ESHRE guideline: number of embryos to transfer during IVF/ICSI†.

STUDY QUESTION: Which clinical and embryological factors should be considered to apply double embryo transfer (DET) instead of elective single embryo transfer (eSET)? SUMMARY ANSWER: No clinical or embryological factor per se justifies a recommendation of DET instead of eSET in IVF/ICSI. WHAT IS KNOWN ALREADY: DET is correlated with a higher rate of multiple pregnancy, leading to a subsequent increase in complications for both mother and babies. These complications include preterm birth, low birthweight, and other perinatal adverse outcomes. To mitigate the risks associated with multiple pregnancy, eSET is recommended by international and national professional organizations as the preferred approach in ART. STUDY DESIGN, SIZE, DURATION: The guideline was developed according to the structured methodology for development and update of ESHRE guidelines. Literature searches were performed in PUBMED/MEDLINE and Cochrane databases, and relevant papers published up to May 2023, written in English, were included. Live birth rate, cumulative live birth rate, and multiple pregnancy rate were considered as critical outcomes. PARTICIPANTS/MATERIALS, SETTING, METHODS: Based on the collected evidence, recommendations were discussed until a consensus was reached within the Guideline Development Group (GDG). A stakeholder review was organized after the guideline draft was finalized. The final version was approved by the GDG and the ESHRE Executive Committee. MAIN RESULTS AND THE ROLE OF CHANCE: The guideline provides 35 recommendations on the medical and non-medical risks associated with multiple pregnancies and on the clinical and embryological factors to be considered when deciding on the number of embryos to transfer. These recommendations include 25 evidence-based recommendations, of which 24 were formulated as strong recommendations and one as conditional, and 10 good practice points. Of the evidence-based recommendations, seven (28%) were supported by moderate-quality evidence. The remaining recommendations were supported by low (three recommendations; 12%), or very low-quality evidence (15 recommendations; 60%). Owing to the lack of evidence-based research, the guideline also clearly mentions recommendations for future studies. LIMITATIONS, REASONS FOR CAUTION: The guideline assessed different factors one by one based on existing evidence. However, in real life, clinicians' decisions are based on several prognostic factors related to each patient's case. Furthermore, the evidence from randomized controlled trials is too scarce to formulate high-quality evidence-based recommendations. WIDER IMPLICATIONS OF THE FINDINGS: The guideline provides health professionals with clear advice on best practice in the decision-making process during IVF/ICSI, based on the best evidence currently available, and recommendations on relevant information that should be communicated to patients. In addition, a list of research recommendations is provided to stimulate further studies in the field. STUDY FUNDING/COMPETING INTEREST(S): The guideline was developed and funded by ESHRE, covering expenses associated with the guideline meetings, the literature searches, and the dissemination of the guideline. The guideline group members did not receive payment. DPB declared receiving honoraria for lectures from Merck, Ferring, and Gedeon Richter. She is a member of ESHRE EXCO, and the Mediterranean Society for reproductive medicine and the president of the Croatian Society for Gynaecological Endocrinology and Reproductive Medicine. CDG is the past Chair of the ESHRE EIM Consortium and a paid deputy member of the Editorial board of Human Reproduction. IR declared receiving reimbursement from ESHRE and EDCD for attending meetings. She holds an unpaid leadership role in OBBCSSR, ECDC Sohonet, and AER. KAR-W declared receiving grants for clinical researchers and funding provision to the institution from the Swedish Cancer Society (200170F), the Senior Clinical Investigator Award, Radiumhemmets Forskningsfonder (Dnr: 201313), Stockholm County Council FoU (FoUI-953912) and Karolinska Institutet (Dnr 2020-01963), NovoNordisk, Merck and Ferring Pharmaceuticals. She received consulting fees from the Swedish Ministry of Health and Welfare. She received honoraria from Roche, Pfizer, and Organon for chairmanship and lectures. She received support from Organon for attending meetings. She participated in advisory boards for Merck, Nordic countries, and Ferring. She declared receiving time-lapse equipment and grants with payment to institution for pre-clinical research from Merck pharmaceuticals and from Ferring. SS-R received research funding from Roche Diagnostics, Organon/MSD, Theramex, and Gedeo-Richter. He received consulting fees from Organon/MSD, Ferring Pharmaceuticals, and Merck Serono. He declared receiving honoraria for lectures from Ferring Pharmaceuticals, Besins, Organon/MSD, Theramex, and Gedeon Richter. He received support for attending Gedeon Richter meetings and participated in the Data Safety Monitoring Board of the T-TRANSPORT trial. He is the Deputy of ESHRE SQART special interest group. He holds stock options in IVI Lisboa and received equipment and other services from Roche Diagnostics and Ferring Pharmaceuticals. KT declared receiving payment for honoraria for giving lectures from Merck Serono and Organon. She is member of the safety advisory board of EDQM. She holds a leadership role in the ICCBBA board of directors. ZV received reimbursement from ESHRE for attending meetings. She also received research grants from ESHRE and Juhani Aaltonen Foundation. She is the coordinator of EHSRE SQART special interest group. The other authors have no conflicts of interest to declare. DISCLAIMER: This guideline represents the views of ESHRE, which were achieved after careful consideration of the scientific evidence available at the time of preparation. In the absence of scientific evidence on certain aspects, a consensus between the relevant ESHRE stakeholders has been obtained. Adherence to these clinical practice guidelines does not guarantee a successful or specific outcome, nor does it establish a standard of care. Clinical practice guidelines do not replace the need for application of clinical judgement to each individual presentation, nor variations based on locality and facility type. ESHRE makes no warranty, express or implied, regarding the clinical practice guidelines and specifically excludes any warranties of merchantability and fitness for a particular use or purpose (full disclaimer available at https://www.eshre.eu/Guidelines-and-Legal).

