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To assess patients' and embryonic characteristics that may have an influence on the decision to transfer a mosaic embryo.Single centre retrospective cohort study including 1247 PGT-A cycles. Demographic and clinical factors associated with a decision to transfer a mosaic embryo were studied. Female age, number of previous cycles, previous availability of euploid embryos, history of miscarriages and parity as well as percentage of mosaicism, type of anomaly and chromosome risk were studied in relation to decision-making. Outcomes after mosaic embryo transfer were assessed.To date, in 7.9% of cycles (99/1247), patients have had to make a decision on the fate of their mosaic embryos. In 23.2% of cycles (23/99), patients decided to transfer. In most cases (79.8%; 79/99), patients underwent genetic counselling before the decision. None of the variables analysed were associated with the patients' decision, although parity and the high-degree mosaicism (50%) seemed to be negatively associated with the decision to transfer (18.2% vs. 29.8%, p = 0.294; 10% vs. 32.2%, p = 0.052).Neither reproductive history nor information on mosaic embryo characteristics through counselling seems to be determinative for patients when deciding to transfer a mosaic embryo. Promising and increasing data on clinical outcomes after mosaic embryo transfer will be of utmost importance to soften risk perception regarding mosaic embryos and give a better, simplified and more evidence-based counselling.

Chromosome territoriality is not random along the cell cycle and it is mainly governed by intrinsic chromosome factors and gene expression patterns. Conversely, very few studies have explored the factors that determine chromosome territoriality and its influencing factors during meiosis. In this study, we analysed chromosome positioning in murine spermatogenic cells using three-dimensionally fluorescence in situ hybridization-based methodology, which allows the analysis of the entire karyotype. The main objective of the study was to decipher chromosome positioning in a radial axis (all analysed germ-cell nuclei) and longitudinal axis (only spermatozoa) and to identify the chromosomal factors that regulate such an arrangement. Results demonstrated that the radial positioning of chromosomes during spermatogenesis was cell-type specific and influenced by chromosomal factors associated to gene activity. Chromosomes with specific features that enhance transcription (high GC content, high gene density and high numbers of predicted expressed genes) were preferentially observed in the inner part of the nucleus in virtually all cell types. Moreover, the position of the sex chromosomes was influenced by their transcriptional status, from the periphery of the nucleus when its activity was repressed (pachytene) to a more internal position when it is partially activated (spermatid). At pachytene, chromosome positioning was also influenced by chromosome size due to the bouquet formation. Longitudinal chromosome positioning in the sperm nucleus was not random either, suggesting the importance of ordered longitudinal positioning for the release and activation of the paternal genome after fertilisation.

“Apenas um ateu pode ser um bom cristão [...]”, declara o filósofo Ernst Bloch em seu livro Ateísmo no cristianismo. Como ele chega a uma tal tese? Que representação de messianismo marca a sua filosofia da esperança? O texto mostra como Bloch quer liberar as motivações rebeldes do religioso a fim de que possam ser explicitados os seus conteúdos de esperança. Também deixa claro por que o autor presume que o ateísmo já está ativo no próprio cristianismo.

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%; P0.01) 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.

SummaryThe aim of this study was to provide a more comprehensive understanding of 1PN intracytoplasmic sperm injection (ICSI) zygotes. To achieve this objective, we assessed whether all 1PN-derived embryos showed a similar morphokinetic pattern, and if the morphokinetic behaviour of 1PN-derived embryos was comparable with that of 2PN-derived embryos. In total, 149 1PN ICSI zygotes (study group) and 195 2PN ICSI zygotes (control group) were included in the study. Embryo development potential was evaluated in terms of blastocyst rate. Morphokinetic parameters, including the pronucleus diameter and kinetics of in vitro development, were also analyzed. Embryos derived from 1PN ICSI zygotes showed impaired development compared with 2PN-derived embryos, with blastocyst rates of 28.9% and 67.2%, respectively. The diameter of the pronucleus of 1PN zygotes was larger than that of 2PN zygotes. When compared with 2PN-derived embryos, those derived from 1PN zygotes had a visible pronucleus for a shorter time, in addition to a longer syngamy time and slower kinetic behaviour from two to nine cells. When 1PN-derived blastocysts and 2PN-derived blastocysts were compared, the developmental kinetics were similar in both groups, except for a delayed and longer duration of the compaction phase in 1PN-derived embryos. In conclusion, monopronucleated ICSI zygotes present differences in developmental capacity and morphokinetic behaviour compared with 2PN ICSI zygotes, showing particular morphokinetic parameters related to pronucleus formation. Only the 1PN ICSI-derived embryos that reached the blastocyst stage have similar morphokinetic development to blastocysts from 2PN zygotes.

