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Structural reorganization carriers usually present compromised fertility accompanied by an increased risk of producing gametes with chromosomal abnormalities that can be transmitted to the offspring. In part these imbalances are ascribed to result from the occurrence of meiotic disturbances produced by the rearrangements in the proper segregation of other chromosome pairs. This phenomenon of interference has been called interchromosomal effect (ICE). Several studies have been performed to assess the occurrence of ICE in structural reorganization carriers by analyzing the frequencies of numerical abnormalities in the gametes. Nevertheless, the occurrence and distribution of these disturbing events still is a controversial issue. In this work we present compiled data from 130 sperm fluorescent in situ hybridization (FISH) studies performed in carriers of the most frequent structural rearrangements in humans: 44 Robertsonian translocations, 66 reciprocal translocations and 13 inversions. Data from 7 complex//multiple rearrangements will be considered in a separate group. Significant increases of gametes with numerical abnormalities have been detected in all types of reorganization carriers. Among the groups of non-complex//multiple rearrangements, Robertsonian translocations appear to be the most prone to produce such interference (54.5%%) closely followed by reciprocal translocations (43.9%%). In contrast, ICE's were only detected in 7.7%% of the inversion carriers analyzed. The presence of complex//multiple rearrangements seems to be an important factor for promoting ICE, as 71.4%% of these carriers presented increased rates of gametes with numerical abnormalities. Altogether, almost half of the structural reorganization carriers (45.4%%) present a higher reproductive risk of producing aneuploid//diploid spermatozoa compared to the general population. This high incidence has been obtained by analyzing a small set of chromosomes, suggesting that underlying meiotic disorders could be present in these individuals. Further ICE studies in structural reorganization carriers will help to clarify the still unknown predisposing cytogenetic features that promote this phenomenon. [ABSTRACT FROM AUTHOR]

Objective: To determine the group of infertile patients that would benefit from sperm fluorescent in situ hybridization (FISH) analysis, the number of chromosomes to be analyzed, and the diagnostic interpretation of the results obtained. Design: A retrospective study of sperm FISH analyses. Setting: Universitat Autònoma de Barcelona. Patient(s): Three hundred nineteen infertile men. Intervention(s): Semen samples were processed for FISH. Main Outcome Measure(s): The frequencies of chromosomal abnormalities for chromosomes 13, 18, 21, X, and Y were compared to the seminogram, the somatic and meiotic karyotype, and the age. Result(s): The highest percentages of patients with an increased rate of sperm chromosome abnormalities were found in the oligozoospermic (50%), oligoasthenozoospermic (33.3%), and oligoasthenoteratozoospermic (21%) individuals. Low sperm count was the only parameter correlated with the percentage of chromosome abnormalities. The 14% of the individuals with a normal somatic karyotype had an increased rate of sperm chromosome abnormalities. This percentage was higher in the group with an altered somatic karyotype (36%) and in patients with meiotic abnormalities (26%). Conclusion(s): Sperm FISH studies are indicated when the oligo condition is present and in individuals with an abnormal somatic or meiotic karyotype. The analysis of chromosomes 21, X, and Y is enough to identify at-risk individuals. Significant differences in the rates of chromosome abnormalities should be taken into consideration regardless of the numerical value. [Copyright &y& Elsevier]

Objective: To evaluate the risk of four inversion carriers for producing unbalanced gametes. Design: Prospective analysis of sperm nuclei by fluorescence in situ hybridization (FISH). Setting: Universitat Autònoma de Barcelona. Patient(s): Four inversion carriers. Intervention(s): A semen sample from each patient was collected and prepared for FISH. Main Outcome Measure(s): The segregation outcome of each inversion was analyzed. The presence of interchromosomal effects (ICE) on chromosomes 13, 18, 21, X, and Y was also evaluated. Result(s): A variable production of unbalanced gametes, which implies a heterogeneous behavior of the inversions, was detected. This variability seems to be directly related to the size of the inversion, indicating that the production of recombinant gametes in inversion carriers would not be relevant when the inverted segment is smaller than 100 Mbp. Conclusion(s): Inversions have a well-defined reproductive effect on carriers. Carriers of inversions up to 100 Mbp have a low genetic reproductive risk and would not usually benefit from preimplantation genetic diagnosis. [Copyright &y& Elsevier]