Shedding light on the ART laboratory.

This commentary examines the impact of light conditions in the assisted reproductive technology (ART) laboratory, specifically considering gametes and embryo culture. While these processes traditionally occur in the absence of light within the female reproductive tract, laboratory conditions often involve exposure to varying wavelengths, intensities and light sources. Although literature reports describe potential detrimental effects of certain wavelengths of light on biological material, these findings are often based on experiments that might not reflect actual laboratory conditions. Current ART laboratory practices aim to minimize light exposure; however, some procedures necessitate light exposure, typically involving microscopy. Results from the authors' cross-sectional survey on light-intensity practices in ART laboratories revealed the frequent use of inadequate lighting, leading to errors and impacting staff well-being. A failure mode and effects analysis was used to identify potential failure modes and their impacts due to poor lighting. Overall, this manuscript stresses the importance of maintaining proper ambient lighting in the ART laboratory, balancing the potentially detrimental effects of light on gametes and embryos against the need for proper lighting for accurate procedures and staff well-being. Adequate lighting not only ensures the safety of reproductive cells, but also improves process management and the operators' psychological conditions.

MicroRNA-146b negatively affects bovine embryo development and quality.

MicroRNAs (miRNAs), which can be carried inside extracellular vesicles (EVs), play a crucial role in regulating embryo development up to the blastocyst stage. Yet, the molecular mechanisms underlying blastocyst development and quality are largely unknown. Recently, our group identified 69 differentially expressed miRNAs in extracellular vesicles (EVs) isolated from culture medium conditioned by bovine embryos that either developed to the blastocyst stage or did not (non-blastocysts). We found miR-146b to be more abundant in the EVs derived from media conditioned by non-blastocyst embryos. Using RT-qPCR, we here confirmed the upregulation of miR-146b in non-blastocyst (arrested at 2-4 cell and morula stage) embryos compared to blastocysts (p<0.005), which coincides with the upregulation of miR-146b in EVs derived from the medium of these non-blastocysts. To evaluate a functional effect, bovine embryo culture media were supplemented with miR-146b mimics, resulting in significantly decreased embryo quality, with lower blastocyst rates at day 7 and lower total cell numbers, while the opposite was found after supplementation with miR-146b inhibitors, which resulted in reduced apoptosis rates (P < 0.01). Transcriptomic analysis of embryos treated with miR-146b mimics or inhibitors showed differential expression (P < 0.01) of genes associated with apoptosis, cell differentiation, and the RNA Pol II transcription complex, including WDR36, MBNL2, ERCC6l2, PYGO1, and SNIP1. Overall, miR-146b is overexpressed in non-blastocyst embryos and in EVs secreted by these embryos, and it regulates genes involved in embryo development and apoptosis, resulting in decreased embryo quality.

Composition and distribution of fatty acids in various lipid fractions in serum and follicular fluid of women undergoing assisted reproductive technology.

Fatty acids (FA) in follicular fluid (FF) are present in an esterified form [triglycerides, cholesterol esters and phospholipids] or as non-esterified FA, which partly originate from blood. However, a comprehensive comparison of blood vs. FF FA in various lipid classes is missing. The aim of this study was to determine the distribution of the FA composition in each lipid class of serum and FF, and to investigate their mutual correlations. A total of 74 patients undergoing assisted reproductive technology treatment were involved in the study. Both in serum as well as FF, saturated FA and mono-unsaturated FA were predominant in non-esterified FA and triglycerides fractions while poly-unsaturated FA were mainly present in phospholipids and cholesterol esters fractions, although phospholipids also contained high proportions of saturated FA. Irrespective of the lipid class, the FA proportions differed between serum and FF (P < 0.05). Despite these differences, most of the FA in triglycerides, phospholipids and cholesterol esters of FF were well correlated with their proportions in serum. Nevertheless, only weak to moderate associations (r < 0.60) were observed for the majority of the FA in the non-esterified FA fraction. Differences in FA product/precursor-ratios were found between serum and FF, such as higher C20:4n-6 to C18:2n-6 and C20:5n-3 to C18:3n-3 in FF. FA metabolism (e.g. desaturation and elongation) takes place in cells of the intrafollicular micro-environment. Moreover, good correlations between esterified FA in serum and FF suggest esterified FA in blood could be representative of esterified FA in FF.

