause of male infertility, often with an unknown aetiology. SPGF can be due to either multifactorial causes, including both common genetic variants in multiple genes and environmental factors, or highly damaging rare variants. Next-generation sequencing methods are useful for identifying rare mutations that explain monogenic forms of SPGF. Genome-wide association studies (GWASs) have become essential approaches for deciphering the intricate genetic landscape of complex diseases, offering a cost-effective and rapid means to genotype millions of genetic variants. Novel methods have demonstrated that GWAS datasets can be used to infer rare coding variants that are causal for male infertility phenotypes. However, this approach has not been previously applied to characterize the genetic component of a whole case-control cohort., Study Design Size Duration: We employed a hypothesis-driven approach focusing on all genetic variation identified, using a GWAS platform and subsequent genotype imputation, encompassing over 20 million polymorphisms and a total of 1571 SPGF patients and 2431 controls. Both common (minor allele frequency, MAF > 0.01) and rare (MAF < 0.01) variants were investigated within a total of 1797 loci with a reported role in spermatogenesis. This gene panel was meticulously assembled through comprehensive searches in the literature and various databases focused on male infertility genetics., Participants/materials Setting Methods: This study involved a European cohort using previously and newly generated data. Our analysis consisted of three independent methods: (i) variant-wise association analyses using logistic regression models, (ii) gene-wise association analyses using combined multivariate and collapsing burden tests, and (iii) identification and characterisation of highly damaging rare coding variants showing homozygosity only in SPGF patients., Main Results and the Role of Chance: The variant-wise analyses revealed an association between SPGF and SHOC1 -rs12347237 ( P  =   4.15E-06, odds ratio = 2.66), which was likely explained by an altered binding affinity of key transcription factors in regulatory regions and the disruptive effect of coding variants within the gene. Three additional genes ( PCSK4 , AP3B1 , and DLK1 ) were identified as novel relevant players in human male infertility using the gene-wise burden test approach ( P  <   5.56E-04). Furthermore, we linked a total of 32 potentially pathogenic and recessive coding variants of the selected genes to 35 different cases., Large Scale Data: Publicly available via GWAS catalog (accession number: GCST90239721)., Limitations Reasons for Caution: The analysis of low-frequency variants presents challenges in achieving sufficient statistical power to detect genetic associations. Consequently, independent studies with larger sample sizes are essential to replicate our results. Additionally, the specific roles of the identified variants in the pathogenic mechanisms of SPGF should be assessed through functional experiments., Wider Implications of the Findings: Our findings highlight the benefit of using GWAS genotyping to screen for both common and rare variants potentially implicated in idiopathic cases of SPGF, whether due to complex or monogenic causes. The discovery of novel genetic risk factors for SPGF and the elucidation of the underlying genetic causes provide new perspectives for personalized medicine and reproductive counselling., Study Funding/competing Interests: This work was supported by the Spanish Ministry of Science and Innovation through the Spanish National Plan for Scientific and Technical Research and Innovation (PID2020-120157RB-I00) and the Andalusian Government through the research projects of 'Plan Andaluz de Investigación, Desarrollo e Innovación (PAIDI 2020)' (ref. PY20&#95;00212) and 'Proyectos de Investigación aplicada FEDER-UGR 2023' (ref. C-CTS-273-UGR23). S.G.-M. was funded by the previously mentioned projects (ref. PY20&#95;00212 and PID2020-120157RB-I00). A.G.-J. was funded by MCIN/AEI/10.13039/501100011033 and FSE 'El FSE invierte en tu futuro' (grant ref. FPU20/02926). IPATIMUP integrates the i3S Research Unit, which is partially supported by the Portuguese Foundation for Science and Technology (FCT), financed by the European Social Funds (COMPETE-FEDER) and National Funds (projects PEstC/SAU/LA0003/2013 and POCI-01-0145-FEDER-007274). S.S. is supported by FCT funds (10.54499/DL57/2016/CP1363/CT0019), ToxOmics-Centre for Toxicogenomics and Human Health, Genetics, Oncology and Human Toxicology, and is also partially supported by the Portuguese Foundation for Science and Technology (UIDP/00009/2020 and UIDB/00009/2020). S. Larriba received support from Instituto de Salud Carlos III (grant: DTS18/00101), co-funded by FEDER funds/European Regional Development Fund (ERDF)-a way to build Europe) and from 'Generalitat de Catalunya' (grant 2021SGR052). S. Larriba is also sponsored by the 'Researchers Consolidation Program' from the SNS-Dpt. Salut Generalitat de Catalunya (Exp. CES09/020). All authors declare no conflict of interest related to this study., Competing Interests: The authors declare that the research was conducted without any conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology.)