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. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10815-022-02453-9.

In a world in which the strategic use of disinformation is becoming more and more the rule, it is necessary to ask how the credibility of information can be measured. The text gives indications as to why rhetorical skills are necessary for this and what the criteria for a credible style of speech are.

Transcriptome analyses using high-throughput methodologies allow a deeper understanding of biological functions in different cell types/tissues. The present study provides an RNA-seq profiling of human sperm mRNAs and lncRNAs (messenger and long non-coding RNAs) in a well-characterized population of fertile individuals. Sperm RNA was extracted from twelve ejaculate samples under strict quality controls. Poly(A)-transcripts were sequenced and aligned to the human genome. mRNAs and lncRNAs were classified according to their mean expression values (FPKM: Fragments Per Kilobase of transcript per Million mapped reads) and integrity. Gene Ontology analysis of the Expressed and Highly Expressed mRNAs showed an involvement in diverse reproduction processes, while the Ubiquitously Expressed and Highly Stable mRNAs were mainly involved in spermatogenesis. Transcription factor enrichment analyses revealed that the Highly Expressed and Ubiquitously Expressed sperm mRNAs were primarily regulated by zinc-fingers and spermatogenesis-related proteins. Regarding the Expressed lncRNAs, only one-third of their potential targets corresponded to Expressed mRNAs and were enriched in cell-cycle regulation processes. The remaining two-thirds were absent in sperm and were enriched in embryogenesis-related processes. A significant amount of post-testicular sperm mRNAs and lncRNAs was also detected. Even though our study is solely directed to the poly-A fraction of sperm transcripts, results indicate that both sperm mRNAs and lncRNAs constitute a footprint of previous spermatogenesis events and are configured to affect the first stages of embryo development.

The aim of this study was to provide a more comprehensive understanding of 1PN intracytoplasmic sperm injection (ICSI) zygotes. To achieve this objective, we assessed whether all 1PN-derived embryos showed a similar morphokinetic pattern, and if the morphokinetic behaviour of 1PN-derived embryos was comparable with that of 2PN-derived embryos. In total, 149 1PN ICSI zygotes (study group) and 195 2PN ICSI zygotes (control group) were included in the study. Embryo development potential was evaluated in terms of blastocyst rate. Morphokinetic parameters, including the pronucleus diameter and kinetics of in vitro development, were also analyzed. Embryos derived from 1PN ICSI zygotes showed impaired development compared with 2PN-derived embryos, with blastocyst rates of 28.9% and 67.2%, respectively. The diameter of the pronucleus of 1PN zygotes was larger than that of 2PN zygotes. When compared with 2PN-derived embryos, those derived from 1PN zygotes had a visible pronucleus for a shorter time, in addition to a longer syngamy time and slower kinetic behaviour from two to nine cells. When 1PN-derived blastocysts and 2PN-derived blastocysts were compared, the developmental kinetics were similar in both groups, except for a delayed and longer duration of the compaction phase in 1PN-derived embryos. In conclusion, monopronucleated ICSI zygotes present differences in developmental capacity and morphokinetic behaviour compared with 2PN ICSI zygotes, showing particular morphokinetic parameters related to pronucleus formation. Only the 1PN ICSI-derived embryos that reached the blastocyst stage have similar morphokinetic development to blastocysts from 2PN zygotes.