Incidence of and indications for sperm donor restriction - analysis of patients continuing treatment: a retrospective single-centre study.

RESEARCH QUESTION: What are the incidence of and indications for sperm donor restriction due to suspected/confirmed disease risk, and the future treatment choices of patients using these sperm donors? DESIGN: This single-centre retrospective study involved donors who had restrictions on the use of their imported spermatozoa from January 2010 to December 2019, and current or previous recipients. Indications for sperm restriction and the characteristics of patients undergoing medically assisted reproduction (MAR) treatment with these specimens at the time of restriction were collected. Differential characteristics of women who decided on whether or not to contintue the procedure were assessed. Characteristics potentially leading to treatment continuation were identified. RESULTS: Of 1124 sperm donors identified, 200 (17.8%) were restricted, most commonly for multifactorial (27.5%) and autosomal recessive (17.5%) disorders. The spermatozoa had been used for 798 recipients, of whom 172, receiving spermatozoa from 100 donors, were informed about the restriction and constituted the 'decision cohort'. The specimens from the restricted donors were accepted by 71 (approximately 40%) patients, with 45 (approximately 63%) eventually using the restricted donor for their future MAR treatment. The odds of accepting the restricted spermatozoa decreased with increasing age (OR 0.857, 95% CI 0.800-0.918, P < 0.001) and the time between MAR treatment and the restriction date (OR 0.806, 95% CI 0.713-0.911, P < 0.001). CONCLUSION: Donor restriction due to suspected/confirmed disease risk is relatively frequent. This affected a relevant number of women (around 800), of whom 172 (approximately 20%) had to decide whether or not to use these donors further. Although donor screening is being performed thoroughly, there remain health risks for donor children. Realistic counselling of all stakeholders involved is necessary.

Reproductive health in transgender and gender diverse individuals: A narrative review to guide clinical care and international guidelines.

Background: Hormonal treatments and surgical interventions practiced with the aim to affirm gender identity in transgender and gender diverse patients may impact their future reproductive ability, family building, and family planning options. Whereas it is recommended by international guidelines to discuss the potential risks of infertility and to present fertility preservation (FP) options to transgender individuals and their families prior to initiating any of these treatments, many barriers still remain. Further, transgender and gender diverse individuals often experience barriers to accessing contraception, abortion, pre-conception care, and comprehensive perinatal care. Aims: In this review we summarize the current literature on reproductive healthcare issues reported in transgender people including fertility issues, fertility preservation (FP), contraception, pregnancy and lactation and perinatal health. Methods: A narrative literature search of major databases (Pubmed, Medline, PsycInfo, Google Scholar, Web of Science) was conducted. Given the paucity and heterogeneity of studies, summative review tactics were not available. The literature was critically reviewed by international experts in the field with focus on the impact of gender-affirming medical interventions on future fertility, current FP options and reproductive health issues in transgender people. Results: The current literature supports that transgender and gender diverse individuals may wish to have genetically related children in the future, rendering the issue of FP relevant to this patient group. The cryopreservation of mature gametes is an efficacious option for FP for post-pubertal adolescents and adults. It is recommended to discuss these options at time of planning for gender-affirming hormonal therapy (GAHT) or engaging with other gender-affirming procedures that can limit future fertility. Discontinuation of GAHT may allow individuals to undergo FP later, but data are limited and there is the concern of symptoms and consequences of stopping GAHT. For pre-pubertal and early pubertal children, FP options are limited to the cryopreservation of gonadal tissue. At present the tissue can become functional only after re-transplantation, which might be undesirable by transgender individuals in the future. Preconception counseling, prenatal surveillance, perinatal support, contraceptive, and pregnancy termination related healthcare need to be meaningfully adapted for this patient population, and many knowledge gaps remain. Discussion: Specialized FP reproductive healthcare for transgender and gender diverse individuals is in early evolution. Research should be conducted to examine effects of medical interventions on fertility, timing of FP, gamete preservation and outcome of the fertility treatments. Strategies to inform and educate transgender and gender diverse patients can lead to optimization of reproductive care and counseling and decision making of FP for this population.

The fatty acid composition in follicles is related to the developmental potential of oocytes up to the blastocyst stage: a single-centre cohort study.