Background: Severe spermatogenic failure (SPGF) represents one of the most relevant causes of male infertility. This pathological condition can lead to extreme abnormalities in the seminal sperm count, such as severe oligozoospermia (SO) or non-obstructive azoospermia (NOA). Most cases of SPGF have an unknown aetiology, and it is known that this idiopathic form of male infertility represents a complex condition. In this study, we aimed to evaluate whether common genetic variation in TEX15 , which encodes a key player in spermatogenesis, is involved in the susceptibility to idiopathic SPGF. Materials and Methods: We designed a genetic association study comprising a total of 727 SPGF cases (including 527 NOA and 200 SO) and 1,058 unaffected men from the Iberian Peninsula. Following a tagging strategy, three tag single-nucleotide polymorphisms (SNPs) of TEX15 (rs1362912, rs323342, and rs323346) were selected for genotyping using TaqMan probes. Case-control association tests were then performed by logistic regression models. In silico analyses were also carried out to shed light into the putative functional implications of the studied variants. Results: A significant increase in TEX15 -rs1362912 minor allele frequency (MAF) was observed in the group of SO patients (MAF = 0.0842) compared to either the control cohort (MAF = 0.0468, OR = 1.90, p = 7.47E-03) or the NOA group (MAF = 0.0472, OR = 1.83, p = 1.23E-02). The genotype distribution of the SO population was also different from those of both control ( p = 1.14E-02) and NOA groups ( p = 4.33-02). The analysis of functional annotations of the human genome suggested that the effect of the SO-associated TEX15 variants is likely exerted by alteration of the binding affinity of crucial transcription factors for spermatogenesis. Conclusion: Our results suggest that common variation in TEX15 is involved in the genetic predisposition to SO, thus supporting the notion of idiopathic SPGF as a complex trait., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Guzmán-Jiménez, González-Muñoz, Cerván-Martín, Rivera-Egea, Garrido, Luján, Santos-Ribeiro, Castilla, Gonzalvo, Clavero, Vicente, Maldonado, Villegas-Salmerón, Burgos, Jiménez, Pinto, Pereira, Nunes, Sánchez-Curbelo, López-Rodrigo, Pereira-Caetano, Marques, Carvalho, Barros, Bassas, Seixas, Gonçalves, Lopes, Larriba, Palomino-Morales, Carmona, Bossini-Castillo, IVIRMA Group and Lisbon Clinical Group.)

We conducted a genome-wide association study in a large population of infertile men due to unexplained spermatogenic failure (SPGF). More than seven million genetic variants were analysed in 1,274 SPGF cases and 1,951 unaffected controls from two independent European cohorts. Two genomic regions were associated with the most severe histological pattern of SPGF, defined by Sertoli cell-only (SCO) phenotype, namely the MHC class II gene HLA-DRB1 (rs1136759, P = 1.32E-08, OR = 1.80) and an upstream locus of VRK1 (rs115054029, P = 4.24E-08, OR = 3.14), which encodes a protein kinase involved in the regulation of spermatogenesis. The SCO-associated rs1136759 allele (G) determines a serine in the position 13 of the HLA-DRβ1 molecule located in the antigen-binding pocket. Overall, our data support the notion of unexplained SPGF as a complex trait influenced by common variation in the genome, with the SCO phenotype likely representing an immune-mediated condition., (© 2022. The Author(s).)