Segregation behaviour studies in t(11;22) carriers have reported controversial results. Whereas some authors have detected a preponderance of 3:1 products, no evidence of such prevalence was found by others. This study reports a fluorescence in-situ hybridization (FISH) segregation analysis on decondensed spermatozoa in two brothers, carriers of the same t(11;22)(q23;11) rearrangement. Data revealed a similar meiotic segregation pattern in both carriers, 2:2 Alternate segregation being the most frequent (42.94 and 45%), while 3:1 genotypes were the least frequent in both patients, with percentages around 10%. The production of three chiasmata, based on the presence of G-light bands along the translocated segments and the presence of recombination sites at 11q and 22q distal regions, are proposed as the cause of a preponderance of the Alternate segregation. Interchromosomal effects involving chromosomes 13, 18, 21, X and Y were also evaluated. An increased frequency of sex chromosome disomies was detected in one patient. Reviewing the literature, a relationship between this phenomenon and the involvement of acrocentric chromosomes in the reorganization is suggested. FISH segregation and interchromosomal effects studies in spermatozoa are encouraged to gather information to establish the best approach for preimplantational genetic diagnosis in reorganization carriers. INSET: Mid-brain human dopaminergic neurons derived from ES cells. [ABSTRACT FROM AUTHOR]

: ObjectiveThe aim of the study was to evaluate the intracytoplasmic sperm injection outcome in a selected group of patients with oligoasthenozoospermia in relation to the results obtained from their meiotic analysis.: DesignRetrospective clinical study.: SettingAn assisted reproduction service and a university department.: Patient(s)One hundred thirty-seven men with oligoasthenozoospermia grouped in relation to their meiotic pattern.: Intervention(s)Two hundred twenty-four intracytoplasmic sperm injection cycles from 137 men with oligoasthenozoospermia in whom diagnostic meiotic analyses had been performed.: Main outcome measure(s)Fertilization, pregnancy, implantation, and abortion rates.: Result(s)There were no significant statistical differences in fertilization, pregnancy, implantation, or abortion rates among the three groups studied.: Conclusion(s)No statistically significant differences in fertilization, pregnancy, implantation, or abortion rates were found in patients with oligoasthenozoospermia in relation to the meiotic pattern. [Copyright &y& Elsevier]

DiGeorge/velocardiofacial syndrome (DGS/VCFS) is a disorder caused by a 22q11.2 deletion mediated by non-allelic homologous recombination (NAHR) between low-copy repeats (LCRs). We have evaluated the role of LCR22 genomic architecture and PRDM9 variants as DGS/VCFS predisposing factors. We applied FISH using fosmid probes on chromatin fibers to analyze the number of tandem repeat blocks in LCR22 in two DGS/VCFS fathers-of-origin with proven 22q11.2 NAHR susceptibility. Results revealed copy number variations (CNVs) of L9 and K3 fosmids in these individuals compared to controls. The total number of L9 and K3 copies was also characterized using droplet digital PCR (ddPCR). Although we were unable to confirm variations, we detected an additional L9 amplicon corresponding to a pseudogene. Moreover, none of the eight DGS/VCFS parents-of-origin was heterozygote for the inv(22)(q11.2) haplotype. PRDM9 sequencing showed equivalent allelic distributions between DGS/VCFS parents-of-origin and controls, although a new PRDM9 allele (L50) was identified in one case. Our results support the hypothesis that LCR22s variations influences 22q11.2 NAHR events, however further studies are needed to confirm this association and clarify the contribution of pseudogenes and rare PDRM9 alleles to NAHR susceptibility. [ABSTRACT FROM AUTHOR]

The production of functional spermatozoa through spermatogenesis requires a spatially and temporally highly regulated gene expression pattern, which in case of alterations, leads to male infertility. Changes of gene expression by chromosome anomalies, gene variants, and epigenetic alterations have been described as the main genetic causes of male infertility. Recent molecular and cytogenetic approaches have revealed that higher order chromosome positioning is essential for basic genome functions, including gene expression. This review addresses this issue by exposing well-founded evidences which support that alterations on the chromosome topology in spermatogenetic cells leads to defective sperm function and could be considered as an additional genetic cause of male infertility. [ABSTRACT FROM AUTHOR]

The purpose of this study is to provide novel information through Next Generation Sequencing (NGS) for the characterization of viral and bacterial RNA cargo of human sperm cells from healthy fertile donors. For this, RNA-seq raw data of poly(A) RNA from 12 sperm samples from fertile donors were aligned to microbiome databases using the GAIA software. Species of viruses and bacteria were quantified in Operational Taxonomic Units (OTU) and filtered by minimal expression level (>1% OTU in at least one sample). Mean expression values (and their standard deviation) of each species were estimated. A Hierarchical Cluster Analysis (HCA) and a Principal Component Analysis (PCA) were performed to detect common microbiome patterns among samples. Sixteen microbiome species, families, domains, and orders surpassed the established expression threshold. Of the 16 categories, nine corresponded to viruses (23.07% OTU) and seven to bacteria (2.77% OTU), among which the Herperviriales order and Escherichia coli were the most abundant, respectively. HCA and PCA displayed four clusters of samples with a differentiated microbiome fingerprint. This work represents a pilot study into the viruses and bacteria that make up the human sperm microbiome. Despite the high variability observed, some patterns of similarity among individuals were identified. Further NGS studies under standardized methodological procedures are necessary to achieve a deep knowledge of the semen microbiome and its implications in male fertility. [ABSTRACT FROM AUTHOR]