BACKGROUND: Advanced maternal age and obesity are associated with impaired female fertility. Moreover, fatty acids (FA) in follicular fluid (FF) play important roles in oocyte maturation and embryo development. However, the effects of body mass index (BMI), age, and FF FA composition on embryo development between days 3 and 5 and blastocyst stage on day 5 are still unclear. METHODS: This study included 138 patients undergoing assisted reproductive technology (ART), which were divided into three BMI groups (18.5-24.9 kg/m2 vs. 25.0-29.9 kg/m2 vs. ≥ 30.0 kg/m2) and three age-related groups (20-30 years vs. 31-34 years vs. ≥ 35 years) which were compared for ART outcomes. Further, observations were divided into quartiles based on either of three parameters related to embryo outcome, i.e. (i) embryos developing between days 3 and 5 (ED3-5) and (ii) expanded blastocysts on day 5 (EB5), both expressed proportionally to the number of oocytes with two pronuclei (2PN), as well as (iii) the embryo utilization rate (EUR). Proportions of FF FA were then compared between Q1 and Q4, representing the quartile with the worst vs. the best embryo outcome, respectively. Finally, regression models were created to assess the relationships between BMI, age, FF total FA (TFA) concentration, relative proportions of specific FA and embryo outcome. RESULTS: Patients of Q1 had higher proportions of FF C20:5n-3, C22:6n-3 and total n-3 PUFA than Q4 patients. Furthermore, Q4 patients tended to be younger than Q1 patients. Within the whole cohort, the proportion of C20:5n-3 negatively correlated with ED3-5/2PN and EUR, while EB5/2PN tended to be negatively correlated with age. Regression models within the overweight and obese group confirmed the negative relation between C20:5n-3 and ED3-5/2PN, but also indicated additional associations: C18:1n-9 and C20:4n-6 were positively associated with ED3-5/2PN and EUR, respectively while the proportion of C18:0 was negatively associated with EUR. CONCLUSION: The proportions of n-3 PUFA, particularly C20:5n-3 and C22:6n-3 were reduced in the patients' quartile with the best embryo outcome. This group of patients was also younger. However, the embryo quality parameters of overweight/obese patients were not associated with age but were positively associated with FF C18:1n-9 and negatively with the proportions of C18:0 or C20:5n-3. TRIAL REGISTRATION: This study' registration number was B670201627735.

Endocrine outcome and seminal parameters in young adult men born with hypospadias: A cross-sectional cohort study.

BACKGROUND: Hypospadias affects around 1/200 newborn males. Intrauterine testicular dysfunction may underlie a subset of cases. The long-term endocrine and reproductive outcomes in these men remain largely unknown. METHODS: Cross-sectional study in Ghent and Vienna University Hospitals to assess the endocrine and seminal parameters of young adult men (16-21 years) born with non-syndromic hypospadias (NSH) (n = 193) compared to healthy typical males (n = 50). Assessments included physical exam, semen analysis, hormone assays and exome-based gene panel analysis (474 genes). FINDINGS: All participants had experienced a spontaneous puberty, in spite of higher LH and INSL3 levels than typical males. Oligo- or azoospermia was observed in 32/172 (18·6%; 99%-CI: 12·2-27·4%) of NSH men; but in 5/16 (31·3%; 99%-CI: 11·1;62·4%) of complex NSH men and in 13/22 (59·1%; 99%-CI: 33·2-80·7%) of those born small for gestational age (SGA). No (likely) pathogenic coding variants were found in the investigated genes. Suboptimal statural growth affected 8/23 (34·8%; 99%-CI: 15·4-61·0%) of men born SGA with NSH. INTERPRETATION: Spermatogenesis is significantly compromised in NSH men, especially in those born SGA or those with complex NSH. Long-term andrological follow-up is recommended, including end-pubertal semen analysis. No clear monogenic causes could be demonstrated in our cohort even in proximal or complex NSH. Being born SGA with NSH is frequently associated with poor catch-up growth, requiring growth hormone therapy in some. FUNDING: Research grants from the European Society of Paediatric Endocrinology, the Belgian Society of Pediatrics, the Belgian Society of Pediatric Endocrinology and Diabetology and the Research Foundation Flanders (FWO).

Screening of chlamydia trachomatis in non-partner donors: situation of stored donations and proposal for periodic screening.

RESEARCH QUESTION: Lack of guidance on the frequency of Chlamydia trachomatis screening in non-partner donors has led to heterogeneous testing protocols. C. trachomatis was checked in sperm donations unscreened for C. trachomatis to determine the risk for C. trachomatis infection in recipients using historic sperm donations unscreened for C. trachomatis. A C. trachomatis screening protocol is proposed to harmonize C. trachomatis screening, for which a cost evaluation is provided. DESIGN: Retrospective study of sperm donations carried out between 2009 and 2019 from healthy non-partner donors for whom at least one straw was available. A straw was selected from the still available donations that had not been tested for C. trachomatis in urine at the time of donation. These sperm samples were screened for C. trachomatis by nucleic acid amplification (NAT). RESULTS: Forty donors were included in the analysis. The 210 analysed straws tested negative for C. trachomatis. A C. trachomatis screening protocol following the European Centre for Disease Prevention and Control (ECDC) protocol for other sexually transmitted diseases (STD), i.e. NAT C. trachomatis screening of donor eligibility and first and last donation, provided these occur within 90 days, is cost advantageous compared with screening of all samples (approximately 75% reduction). CONCLUSION: A negligible risk for C. trachomatis infection was found in recipients when using historical sperm samples stored at the sperm bank. C. trachomatis screening following the ECDC protocol for other STDs is supported as it significantly reduces workload and cost compared with screening all samples.