Background: Previous studies in animal models evidenced that genetic mutations of KATNAL1, resulting in dysfunction of its encoded protein, lead to male infertility through disruption of microtubule remodelling and premature germ cell exfoliation. Subsequent studies in humans also suggested a possible role of KATNAL1 single-nucleotide polymorphisms in the development of male infertility as a consequence of severe spermatogenic failure., Objectives: The main objective of the present study is to evaluate the effect of the common genetic variation of KATNAL1 in a large and phenotypically well-characterised cohort of infertile men because of severe spermatogenic failure., Materials and Methods: A total of 715 infertile men because of severe spermatogenic failure, including 210 severe oligospermia and 505 non-obstructive azoospermia patients, as well as 1058 unaffected controls were genotyped for three KATNAL1 single-nucleotide polymorphism taggers (rs2077011, rs7338931 and rs2149971). Case-control association analyses by logistic regression assuming different models and in silico functional characterisation of risk variants were conducted., Results: Genetic associations were observed between the three analysed taggers and different severe spermatogenic failure groups. However, in all cases, the haplotype model (rs2077011*C | rs7338931*T | rs2149971*A) better explained the observed associations than the three risk alleles independently. This haplotype was associated with non-obstructive azoospermia (adjusted p = 4.96E-02, odds ratio = 2.97), Sertoli-cell only syndrome (adjusted p = 2.83E-02, odds ratio = 5.16) and testicular sperm extraction unsuccessful outcomes (adjusted p = 8.99E-04, odds ratio = 6.13). The in silico analyses indicated that the effect on severe spermatogenic failure predisposition could be because of an alteration of the KATNAL1 splicing pattern., Conclusions: Specific allelic combinations of KATNAL1 genetic polymorphisms may confer a risk of developing severe male infertility phenotypes by favouring the overrepresentation of a short non-functional transcript isoform in the testis., (© 2022 The Authors. Andrology published by Wiley Periodicals LLC on behalf of American Society of Andrology and European Academy of Andrology.)

We aimed to analyze the role of the common genetic variants located in the PIN1 locus, a relevant prolyl isomerase required to control the proliferation of spermatogonial stem cells and the integrity of the blood-testis barrier, in the genetic risk of developing male infertility due to a severe spermatogenic failure (SPGF). Genotyping was performed using TaqMan genotyping assays for three PIN1 taggers (rs2287839, rs2233678 and rs62105751). The study cohort included 715 males diagnosed with SPGF and classified as suffering from non-obstructive azoospermia (NOA, n = 505) or severe oligospermia (SO, n = 210), and 1058 controls from the Iberian Peninsula. The allelic frequency differences between cases and controls were analyzed by the means of logistic regression models. A subtype specific genetic association with the subset of NOA patients classified as suffering from the Sertoli cell-only (SCO) syndrome was observed with the minor alleles showing strong risk effects for this subset (OR add rs2287839 = 1.85 (1.17-2.93), OR add rs2233678 = 1.62 (1.11-2.36), OR add rs62105751 = 1.43 (1.06-1.93)). The causal variants were predicted to affect the binding of key transcription factors and to produce an altered PIN1 gene expression and isoform balance. In conclusion, common non-coding single-nucleotide polymorphisms located in PIN1 increase the genetic risk to develop SCO.

Background: Despite advances in routine prenatal cytogenetic testing, most anomalous fetuses remain without a genetic diagnosis. Exome sequencing (ES) is a molecular technique that identifies sequence variants across protein-coding regions and is now increasingly used in clinical practice. Fetal phenotypes differ from postnatal and, therefore, prenatal ES interpretation requires a large amount of data deriving from prenatal testing. The aim of our study was to present initial results of the implementation of ES to prenatal diagnosis in Polish patients and to discuss its possible clinical impact on genetic counseling., Methods: In this study we performed a retrospective review of all fetal samples referred to our laboratory for ES from cooperating centers between January 2017 and June 2021., Results: During the study period 122 fetuses were subjected to ES at our institution. There were 52 abnormal ES results: 31 in the group of fetuses with a single organ system anomaly and 21 in the group of fetuses with multisystem anomalies. The difference between groups was not statistically significant. There were 57 different pathogenic or likely pathogenic variants reported in 33 different genes. The most common were missense variants. In 17 cases the molecular diagnosis had an actual clinical impact on subsequent pregnancies or other family members., Conclusions: Exome sequencing increases the detection rate in fetuses with structural anomalies and improves genetic counseling for both the affected couple and their relatives.