SummaryShortly after the implementation of comprehensive chromosome screening (CCS) techniques for preimplantation genetic testing for aneuploidies (PGT-A), the discussion about the transition from day 3 to blastocyst stage biopsy was initiated. Trophectoderm biopsy with CCS is meant to overcome the limitations of cleavage-stage biopsy and single-cell analysis. The aim of this study was to assess the results obtained in our PGT-A programme after the implementation of this new strategy. Comparisons between the results obtained in 179 PGT-A cycles with day 3 biopsy (D+3) and fresh embryo transfer, and 204 cycles with trophectoderm biopsy and deferred (frozen–thawed) embryo transfer were established. Fewer embryos were biopsied and a higher euploidy rate was observed in the trophectoderm biopsy group. No differences in implantation (50.3% vs. 61.4%) and clinical pregnancy rate per transfer (56.1% vs. 65.3%) were found. Although the mean number of euploid embryos per cycle did not differ between groups (1.5 ± 1.7 vs. 1.7 ± 1.8), the final number of euploid blastocysts available for transfer per cycle was significantly higher in the trophectoderm biopsy group (1.1 ± 1.3 vs. 1.7 ± 1.8). This factor led to an increased cumulative live birth rate in this last group (34.1% vs. 44.6%). Although both strategies can offer good results, trophectoderm biopsy offers a more robust diagnosis and the intervention is less harmful for the embryos so more euploid blastocysts are finally available for transfer and/or vitrification.

Colloidal super-paramagnetic microbeads conjugated with annexin V are effective for separating apoptotic spermatozoa by MACS as a result of the high affinity of annexin V for externalized PS molecules. The effectiveness of the procedure in reducing the percentage of sperm with fragmented DNA and abnormal morphology has also been reported. However, it is still unknown if it could decrease the percentage of aneuploid spermatozoa. The objective of our prospective study, performed on 16 males with abnormal FISH on spermatozoa, was to assess if MACS columns were useful tools to retain spermatozoa carrying chromosomal abnormalities in semen samples processed after density gradient centrifugation (DGC). The pellet obtained after DGC was subjected to MACS, and sperm FISH analyses were performed both in the eluded fraction and in the fraction retained in the column. The observed frequencies of disomy and nullisomy 13, 18, and 21, X and Y, as well as the diploidy rates in the MACS eluded fraction and the fraction retained in the MACS column were recorded. We observed that the frequencies of aneuploidies in the eluded fraction were lower than in the fraction retained in the MACS column (0.59% vs. 0.75%; p = 0.010). DGC determined a significant reduction in sperm concentration (z-ratio = 2.83; p = 0.005) and a significant increase in sperm progressive motility (z-ratio = -3.5; p

Resumen Introduccion La multinucleacion es un fenomeno frecuente en los embriones obtenidos por fecundacion in vitro y suele asociarse a un pobre pronostico reproductivo. La monitorizacion embrionaria en sistemas de cultivo time-lapse permite la observacion detallada y continua del desarrollo embrionario. En nuestro trabajo se ha evaluado el impacto de la presencia de multinucleacion en la morfocinetica, la capacidad de desarrollo y la implantacion embrionaria. Material y metodos Estudio retrospectivo de cohorte. Se han incluido 5.540 embriones cultivados en TL (Embryoscope®, Unisense Fertilitech®) procedentes de 763 ciclos de FIV-ICSI realizados durante el periodo 2012-2014. La presencia de multinucleacion se evaluo en todas las etapas del desarrollo. La transferencia embrionaria se realizo en D + 3, y los embriones multinucleados se cultivaron hasta alcanzar el estadio de blastocisto. Resultados Un 23,6% de los embriones mostraron multinucleacion en algun momento del desarrollo embrionario. Los valores de los parametros morfocineticos de los embriones multinucleados y no multinucleados mostraron diferencias estadisticamente significativas. El 18% de los embriones multinucleados alcanzaron el estadio de blastocisto y fueron congelados. No se observo efecto ni del tipo de multinucleacion ni del porcentaje de celulas multinucleadas sobre la tasa de blastocisto. La tasa de implantacion de los blastocistos procedentes de embriones multinucleados fue del 39,1%. Discusion La monitorizacion embrionaria continua permite una mejor deteccion de la multinucleacion. La presencia de multinucleacion embrionaria se asocia a una morfocinetica alterada y una capacidad de desarrollo reducida. No obstante, los embriones multinucleados que alcanzan el estadio de blastocisto muestran una elevada capacidad de implantacion.