Hatching is modulated by microRNA-378a-3p derived from extracellular vesicles secreted by blastocysts.

SignificanceHatching from the zona pellucida is a prerequisite for embryo implantation and is less likely to occur in vitro for reasons unknown. Extracellular vesicles (EVs) are secreted by the embryo into the culture medium. Yet the role that embryonic EVs and their cargo microRNAs (miRNAs) play in blastocyst hatching has not been elucidated, partially due to the difficulties of isolating them from low amounts of culture medium. Here, we optimized EV-miRNA isolation from medium conditioned by individually cultured bovine embryos and subsequently showed that miR-378a-3p, which was up-regulated in EVs secreted by blastocysts, plays a crucial role in promoting blastocyst hatching. This demonstrates the regulatory effect of miR-378-3p on hatching, which is an established embryo quality parameter linked with implantation.

Good practice recommendations for information provision for those involved in reproductive donation†.

STUDY QUESTION: What information and support should be offered to donors, intended parents and donor-conceived people, in general and in consideration of the availability of direct-to-consumer genetic testing and matching services? SUMMARY ANSWER: For donors, intended parents and donor-conceived offspring, recommendations are made that cover information needs and informed consent, psychosocial implications and disclosure. WHAT IS KNOWN ALREADY: Trends indicate that the use of donor-assisted conception is growing and guidance is needed to help these recipients/intended parents, the donors and offspring, navigate the rapidly changing environment in which donor-assisted conception takes place. STUDY DESIGN SIZE DURATION: A working group (WG) collaborated on writing recommendations based, where available, on evidence collected from a literature search and expert opinion. Draft recommendations were published for stakeholder review and adapted where relevant based on the comments received. PARTICIPANTS/MATERIALS SETTING METHODS: Papers retrieved from PUBMED were included from 1 January 2014 up to 31 August 2020, focusing on studies published since direct-to-consumer genetic testing has become more widespread and accessible. The current paper is limited to reproductive donation performed in medically assisted reproduction (MAR) centres (and gamete banks): donation outside the medical context was not considered. MAIN RESULTS AND THE ROLE OF CHANCE: In total, 32 recommendations were made for information provision and support to donors, 32 for intended parents and 27 for donor-conceived offspring requesting information/support. LIMITATIONS REASONS FOR CAUTION: The available evidence in the area of reproductive donation is limited and diverse with regards to the context and types of donation. General conclusions and recommendations are largely based on expert opinion and may need to be adapted in light of future research. WIDER IMPLICATIONS OF THE FINDINGS: These recommendations provide guidance to MAR centres and gamete banks on good practice in information provision and support but should also be considered by regulatory bodies and policymakers at a national and international level to guide regulatory and legislative efforts towards the protection of donors and donor-conceived offspring. STUDY FUNDING/COMPETING INTERESTS: The development of this good practice paper was funded by European Society of Human Reproduction and Embryology (ESHRE), covering expenses associated with the WG meetings, the literature searches and dissemination. The WG members did not receive any payment. The authors have no conflicts of interest to declare. DISCLAIMER: This document represents the views of ESHRE, which are the result of consensus between the relevant ESHRE stakeholders and where relevant based on the scientific evidence available at the time of preparation. The recommendations should be used for informational and educational purposes. They should not be interpreted as setting a standard of care, or be deemed inclusive of all proper methods of care nor exclusive of other methods of care reasonably directed to obtaining the same results. They do not replace the need for application of clinical judgement to each individual presentation, nor variations based on locality and facility type. †ESHRE pages content is not externally peer reviewed. The manuscript has been approved by the Executive Committee of ESHRE.

Machine learning for prediction of euploidy in human embryos: in search of the best-performing model and predictive features.

OBJECTIVE: To assess the best-performing machine learning (ML) model and features to predict euploidy in human embryos. DESIGN: Retrospective cohort analysis. SETTING: Department for reproductive medicine in a university hospital. PATIENT(S): One hundred twenty-eight infertile couples treated between January 2016 and December 2019. Demographic and clinical data and embryonic developmental and morphokinetic data from 539 embryos (45% euploid, 55% aneuploid) were analyzed. INTERVENTION(S): Random forest classifier (RFC), scikit-learn gradient boosting classifier, support vector machine, multivariate logistic regression, and naïve Bayes ML models were trained and used in 9 databases containing either 26 morphokinetic features (as absolute [A1] or interim [A2] times or combined [A3]) alone or plus 19 standard development features [B1, B2, and B3] with and without 40 demographic and clinical characteristics [C1, C2, and C3]. Feature selection and model retraining were executed for the best-performing combination of model and dataset. MAIN OUTCOME MEASURE(S): The main outcome measures were overall accuracy, precision, recall or sensitivity, F1 score (the weighted average of precision and recall), and area under the receiver operating characteristic curve (AUC) of ML models for each dataset. The secondary outcome measure was ranking of feature importance for the best-performing combination of model and dataset. RESULT(S): The RFC model had the highest accuracy (71%) and AUC (0.75) when trained and used on dataset C1. The precision, recall or sensitivity, F1 score, and AUC were 66%, 86%, 75%, and 0.75, respectively. The accuracy, recall or sensitivity, and F1 score increased to 72%, 88%, and 76%, respectively, after feature selection and retraining. Morphokinetic features had the highest relative predictive weight. CONCLUSION(S): The RFC model can predict euploidy with an acceptable accuracy (>70%) using a dataset including embryos' morphokinetics and standard embryonic development and subjects' demographic and clinical features.