Background: Severe spermatogenic failure (SpF) represents the most extreme manifestation of male infertility, as it decreases drastically the semen quality leading to either severe oligospermia (SO, <5 million spermatozoa/mL semen) or non-obstructive azoospermia (NOA, complete lack of spermatozoa in the ejaculate without obstructive causes)., Objectives: The main objective of the present study is to analyze in the Iberian population the effect of 6 single-nucleotide polymorphisms (SNPs) previously associated with NOA in Han Chinese through genome-wide association studies (GWAS) and to establish their possible functional relevance in the development of specific SpF patterns., Materials and Methods: We genotyped 674 Iberian infertile men (including 480 NOA and 194 SO patients) and 1058 matched unaffected controls for the GWAS-associated variants PRMT6-rs12097821, PEX10-rs2477686, CDC42BPA-rs3000811, IL17A-rs13206743, ABLIM1-rs7099208, and SOX5-rs10842262. Their association with SpF, SO, NOA, and different NOA phenotypes was evaluated by logistic regression models, and their functional relevance was defined by comprehensive interrogation of public resources., Results: ABLIM1-rs7099208 was associated with SpF under both additive (OR = 0.86, p = 0.036) and dominant models (OR = 0.78, p = 0.026). The CDC42BPA-rs3000811 minor allele frequency was significantly increased in the subgroup of NOA patients showing maturation arrest (MA) of germ cells compared to the remaining NOA cases under the recessive model (OR = 4.45, p = 0.044). The PEX10-rs2477686 SNP was associated with a negative testicular sperm extraction (TESE) outcome under the additive model (OR = 1.32, p = 0.034). The analysis of functional annotations suggested that these variants affect the testis-specific expression of nearby genes and that lincRNA may play a role in SpF., Conclusions: Our data support the association of three previously reported NOA risk variants in Asians (ABLIM1-rs7099208, CDC42BPA-rs3000811, and PEX10-rs2477686) with different manifestations of SpF in Iberians of European descent, likely by influencing gene expression and lincRNA deregulation., (© 2021 American Society of Andrology and European Academy of Andrology.)

Infertility is a growing concern in developed societies. Two extreme phenotypes of male infertility are non-obstructive azoospermia (NOA) and severe oligospermia (SO), which are characterized by severe spermatogenic failure (SpF). We designed a genetic association study comprising 725 Iberian infertile men as a consequence of SpF and 1058 unaffected controls to evaluate whether five single-nucleotide polymorphisms (SNPs), previously associated with reduced fertility in Hutterites, are also involved in the genetic susceptibility to idiopathic SpF and specific clinical entities. A significant difference in the allele frequencies of USP8 -rs7174015 was observed under the recessive model between the NOA group and both the control group ( p = 0.0226, OR = 1.33) and the SO group ( p = 0.0048, OR = 1.78). Other genetic associations for EPSTI1 -rs12870438 and PSAT1 -rs7867029 with SO and between TUSC1 -rs10966811 and testicular sperm extraction (TESE) success in the context of NOA were observed. In silico analysis of functional annotations demonstrated cis-eQTL effects of such SNPs likely due to the modification of binding motif sites for relevant transcription factors of the spermatogenic process. The findings reported here shed light on the molecular mechanisms leading to severe phenotypes of idiopathic male infertility, and may help to better understand the contribution of the common genetic variation to the development of these conditions.

Objective: To evaluate whether SOHLH2 intronic variation contributes to the genetic predisposition to male infertility traits, including severe oligospermia (SO) and different nonobstructive azoospermia (NOA) clinical phenotypes., Design: Genetic association study., Setting: Not applicable., Patient(s): Five hundred five cases (455 infertile patients diagnosed with NOA and 50 with SO) and 1,050 healthy controls from Spain and Portugal., Intervention(s): None., Main Outcome Measure(s): Genomic DNA extraction from peripheral blood mononuclear cells, genotyping of the SOHLH2 polymorphisms rs1328626 and rs6563386 using the TaqMan allelic discrimination technology, case-control association analyses using logistic regression models, and exploration of functional annotations in publicly available databases., Result(s): Evidence of association was observed for both rs6563386 with SO and rs1328626 with unsuccessful sperm retrieval after testicular sperm extraction (TESE-) in the context of NOA. A dominant effect of the minor alleles was suggested in both associations, either when the subset of patients with the manifestation were compared against the control group (rs6563386/SO: P=.021, odds ratio [OR] = 0.51; rs1328626/TESE-: P=.066, OR = 1.46) or against the group of patients without the manifestation (rs6563386/SO: P=.014, OR = 0.46; rs1328626/TESE-: P=.012, OR = 2.43). The haplotype tests suggested a combined effect of both polymorphisms. In silico analyses evidenced that this effect could be due to alteration of the isoform population., Conclusion(s): Our data suggest that intronic variation of SOHLH2 is associated with spermatogenic failure. The genetic effect is likely caused by different haplotypes of rs6563386 and rs1328626, which may predispose to SO or TESE- depending on the specific allelic combination., (Copyright © 2020 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)