Resumen Los hallazgos acerca de la relevancia biologica de los espermatozoides, mas alla de la mera aportacion del genoma paterno al embrion, han abierto las puertas al estudio del RNA espermatico bajo nuevos enfoques. Lejos de constituir una mera reserva residual, esta fraccion ha demostrado ser una poblacion compleja y multifuncional, con una amplia gama de implicaciones en procesos biologicos tanto espermatogenicos como de desarrollo embrionario tras la fecundacion del ovocito. Mas alla de la poblacion de RNA codificante, el papel ejercido por los RNA no codificantes comienza a desentramarse e integrarse con el funcionamiento celular global, demostrando su relevancia sobre la regulacion celular e incluso su influencia epigenetica. Los estudios comparativos de expresion a gran escala permiten obtener informacion global sobre los cambios en el transcriptoma espermatico en diferentes poblaciones, lo que aporta al RNA un nuevo valor diagnostico. En esta revision se ofrece una vision general sobre la biosintesis, las funciones y los mecanismos biologicos relacionados con los diferentes subtipos de RNA espermaticos, asi como sobre su importancia durante la espermatogenesis, su implicacion tras la fecundacion y su valor como biomarcadores de fertilidad.

STUDY QUESTION What has the ESHRE programme ‘ESHRE Certification for Clinical Embryologists’ achieved after 10 years? SUMMARY ANSWER The post-exam analysis showed a pass rate of 60% for Clinical and 50% for Senior Clinical Embryologists and a high level of internal consistency of all exams, leading to a total of 773 certified Clinical and 493 Senior Clinical Embryologists over the decade. WHAT IS KNOWN ALREADY In an ESHRE survey on the educational and professional status of Clinical Embryology in Europe, it was found that education of laboratory personnel working in the field of assisted reproduction is highly variable between countries. In 2008, ESHRE introduced a programme, curriculum and certification in the field of Clinical Embryology. Knowledge gained by postgraduate study of recommended literature, following a clear curriculum, is verified by a written two-level exam for obtaining a certificate for Clinical (basic) or Senior Clinical (advanced) Embryologists. With a total of 1266 certificates awarded over a period of 10 years and recognition by the Union Européenne des Médecins Spécialistes and their Council for European Specialists Medical Assessment, the ESHRE Clinical Embryology exams have become an internationally recognized educational standard in the field of Clinical Embryology. STUDY DESIGN, SIZE, DURATION A retrospective analysis of all applications for ESHRE Clinical (2009–2018) and Senior Clinical Embryologist Certification (2008–2018) and exam results of the first decade was carried out by the Steering Committee for Clinical Embryologist Certification. PARTICIPANTS/MATERIALS, SETTING, METHODS A total of 2894 applications for ESHRE Certification for Clinical Embryologists and the results of 10 exams for the Clinical (1478 candidates) and 11 exams for Senior Clinical (987 candidates) levels were analysed. A detailed post-exam retrospective analysis was performed regarding difficulty, discrimination and reliability levels of 1600 multiple-choice questions (MCQs) with a single best answer among four options, from eight different curriculum topics (Basic cell biology, Genetics, Developmental biology, Female reproduction, Male reproduction, IVF laboratory, Cryopreservation and Laboratory management), representing the core theoretical knowledge of Clinical Embryology. Difficulty levels of the MCQs were subsequently compared regarding each topic and each yearly exam. The participation and success rates in the ESHRE Clinical Embryology exams were also assessed in terms of the educational and geographic backgrounds of candidates. MAIN RESULTS AND THE ROLE OF CHANCE Over the 10 years studied, the mean pass rate for the Clinical Embryologist exam was 60% (range 41–86%), and for the Senior Clinical Embryologist exam was 50% (range 34–81%). On average, 63% European candidates and 35% non-European candidates passed the Clinical Embryologist exam, while 52% European candidates and 31% non-European candidates passed the Senior Clinical Embryologist exam. The candidates’ educational level impacted on the success of the Clinical Embryologist exam but not of the Senior Clinical Embryologist exam. The mean difficulty indices by study topic showed that in the period of 10 years, there were no statistically significant differences between topics, for either the Clinical or Senior Clinical Embryologist exams. However, the overall exam difficulty varied between years. Reassuringly, the exam MCQ discrimination and reliability indices always showed a high level of internal consistency in all exams. LIMITATIONS, REASONS FOR CAUTION Some data from the initial ESHRE certification programme were not obtained electronically, in particular data for education, implying tables and figures reflect the specified valid data periods. Several countries exhibit different study profiles for those working in ART laboratories, such that laboratory technicians/technologists predominate in some countries, while in others only biologists and medical doctors are allowed to work with human embryos. Such differences could consequently affect the exam performance of candidates from specific countries. WIDER IMPLICATIONS OF THE FINDINGS The ESHRE exams on Clinical Embryology are the most widely, internationally accepted tests of knowledge in the rapidly growing area of human reproduction. Clinical Embryology is increasingly recognized as a specific discipline for scientific staff who are collaborating closely with clinicians in managing human infertility through medically assisted reproduction. The analysis of the first 10 years of application of a two-level exam for Clinical Embryology shows a consistent high quality and reliability of the exam and MCQs used. These results represent an important follow-up of the quality of the ESHRE Certification programme for Clinical Embryologists, and convincingly position Clinical Embryology in the wider group of health disciplines that are harmonized through professional bodies such as ESHRE and European Board & College of Obstetrics and Gynaecology. The exams provide a clear step towards the increasing professional recognition and establishment of Clinical Embryology within health systems at both European and international level. STUDY FUNDING/COMPETING INTEREST(S) No competing interest. All costs of the Steering Committee meetings were covered by ESHRE.