In Vitro Maturation of Oocytes Retrieved from Ovarian Tissue: Outcomes from Current Approaches and Future Perspectives.

In vitro maturation (IVM) of transvaginally aspirated immature oocytes is an effective and safe assisted reproductive treatment for predicted or high responder patients. Currently, immature oocytes are also being collected from the contralateral ovary during laparoscopy/laparotomy and even ex vivo from the excised ovary or the spent media during ovarian tissue preparation prior to ovarian cortex cryopreservation. The first live births from in vitro-matured ovarian tissue oocytes (OTO-IVM) were reported after monophasic OTO-IVM, showing the ability to achieve mature OTO-IVM oocytes. However, fertilisations rates and further embryological developmental capacity appeared impaired. The introduction of a biphasic IVM, also called capacitation (CAPA)-IVM, has been a significant improvement of the oocytes maturation protocol. However, evidence on OTO-IVM is still scarce and validation of the first results is of utmost importance to confirm reproducibility, including the follow-up of OTO-IVM children. Differences between IVM and OTO-IVM should be well understood to provide realistic expectations to patients.

The Maribor consensus: report of an expert meeting on the development of performance indicators for clinical practice in ART.

STUDY QUESTION: Is it possible to define a set of performance indicators (PIs) for clinical work in ART, which can create competency profiles for clinicians and for specific clinical process steps? SUMMARY ANSWER: The current paper recommends six PIs to be used for monitoring clinical work in ovarian stimulation for ART, embryo transfer, and pregnancy achievement: cycle cancellation rate (before oocyte pick-up (OPU)) (%CCR), rate of cycles with moderate/severe ovarian hyperstimulation syndrome (OHSS) (%mosOHSS), the proportion of mature (MII) oocytes at ICSI (%MII), complication rate after OPU (%CoOPU), clinical pregnancy rate (%CPR), and multiple pregnancy rate (%MPR). WHAT IS KNOWN ALREADY: PIs are objective measures for evaluating critical healthcare domains. In 2017, ART laboratory key PIs (KPIs) were defined. STUDY DESIGN SIZE DURATION: A list of possible indicators was defined by a working group. The value and limitations of each indicator were confirmed through assessing published data and acceptability was evaluated through an online survey among members of ESHRE, mostly clinicians, of the special interest group Reproductive Endocrinology. PARTICIPANTS/MATERIALS SETTING METHODS: The online survey was open for 5 weeks and 222 replies were received. Statements (indicators, indicator definitions, or general statements) were considered accepted when ≥70% of the responders agreed (agreed or strongly agreed). There was only one round to seek levels of agreement between the stakeholders.Indicators that were accepted by the survey responders were included in the final list of indicators. Statements reaching less than 70% were not included in the final list but were discussed in the paper. MAIN RESULTS AND THE ROLE OF CHANCE: Cycle cancellation rate (before OPU) and the rate of cycles with moderate/severe OHSS, calculated on the number of started cycles, were defined as relevant PIs for monitoring ovarian stimulation. For monitoring ovarian response, trigger and OPU, the proportion of MII oocytes at ICSI and complication rate after OPU were listed as PIs: the latter PI was defined as the number of complications (any) that require an (additional) medical intervention or hospital admission (apart from OHSS) over the number of OPUs performed. Finally, clinical pregnancy rate and multiple pregnancy rate were considered relevant PIs for embryo transfer and pregnancy. The defined PIs should be calculated every 6 months or per 100 cycles, whichever comes first. Clinical pregnancy rate and multiple pregnancy rate should be monitored more frequently (every 3 months or per 50 cycles). Live birth rate (LBR) is a generally accepted and an important parameter for measuring ART success. However, LBR is affected by many factors, even apart from ART, and it cannot be adequately used to monitor clinical practice. In addition to monitoring performance in general, PIs are essential for managing the performance of staff over time, and more specifically the gap between expected performance and actual performance measured. Individual clinics should determine which indicators are key to the success in their organisation based on their patient population, protocols, and procedures, and as such, which are their KPIs. LIMITATIONS REASONS FOR CAUTION: The consensus values are based on data found in the literature and suggestions of experts. When calculated and compared to the competence/benchmark limits, prudent interpretation is necessary taking into account the specific clinical practice of each individual centre. WIDER IMPLICATIONS OF THE FINDINGS: The defined PIs complement the earlier defined indicators for the ART laboratory. Together, both sets of indicators aim to enhance the overall quality of the ART practice and are an essential part of the total quality management. PIs are important for education and can be applied during clinical subspecialty. STUDY FUNDING/COMPETING INTERESTS: This paper was developed and funded by ESHRE, covering expenses associated with meetings, literature searches, and dissemination. The writing group members did not receive payment.Dr G.G. reports personal fees from Merck, MSD, Ferring, Theramex, Finox, Gedeon-Richter, Abbott, Biosilu, ReprodWissen, Obseva, PregLem, and Guerbet, outside the submitted work. Dr A.D. reports personal fees from Cook, outside the submitted work; Dr S.A. reports starting a new employment in May 2020 at Vitrolife. Previously, she has been part of the Nordic Embryology Academic Team, with meetings were sponsored by Gedeon Richter. The other authors have no conflicts of interest to declare. DISCLAIMER: This document represents the views of ESHRE, which are the result of consensus between the relevant ESHRE stakeholders and where relevant based on the scientific evidence available at the time of preparation.The recommendations should be used for informational and educational purposes. They should not be interpreted as setting a standard of care, or be deemed inclusive of all proper methods of care nor exclusive of other methods of care reasonably directed to obtaining the same results. They do not replace the need for application of clinical judgment to each individual presentation, nor variations based on locality and facility type.Furthermore, ESHREs recommendations do not constitute or imply the endorsement, recommendation, or favouring of any of the included technologies by ESHRE.