The transition in biopsy timing from blastomere to trophectoderm biopsy has led to a remarkable decrease in the percentage of undiagnosed blastocysts. However, patients with few or no euploid blastocysts can be affected by this residual percentage of diagnosis failure. The aim of this study is to assess whether blastocyst rebiopsy and revitrification is an efficient and safe procedure to be applied in cases of no results after analysis. Fifty-three patients agreed to the warming of 61 blastocysts to perform a second biopsy and PGT-A by aCGH. Only 75.4% of the blastocysts survived, reexpanded, and could be rebiopsied. After the second biopsy and analysis, 95.6% of the blastocysts were successfully diagnosed with an euploidy rate of 65.9%. Eighteen euploid blastocysts were warmed and transferred to 18 patients with a 100% survival and reexpansion rate. Seven clinical pregnancies have been achieved with 4 live births, 1 ongoing pregnancy, and 2 miscarriages. Thus, although few transfers of rebiopsied and revitrified blastocysts have been performed till date, our preliminary results show that this approach is efficient and safe to be applied for undiagnosed blastocysts, as it ultimately allows the transfer of euploid blastocysts and good clinical outcomes. [ABSTRACT FROM AUTHOR]

Objective To compare the microRNA (miRNA) expression profile in spermatozoa from three infertile populations vs. a group of fertile men. Design Evaluation of the expression level of 736 miRNAs in human spermatozoa using TaqMan quantitative reverse transcription–polymerase chain reaction. Setting University research facility. Patient(s) Semen samples with a single seminal alteration were collected from infertile individuals: asthenozoospermic (n = 10), teratozoospermic (n = 10), and oligozoospermic (n = 10). Intervention(s) None. Main Outcome Measure(s) Correlation of the expression level of each miRNA with seminal parameters, age, and chromosome instability; clustering of the individuals according to their miRNA expression profiles and influence of the seminogram, age, chromosome instability, and assisted reproductive technology outcome in the clustering; analysis of the differentially expressed miRNAs (DE-miRNAs) in each infertile population; genome annotation of these DE-miRNAs; and ontological analysis of their predicted targets. Result(s) The hsa-miR-34b-3p correlated with age, the hsa-miR-629-3p with sperm motility, and the hsa-miR-335-5p, hsa-miR-885-5p, and hsa-miR-152-3p with sperm concentration. The individuals clustered into two groups, and only the seminogram was differentially distributed. We identified 32 DE-miRNAs in the asthenozoospermic group, 19 in the teratozoospermic group, and 18 in the oligozoospermic group. The up-regulated miRNAs presented an enriched localization in introns, affecting relevant genes for spermatogenesis. The predicted targets of the DE-miRNAs contained critical genes associated to infertility, and their ontological analysis revealed significantly associated functions related to the seminal alterations of each group. Conclusion(s) Spermatozoa from patients with seminal alterations exhibit a differential miRNA profile. This provides new evidence that miRNAs have an essential role in spermatogenesis, contributing to the mechanisms involved in human fertility. [ABSTRACT FROM AUTHOR]

Purpose: To determine whether embryo mosaicism prevalence in preimplantation genetic testing for aneuploidy (PGT-A) cycles is associated with the trophectoderm biopsy technique used (a. number of laser pulses or b. the use of flicking or pulling) or the time to tubing. Methods: Prospective observational study performed in a single IVF-PGT-A setting from May 2019 to May 2021. Trophectoderm biopsies were analysed by next-generation sequencing. Mosaicism was analysed in relation to the biopsy methodology (number of laser pulses and pulling vs flicking), time elapsed from biopsy to tubing (min), and time of sample cryostorage from tubing to amplification (days). As a secondary objective, the number of laser pulses and biopsy methodology were studied in relation to clinical outcomes of transferred euploid blastocysts. Results: None of the analysed variables were associated to mosaicism prevalence. Multivariable regression analysis demonstrated that mosaicism prevalence was comparable either when > 3 laser pulses were used as compared to ≤ 3 (13.9% vs 13.8%, aOR = 0.8726 [0.60–1.28]) and pulling compared to flicking (13.1% vs 14.0%, aOR = 0.86 [0.60–1.23]). Moreover, neither the number of laser pulses during biopsy (> 3 vs ≤ 3) nor the technique used (pulling vs flicking) were associated with clinical pregnancy after the transfer of frozen-thawed euploid blastocysts (54.9% vs 55.2%, aOR = 1.05 [0.53–2.09]; 61.1% vs 52.9%, aOR = 1.11 [0.55–2.25], respectively). Conclusion: Our results suggest that, as long as the biopsy and tubing procedures are performed following standardized high quality procedures, no specific approach would increase the generation of artefactual mosaicism as a result of trophectoderm biopsy. Trophectoderm biopsies should be performed regardless of the methodology but always aiming on minimising blastocyst manipulation. [ABSTRACT FROM AUTHOR]