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.

Study question What is the most reliable normalization strategy for sperm microRNA (miRNA) quantitative Reverse Transcription Polymerase Chain Reactions (qRT-PCR) using singleplex assays? Summary answer The use of the average expression of hsa-miR-100-5p and hsa-miR-30a-5p as sperm miRNA qRT-PCR data normalizer is suggested as an optimal strategy. What is known already Mean-centering methods are the most reliable normalization strategies for miRNA high-throughput expression analyses. Nevertheless, specific trustworthy reference controls must be established in singleplex sperm miRNA qRT-PCRs. Study design, size duration Cycle threshold (Ct) values from previously published sperm miRNA expression profiles were normalized using four approaches: (i) Mean-Centering Restricted (MCR) method (taken as the reference strategy); (ii) expression of the small nuclear RNA RNU6B; (iii) expression of four miRNAs selected by the Concordance Correlation Restricted (CCR) algorithm: hsa-miR-100-5p, hsa-miR-146b-5p, hsa-miR-92a-3p and hsa-miR-30a-5p; (iv) the combination of two of these miRNAs that achieved the highest proximity to MCR. Participants/materials, setting, methods Expression profile data from 736 sperm miRNAs were taken from previously published studies performed in fertile donors (n = 10) and infertile patients (n = 38). For each tested normalizer molecule, expression ubiquity and uniformity across the different samples and populations were assessed as indispensable requirements for being considered as valid candidates. The reliability of the different normalizing strategies was compared to MCR based on the set of differentially expressed miRNAs (DE-miRNAs) detected between populations, the corresponding predicted targets and the associated enriched biological processes. Main results and the role of chance All tested normalizers were found to be ubiquitous and non-differentially expressed between populations. RNU6B was the least uniformly expressed candidate across samples. Data normalization through RNU6B led to dramatically misguided results when compared to MCR outputs, with a null prediction of target genes and enriched biological processes. Hsa-miR-146b-5p and hsa-miR-92a-3p were more uniformly expressed than RNU6B, but their results still showed scant proximity to the reference method. The highest resemblance to MCR was achieved by hsa-miR-100-5p and hsa-miR-30a-5p. Normalization against the combination of both miRNAs reached the best proximity rank regarding the detected DE-miRNAs (Area Under the Curve = 0.8). This combination also exhibited the best performance in terms of the target genes predicted (72.3% of True Positives) and their corresponding enriched biological processes (70.4% of True Positives). Large scale data Not applicable. Limitations, reasons for caution This study is focused on sperm miRNA qRT-PCR analysis. The use of the selected normalizers in other cell types or tissues would still require confirmation. Wider implications of the findings The search for new fertility biomarkers based on sperm miRNA expression using high-throughput assays is one of the upcoming challenges in the field of reproductive genetics. In this context, validation of the results using singleplex assays would be mandatory. The normalizer strategy suggested in this study would provide a universal option in this area, allowing for normalization of the validated data without causing meaningful variations of the results. Instead, qRT-PCR data normalization by RNU6B should be discarded in sperm-miRNA expression studies. Study funding/competing interests This work was supported by the 2014/SGR00524 project (Agencia de Gestio d'Ajuts Universitaris i de Recerca, Generalitat de Catalunya, Spain) and UAB CF-180034 grant (Universitat Autonoma de Barcelona). Celia Corral-Vazquez is a recipient of a Personal Investigador en Formacio grant UAB/PIF2015 (Universitat Autonoma de Barcelona). The authors report no conflict of interest.