Low feasibility of in vitro matured oocytes originating from cumulus complexes found during ovarian tissue preparation at the moment of gender confirmation surgery and during testosterone treatment for fertility preservation in transgender men.

OBJECTIVE: To study the feasibility of in vitro maturation of ovarian tissue oocytes for fertility preservation in transgender men on testosterone treatment. DESIGN: Cross-sectional study SETTING: University hospital PATIENT(S): Eighty-three transgender men enrolled from November 2015 to January 2019 INTERVENTION(S): In vitro maturation of cumulus-oocyte complexes (COCs) harvested at the time of gender confirmation surgery, and fertilization through intracytoplasmic sperm injection. MAIN OUTCOME MEASURE(S): In vitro maturation, fertilization, and blastulation rates; comparison of morphokinetics with vitrified-warmed oocytes; and analysis of the genetic profiles of embryos. SECONDARY OUTCOMES: association between serum hormone levels; COCs' morphologic characteristics, and vitrification rate. RESULT(S): All participants were on testosterone treatment for a median of 83 (64[Quartile 1]; 113.2[Quartile 2]) weeks. A total of 1,903 COCs (mean per participant, 23 ± 15.8) were collected. The in vitro maturation rate was 23.8%, vitrification rate was 21.5%, and survival rate after warming was 72.6% (n = 151). Intracytoplasmic sperm injection was performed in 139 oocytes. The rate of normal fertilized oocytes was 34.5%, and 25 (52.1%) embryos reached day 3. One blastocyst was achieved on day 5. Aberrant cleavage patterns and early embryo arrest were observed in 22 (45.8%) and 44 (91.7%) zygotes, respectively. Compared with vitrified-warmed donor oocytes, a delay was observed in pronuclei disappearance, t2 (time to reach 2 cell stage) timings, and CC1 (the duration of the 1st cell cycle) and SS3 (synchronization of cleavage pattern (calculated as t8-t5) time intervals. A normal genetic pattern was seen in 42% embryos. The proportion of vitrified oocytes was negatively associated with progesterone (odds ratio, 0.76) and positively associated with antimüllerian hormone serum levels (odds ratio, 1.23). The highest vitrification rate was achieved by the morphologic characteristic 344 at day 0 and by 433 at day 2. CONCLUSION(S): Ovarian tissue oocytes matured in vitro show low developmental capacity in transgender men, when collected under testosterone treatment.

ESHRE guideline: medically assisted reproduction in patients with a viral infection/disease.