The aim of this study was to assess whether there is a relationship between numerical chromosome abnormalities and certain segregation modes in spermatozoa from Robertsonian translocation carriers. A sequential fluorescence in-situ hybridization protocol based on two successive hybridization rounds was performed on sperm samples from one t(13;22) and ten t(13;14) carriers. Patient inclusion criteria included the presence of a positive interchromosomal effect (ICE). In the first round, numerical abnormalities for chromosomes 15/22, 18, 21, X and Y were analysed. In the second round, the segregation outcome of the rearranged chromosomes was evaluated in the numerically abnormal spermatozoa detected in the first round, as well as in randomly assessed spermatozoa. Aneuploid spermatozoa showed statistical differences in all segregation modes when compared with randomly assessed spermatozoa: alternate (50.7% versus 84.3%), adjacent (36.6% versus 14.6%) and 3:0 (10.2% versus 1%). Diploid/multiple disomic spermatozoa showed differences in alternate (3.7% versus 84.3%) and 3:0 (67.6% versus 1%). We concluded that in Robertsonian translocation carriers that exhibit ICE, numerically abnormal spermatozoa preferentially contain unbalanced segregation products. This might be explained by heterosynapsis acting as a rescue mechanism that would lead to aberrant recombination, which is a predisposing factor for non-disjunction events. [ABSTRACT FROM AUTHOR]

Purpose: To find out the meiotic segregation behaviour of the t(1;8;2)(q42;p21;p15), to evaluate the occurrence of interchromosomal effects, and to determine whether there is an accumulation of unbalanced products in aneuploid/diploid gametes. Methods: A sequential FISH protocol based on two successive hybridization rounds over the same spermatozoa was performed to determine the segregation outcome of the rearranged chromosomes. The presence of numerical abnormalities for 13, 18, 21, X and Y was also evaluated by sperm FISH. Those aneuploid/diploid gametes were subsequently relocalized and analyzed for their segregation content through additional hybridization rounds. Results: The segregation pattern observed reported a very low production of normal/balanced gametes (11.7 %). Significant increased frequencies of diploidies and disomies for chromosomes X/Y and 18 were detected ( p < 0.001). Aneuploid and diploid spermatozoa displayed significant increases of 5:1, 6:0 and other unexpected disjunction modes ( p < 0.001). Conclusions: The strategy developed in this study is a reliable new approach to establish the full segregation pattern of complex chromosome rearrangements (CCR). Results corroborate the low number of normal/balanced spermatozoa produced by CCR carriers and support previous findings regarding an altered segregation pattern in gametes with numerical abnormalities. Altogether this confirms the importance of PGD as a tool to prevent the transmission of chromosomal abnormalities to the offspring in CCR patients. [ABSTRACT FROM AUTHOR]

Is partial compaction during morula formation associated with an embryo's developmental ability and implantation potential? Retrospective analysis of data from 196 preimplantation genetic testing for aneuploidy (PGT-A) cycles. Embryos starting compaction were grouped according to the inclusion or not of all the blastomeres in the forming morula (full compaction or partial compaction). The possible effect of maternal age and ovarian response on compaction was analysed. Morphokinetic characteristics, blastocyst formation rate, morphology and cytogenetic constitution of the obtained blastocysts were compared. Comparisons of reproductive outcomes after the transfer of euploid blastocysts from both groups were established. Finally, in a subset of embryos, the chromosomal constitution concordance of the abandoned cells and the corresponding blastocyst through trophectoderm biopsies was assessed. A total of 430 embryos failed to include at least one cell during compaction (partial compaction group [49.3%]), whereas the 442 remaining embryos formed a fully compacted morula (full compaction group [50.7%]). Neither female age nor the number of oocytes collected affected the prevalence of partial compaction morulae. Morphokinetic parameters were altered in embryos from partial compaction morulae compared with full compaction. Although an impairment in blastocyst formation rate was observed in partial compaction morulae (57.2% versus 70.8%, P < 0.001), both chromosomal constitution (euploidy rate: partial compaction [38.4%] versus full compaction [34.2%]) and reproductive outcomes (live birth rate: partial compaction [51.9%] versus full compaction [46.2%]) of the obtained blastocysts were equivalent between groups. A high ploidy correlation of excluded cells-trophectoderm duos was observed. Partial compaction morulae show a reduced developmental ability compared with full compaction morulae. Resulting blastocysts from both groups, however, have similar euploidy rates and reproductive outcomes. Cell exclusion might be a consequence of a compromised embryo development regardless of the chromosomal constitution of the excluded cells. [ABSTRACT FROM AUTHOR]