The study aims to determine whether there is an altered bivalent positioning in metaphase I human spermatocytes from Robertsonian translocation carriers.Metaphase I human spermatocytes from three 45,XY,der(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.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.The territorial organization of bivalents in metaphase I human spermatocytes changes in the presence of a Robertsonian translocation.

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.

Altres ajuts: Universitat Autònoma de Barcelona (CF-180034)

Introduction The development of new methodologies for PGT-A has led to the detection of a certain percentage of mosaic embryos. It has been postulated that this could be a normal feature in human embryos. To date, the real incidence of mosaicism in blastocysts is unknown as illustrated by the very variable incidences reported between groups. Certain factors could influence mosaicism occurrence such as patients’ characteristics, ovarian stimulation and in vitro culture conditions. Biopsy sample manipulation could also induce artefactual mosaicism. Moreover, different diagnostic platforms, algorithms and detection limits are used by different groups, thus making comparisons difficult. Material & methods The cytogenetic results of 1339 blastocysts obtained from PGT-A cycles performed in our centre from September 2017 to January 2019 were retrospectively analysed. Embryos were biopsied at the blastocyst stage between day 5 and 7 of development and genetic analysis was performed by next-generation sequencing (NGS) according to VeriSeq™-PGS protocols (Illumina). Embryos were diagnosed as euploid, aneuploid or mosaic. An embryo was classified as mosaic when the percentage of affected cells ranged from 30-70%. Both the incidence of mosaicism in this cohort and the specific chromosomes affected were studied. Mosaicism was studied versus patients’ factors (age, indication for PGT-A, ovarian reserve), embryonic factors (blastocyst development and quality), potential iatrogenic factors (ovarian stimulation treatment and the use of two different single-step culture media), and the biopsy operator as a proposed source of artefactual mosaicism. A logistic mixed multivariable model was applied to estimate the odds for every endpoint. Patient and biopsy procedure were treated as random factor to control the correlated observations effect. Results The overall incidence of mosaic embryos was 13.0% (7.5% of euploid-aneuploid mosaic embryos and 5.5% of aneuploid-aneuploid mosaic embryos). Most mosaic embryos showed just one chromosome affected (85.5%). No chromosomes were especially associated with mosaic aneuploidy. However, segmental mosaicism was more common in larger chromosomes. No association could be stablished between embryo mosaicism and the factors studied. Neither maternal nor paternal ages showed any association with mosaicism (ORa=0.96(0.88-1.04); ORa=1.04(0.99-1.09)). The antral follicle count did not show any correlation either (ORa=0.987(0.95-1.02)) nor the indication for PGT-A. In terms of embryo development, no associations were found either: ORa for blastocyst formation in day 5 vs day 7 was 0.72(0.31-1.68); ORa for blastocyst formation in day 6 vs day 7 was 0.53(0.23-1.22); ORa for excellent and good quality embryos vs fair and poor quality embryos was 0.86(0.54-1.36). Regarding the studied factors that were potentially associated with iatrogenic mosaicism, neither type of stimulation (agonists vs antagonists), nor the use of two different single-step media showed an association. No impact on the incidence on mosaicism has been related to a given operator. Conclusions There is no preferential distribution of mosaicism among the chromosomes although large chromosomes seem to be more sensitive to segmental mosaicism. None of the studied potential sources of intrinsic, iatrogenic, or artefactual mosaicism showed an association with such feature.

Trabajo presentado en el Congreso 25 aniversario crea nuevos desafios en medicina reproductiva, celebrado en Valencia (España) del 27 al 28 de septiembre de 2018.

At re: publica 2010 Sascha Lobo gave a lecture entitled ‘How to Survive a Shitstorm’. Thus he established the term. The term ‘Shitstorm’ describe a phenomenon often observed on the social web: One or two critical remarks trigger aggressive, insultung and even threatening verbal attacks which culminate in a ‘wave of indignation’ through active as well as seemingly passive actors.