The objective of this study was to develop a methodology that permits the detection and separation of apoptotic cells in human testicular tissue and their subsequent cytogenetic analysis by fluorescence in situ hybridization (FISH). The sequential methodology consisted of five steps: 1) enzymatic disaggregation of testicular tissue, 2) specific staining of apoptotic cells, 3) cell sorting by flow cytometry, 4) cell fixation, and 5) FISH. Enzymatic disaggregation yielded cell counts that ranged from 1.7x105 to 5x106 cells, and viability values greater than 72%. The apoptotic (mean ± SD: 22% ± 5.3%) and viable (45.5% ± 7.3%) populations were identified and selected by flow cytometry and demonstrated purity values ranging between 62% and 100%. The paraformaldehyde fixation of the selected fractions resulted in cell loss values of less than 10%. The application of three treatments before FISH (membrane permeabilization, elimination of cytoplasmic components, and re-fixation of the sample) resulted in hybridization frequencies of greater than 98%. In both selected fractions, cells of all spermatogenic stages and Sertoli cells were identified. The methodology developed has enabled the preparation of a cellular suspension with optimal viability and counting, the efficient selection of the apoptotic population, and its analysis by cytogenetic techniques. The application of this methodology in testicular cells should help establish whether there is a direct relationship between chromosome anomalies and apoptosis. [ABSTRACT FROM AUTHOR]

Objective: To analyze whether the preferential proximity between acrocentric bivalents and the XY pair described at pachytene was maintained in metaphase I human spermatocytes. Design: Proximity frequencies of autosomic bivalents to the sex bivalent were evaluated with the analysis of meiotic preparations combining sequentially standard techniques and multiplex fluorescence in situ hybridization. Setting: Assisted reproduction centers. Patient(s): Thirty-seven men consulting for fertility problems. Intervention(s): Unilateral testicular biopsies. Main Outcome Measure(s): Proximity frequencies analysis to the XY pair, evaluated individually and grouping bivalents, was carried out using a logistical regression model with repeated measures. Result(s): Bivalents 22 and 15 were observed more frequently near to the sex bivalent than the others. Significant interindividual differences were not observed. Conclusion(s): Results suggest that bivalents distribution to the metaphase plate is nonrandom. The maintenance of the acrocentric chromosomes’ proximity to the sex bivalent from pachytene to metaphase I would indicate that the relative bivalents position would be notably preserved. The observation of non-interindividual variability, despite different infertility etiology, suggests that the nuclear organization pattern remains largely unaffected even if spermatogenesis is compromised. [ABSTRACT FROM AUTHOR]

A prospective study was performed to assess the impact of sperm DNA fragmentation on the outcome of IVF with own or donated oocytes. The study population included 178 couples (62 cycles of IVF, 116 of intracytoplasmic sperm injection (ICSI)) with own (n = 77) and donor (n = 101) oocytes. DNA fragmentation was evaluated by TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling assay. Correlation between DNA damage to oocyte fertilization, embryo quality and clinical pregnancy, implantation and miscarriage rates was evaluated. DNA fragmentation was not related to fertilization rates in either IVF (r = 0.08) or ICSI (r = 0.04) cycles. DNA fragmentation was similar in patients with <50% embryo utilization rate compared with ≥-50%, in cancelled and in embryo transfer cycles and in miscarriages and in successful deliveries. Moreover, DNA fragmentation was similar in pregnant and non-pregnant women as well as in IVF with own or donor oocytes. In the multivariabte analysis, the odds ratio of DNA after controlling by age was 1.0. Using a 36% sperm fragmentation threshold, results did not vary. It is concluded that DNA damage was not related to outcomes of IVF or ICSI with own or donor oocytes. [ABSTRACT FROM AUTHOR]

Genomic disorders are human diseases caused by meiotic chromosomal rearrangements of unstable regions flanked by Low Copy Repeats (LCRs). LCRs act as substrates for Non-Allelic Homologous Recombination (NAHR) leading to deletions and duplications. The aim of this study was to assess the basal frequency of deletions and duplications of the 7q11.23, 15q11-q13 and 22q11.2 regions in spermatozoa from control donors to check differences in the susceptibility to generate anomalies and to assess the contribution of intra- and inter-chromatid NAHR events. Semen samples from ten control donors were processed by FISH. A customized combination of probes was used to discriminate among normal, deleted and duplicated sperm genotypes. A minimum of 10,000 sperm were assessed per sample and region. There were no differences in the mean frequency of deletions and duplications (del + dup) among the 7q11.23, 15q11-q13 and 22q11.2 regions (frequency ± SEM, 0.37 ± 0.02; 0.46 ± 0.07 and 0.27 ± 0.07%, respectively) ( P = 0.122). Nevertheless, hierarchical cluster analysis reveals interindividual differences suggesting that particular haplotypes could be the main source of variability in NAHR rates. The mean frequency of deletions was not different from the mean frequency of duplications in the 7q11.23 ( P = 0.202) and 15q11-q13 ( P = 0.609) regions, indicating a predominant inter-chromatid NAHR. By contrast, in the 22q11.2 region the frequency of deletions slightly exceed duplications ( P = 0.032), although at the individual level any donor showed differences. Altogether, our results support the inter-chromatid NAHR as the predominant mechanism involved in the generation of sperm deletions and duplications. [ABSTRACT FROM AUTHOR]

Robertsonian translocations are one of the most frequent reorganizations in humans. Their segregational behavior and their implication in the occurrence of interchromosomal effects (ICEs) has been an important topic of research for the past 10 years. Most of the cases analyzed correspond to rearrangements with chromosomes from the D-group (chromosomes 13, 14 and 15), whereas some rare Robertsonian translocations are scarcely found in the literature, mainly those with both chromosomes from the G-group (chromosomes 21 and 22) and those involving chromosomes from both groups (D;G translocations). Results supporting/rejecting the existence of the ICE phenomenon have been reported, showing the need for more studies to characterize its distribution. In this study, sperm fluorescent in situ hybridization studies have been performed in three D;G Robertsonian translocation carriers: two men with the translocation t(14;21) and a third individual with the rare t(13;22) reorganization. Segregation and ICE results have been considered in relation to their cytogenetic features and all previously published data. The compiled information discards a particular segregation behavior related to the chromosomes involved. In contrast with this segregational homogeneity, heterogeneous ICE results were observed, indicating a significant but random distribution of such phenomenon. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the reliability and applicability of the spot-counting system (Cytovision Spot AX workstation) which offers an alternative to the tedious manual analysis of sperm fluorescence in situ hybridization (FISH).Design: Manual and automatic analyses were performed and compared.Setting: Universitat Autònoma de Barcelona.Patient(s): Twenty-four men who asked for information on infertility showing different seminal parameters.Intervention(s): A semen sample for each patient was collected and prepared for FISH.Main Outcome Measure(s): A dual-color FISH using specific probes for chromosomes 13 and 21 and a triple-color FISH with centromeric probes for chromosomes 18, X, and Y were used (Vysis). Standard FISH analysis was carried out. Automatic analysis was subsequently performed using a Spot AX system.Result(s): Overall, we performed 120 comparisons. In 116 out of 120 (96.67%), the percentage of anomalies reported using manual counting fell within the incidence detected using the automatic system. In the remaining comparisons, statistical differences were detected (4 out of 120; 3.33%). Time consumed by the automatic analysis was always higher than the manual one, being influenced by the characteristics of the preparations.Conclusion(s): The spot-counting system has potential, but before the service is ready to be offered, we still need to overcome some limitations associated with it. [ABSTRACT FROM AUTHOR]

Are intrinsic or extrinsic factors associated with embryo mosaicism prevalence in IVF cycles? Retrospective cohort study of preimplantation genetic testing for aneuploidy (PGT-A) cycles carried out at a university-affiliated IVF clinic between October 2017 and October 2019. Trophectoderm biopsies were analysed by next generation sequencing. Mosaicism prevalence, type of anomaly and the chromosomes involved were analysed. Intrinsic and extrinsic factors potentially inducing mosaicism were studied: maternal and paternal age, antral follicle count, cumulus–oocyte complexes retrieved, female body mass index, PGT-A indication, sperm concentration, total dosage of gonadotrophins, embryo quality and day of blastocyst formation, single-step commercial media used and biopsy operator. Overall prevalence of mosaicism in our PGT-A setting was 13.9%. In segmental mosaicism, larger chromosomes tended to be more affected, which was not observed in whole-chromosome mosaicism. Additionally, segmental mosaicism was mostly observed in monosomy (69.6%; P < 0.01) compared with whole-chromosome mosaicism (49.7% monosomies versus 50.3% trisomies; P = 0.83). Although a high inter-patient variability was observed, only paternal age showed a positive association with mosaicism (adjusted OR 1.26, 95% CI 1.02 to 1.54) among the analysed variables. Our results suggest remarkable differences in the mechanisms generating segmental and whole-chromosome mosaicism, indicating that they may deserve different consideration when studying them and when prioritizing them for transfer. Male factor seems to be associated with mosaicism and may be worthy of specific assessment in future studies. [ABSTRACT FROM AUTHOR]

Objective: To identify candidates of fertility biomarkers among pairs of human sperm microRNAs.Design: Expression data of 736 sperm microRNAs from fertile and infertile individuals characterized in previous published studies by means of TaqMan quantitative polymerase chain reaction (PCR) were reexamined. A set of microRNA pairs with the best biomarker potential were selected and validated by means of quantitative real-time (qRT) PCR in an independent cohort.Setting: University laboratory.Patient(s): Semen samples were obtained from fertile (n = 10) and infertile (asthenozoospermia, n = 10; teratozoospermia, n = 10; oligozoospermia, n = 10; unexplained male infertility [UMI], n = 8) individuals. The validation cohort included 9 fertile donors and 14 infertile patients with different seminal alterations.Intervention(s): None.Main Outcome Measure(s): Spearman test was used to select microRNA pairs with a correlated expression in fertile individuals and a noncorrelated expression in each infertile group. The biomarker potential of these pairs was determined with the use of receiver operating characteristic curves. The differential relative expression of each pair in fertile and infertile populations was verified (Mann-Whitney test). Those pairs with best results were validated by qRT-PCR.Result(s): Forty-eight pairs showed significant correlations in the fertile group. The pairs that were uncorrelated in the infertile populations and displayed the best biomarker potential were hsa-miR-942-5p/hsa-miR-1208 (asthenozoospermia), hsa-miR-296-5p/hsa-miR-328-3p (teratozoospermia), hsa-miR-139-5p/hsa-miR-1260a (oligozoospermia), and hsa-miR-34b-3p/hsa-miR-93-3p (UMI). The hsa-miR-942-5p/hsa-miR-1208 pair showed the greatest potential for detecting seminal alterations in the validation cohort (85.71% true positives).Conclusion(s): The pairs hsa-miR-942-5p/hsa-miR-1208 and hsa-miR-34b-3p/hsa-miR-93-3p have the potential to become new molecular biomarkers that could help to diagnose male infertility, especially in cases of UMI or when seminal parameters are close to the threshold values. [ABSTRACT FROM AUTHOR]

Purpose The study aims to determine whether there is an altered bivalent positioning in metaphase I human spermatocytes from Robertsonian translocation carriers. Methods Metaphase I human spermatocytes from three 45,XY, d e r(13;14)(q10;q10) individuals and a 45,XY,der(14;15)(q10;q10) individual were analyzed. Proximity relationships of bivalents were established by analyzing meiotic preparations combining Leishman staining and multiplex-FISH procedures. Poisson regression model was used to determine proximity frequencies between bivalents and to assess associations with chromosome size, gene density, acrocentric morphology, and chromosomes with heterochromatic blocks. The hierarchical cluster Ward method was used to characterize the groups of bivalents with preferred proximities in a cluster analysis. Bivalent groups obtained were individually compared with those obtained in normal karyotype individuals evaluated in a previous study. Results A total of 1288 bivalents were examined, giving a total of 2289 proximity data. Only four positive significant proximities were detected for each type of Robertsonian translocation. Significant bivalent associations were only observed by small-size chromosomes for MI,22,XY,III(13q14q). These results were clearly divergent from 46,XY individuals. Moreover, cluster analysis revealed that about 30 % of the bivalents showed changes in their proximity relationships in metaphase I. Conclusions The territorial organization of bivalents in metaphase I human spermatocytes changes in the presence of a Robertsonian translocation. [ABSTRACT FROM AUTHOR]

Objective: To characterize the microRNA (miRNA) expression profile in spermatozoa from human fertile individuals and their implications in human fertility. Design: The expression levels of 736 miRNAs were evaluated using TaqMan arrays. Ontologic analyses were performed to determine the presence of enriched biological processes among their targets. Setting: University research and clinical institutes. Patient(s): Ten individuals with normal seminogram, standard karyotype, and proven fertility. Intervention(s): None. Main Outcome Measure(s): Expression levels of 736 miRNAs, presence of enriched metabolic routes among their targets, homogeneity of the population, influence of demographic features in the results, presence of miRNA stable pairs, and best miRNA normalizing candidates. Result(s): A total of 221 miRNAs were consistently present in all individuals, 452 were only detected in some individuals, and 63 did not appear in any sample. The ontologic analysis of the 2,356 potential targets of the ubiquitous miRNAs showed an enrichment of processes related to cell differentiation, development, morphogenesis, and embryogenesis. None of the miRNAs were significantly correlated with age, semen volume, sperm concentration, motility, or morphology. Correlations between samples were statistically significant, indicating a high homogeneity of the population. A set of 48 miRNA pairs displayed a stable expression, a particular behavior that is discussed in relationship to their usefulness as fertility biomarkers. Hsa-miR-532-5p, hsa-miR-374b-5p, and hsa-miR-564 seemed to be the best normalizing miRNA candidates. Conclusion(s): Human sperm contain a stable population of miRNAs potentially related to embryogenesis and spermatogenesis. [ABSTRACT FROM AUTHOR]

To characterize meiotic anomalies in infertile men by multiplex fluorescence in situ hybridization (M-FISH) and to determine whether synaptic problems affect specific bivalents or whether anomalies are random.Analysis of meiotic preparations with standard techniques and M-FISH.Assisted reproduction centers and Universitat Autònoma de Barcelona.Three fertile men undergoing vasectomy, four sterile patients with oligoasthenoteratozoospermia, and one patient with a Robertsonian translocation t(13;14).Unilateral testicular biopsy in controls and patients with oligoasthenoteratozoospermia and collection of a semen sample from the translocation carrier.Identification of bivalents in metaphase I and chromosomes in metaphase II and characterization of chromosome abnormalities.All bivalents in metaphase I and all chromosomes in metaphase II could be identified. In controls and in one patient with oligoasthenoteratozoospermia, meiosis was normal. Other patients with oligoasthenoteratozoospermia showed different types of anomaly: desynapsis, breaks, precocious XY separation, or cryptic reorganizations. The Robertsonian translocation t(13;14) was easily identified.Results confirm the high incidence of synaptic errors in oligoasthenoteratozoospermia patients. Bivalents in metaphase I and chromosomes in metaphase II were individually identifiable. Nondisjunctional errors or small reorganizations overlooked in classic meiotic preparations were identified. Synaptic anomalies seem to affect meiotic bivalents at random. [Copyright &y& Elsevier]

: ObjectiveTo determine the meiotic behavior of a ring Y chromosome in a semen sample from a 45,X/46,X,r(Y)/46,X,dic r(Y) patient and the possible interchromosomal effects of the ring on other chromosome pairs.: DesignRetrospective analysis.: SettingUniversitat Auto&grave;noma de Barcelona.: PatientAn oligoasthenoteratozoospermic patient who presented for infertility consultation.: Main outcome measure(s)The sex chromosome content of spermatogenic cells, meiotic figures, and spermatozoa in the ejaculate and the possible interchromosomal effects on chromosomes 13, 18, and 21 were analyzed by using multicolor fluorescence in situ hybridization. Germ-cell aneuploidies were scored.: Result(s)X0 cells are meiotically incompetent. All meiotic figures were exclusively XY, and 80% showed unpaired sex chromosomes. A high proportion of postreductional cells were XY (45.5%) or nullisomic for sex chromosomes (13.92%). This percentage decreased in spermatozoa to 14.89% and 27.66%, respectively. A statistically significant increase in X-bearing versus Y-bearing cells both in postreductional cells (23.9% vs. 14.3%) and spermatozoa (41.9% vs. 19.3%) was also observed. Evidence for an interchromosomal effect on chromosome 21 was detected.: Conclusion(s)Data suggest that this patient had a generalized increase incidence of chromosome anomalies, underscoring the importance of incorporating screening for sperm aneuploidies in genetic analysis of affected patients. [Copyright &y& Elsevier]

Objective: To determine whether there is a preferential bivalent distribution pattern in metaphase I human spermatocytes and to analyze whether this positioning is influenced by chiasmata count, chromosome size, gene density, acrocentric morphology, and heterochromatic blocks. Design: Proximity frequencies of bivalents were evaluated with the analysis of meiotic preparations combining sequentially standard techniques and multiplex fluorescence in situ hybridization. Setting: University. Patient(s): Twenty-five men consulting for fertility problems. Intervention(s): Unilateral testicular biopsies. Main Outcome Measure(s): Proximity analyses were performed for each bivalent considering as nearby bivalents those that were part of the first ring around the bivalent studied. Data were analyzed using Poisson regression models, multidimensional scaling, and cluster analysis. Result(s): Some bivalents have a preferential relative position. Significant associations among bivalents related to chromosome size, high gene density, and acrocentric morphology were observed. Chiasmata count and heterochromatic blocks were nonconditioning parameters of the bivalent organization. Conclusion(s): This study demonstrates that distribution in metaphase I is nonrandom and influenced by chromosome size, gene density, and acrocentric chromosome morphology. Results support that some features defining chromosome territories are maintained during meiosis. [ABSTRACT FROM AUTHOR]