STUDY QUESTION: What is the recommended management for medically assisted reproduction (MAR) in patients with a viral infection or disease, based on the best available evidence in the literature? SUMMARY ANSWER: The ESHRE guideline on MAR in patients with a viral infection/disease makes 78 recommendations on prevention of horizontal and vertical transmission before, during and after MAR, and the impact on its outcomes, and these also include recommendations regarding laboratory safety on the processing and storage of gametes and embryos testing positive for viral infections. WHAT IS KNOWN ALREADY: The development of new and improved anti-viral medications has resulted in improved life expectancy and quality of life for patients with viral infections/diseases. Patients of reproductive age are increasingly exploring their options for family creation. STUDY DESIGN SIZE DURATION: The guideline was developed according to the structured methodology for the development of ESHRE guidelines. After the formulation of nine key questions for six viruses (hepatitis B virus, hepatitis C virus, human immunodeficiency virus, human papilloma virus, human T-lymphotropic virus I/II and Zika virus) by a group of experts, literature searches and assessments were performed. Papers published up to 2 November 2020 and written in English were included in the review. Evidence was analyzed by female, male or couple testing positive for the virus. PARTICIPANTS/MATERIALS SETTING METHODS: Based on the collected evidence, recommendations were formulated and discussed until consensus was reached within the guideline group. There were 61 key questions to be answered by the guideline development group (GDG), of which 12 were answered as narrative questions and 49 as PICO (Patient, Intervention, Comparison, Outcome) questions. A stakeholder review was organized after the finalization of the draft. The final version was approved by the GDG and the ESHRE Executive Committee. MAIN RESULTS AND THE ROLE OF CHANCE: This guideline aims to help providers meet a growing demand for guidance on the management of patients with a viral infection/disease presenting in the fertility clinic.The guideline makes 78 recommendations on prevention of viral transmission before and during MAR, and interventions to reduce/avoid vertical transmission to the newborn. Preferred MAR treatments and interventions are described together with the effect of viral infections on outcomes. The GDG formulated 44 evidence-based recommendations-of which 37 were formulated as strong recommendations and 7 as weak-33 good practice points (GPP) and one research only recommendation. Of the evidence-based recommendations, none were supported by high-quality evidence, two by moderate-quality evidence, 15 by low-quality evidence and 27 by very low-quality evidence. To support future research in the field of MAR in patients with a viral infection/disease, a list of research recommendations is provided. LIMITATIONS REASONS FOR CAUTION: Most interventions included are not well-studied in patients with a viral infection/disease. For a large proportion of interventions, evidence was very limited and of very low quality. More evidence is required for these interventions, especially in the field of human papilloma virus (HPV). Such future studies may require the current recommendations to be revised. WIDER IMPLICATIONS OF THE FINDINGS: The guideline provides clinicians with clear advice on best practice in MAR for patients with a viral infection/disease, based on the best evidence currently available. In addition, a list of research recommendations is provided to stimulate further studies in the field. STUDY FUNDING/COMPETING INTERESTS: The guideline was developed and funded by ESHRE, covering expenses associated with the guideline meetings, with the literature searches and with the dissemination of the guideline. The guideline group members did not receive any financial incentives, all work was provided voluntarily. A.D. reports research fees from Ferring and Merck, consulting fees from Ferring, outside the submitted work. C.P. reports speakers fees from Merck and MSD outside the submitted work. K.T. reports speakers fees from Cooper Surgical and Ferring and consultancy fees as member of the advisory board BioTeam of Ferring, outside the submitted work. The other authors have no conflicts of interest to declare. DISCLAIMER: This guideline represents the views of ESHRE, which were achieved after careful consideration of the scientific evidence available at the time of preparation. In the absence of scientific evidence on certain aspects, a consensus between the relevant ESHRE stakeholders has been obtained. Adherence to these clinical practice guidelines does not guarantee a successful or specific outcome, nor does it establish a standard of care. Clinical practice guidelines do not replace the need for application of clinical judgment to each individual presentation, nor variations based on locality and facility type. ESHRE makes no warranty, express or implied, regarding the clinical practice guidelines and specifically excludes any warranties of merchantability and fitness for a particular use or purpose. (Full disclaimer available at  www.eshre.eu/guidelines.).

Charting Extracellular Transcriptomes in The Human Biofluid RNA Atlas.

Extracellular RNAs present in biofluids have emerged as potential biomarkers for disease. Where most studies focus on blood-derived fluids, other biofluids may be more informative. We present an atlas of messenger, circular, and small RNA transcriptomes of a comprehensive collection of 20 human biofluids. By means of synthetic spike-in controls, we compare RNA content across biofluids, revealing a 10,000-fold difference in concentration. The circular RNA fraction is increased in most biofluids compared to tissues. Each biofluid transcriptome is enriched for RNA molecules derived from specific tissues and cell types. Our atlas enables an informed selection of the most relevant biofluid to monitor particular diseases. To verify the biomarker potential in these biofluids, four validation cohorts representing a broad spectrum of diseases were profiled, revealing numerous differential RNAs between case and control subjects. Spike-normalized data are publicly available in the R2 web portal for further exploration.

Parental desire and fertility preservation in assigned male at birth transgender people living in Belgium.

Introduction: Transgender people who chose to proceed with gender affirming hormonal and/or surgical therapy, may face reduced options for fulfilling their parental desire in the future. The ideas and concerns of adult transgender people regarding fertility preservation and parental desire have never been reported in a large, non-clinical sample of assigned male at birth (AMAB) transgender people. Methods: A web-based survey on fertility and parenthood in (binary and non-binary) transgender people was conducted in Belgium. AMAB people were selected for this analysis. Results: We included 254 AMAB persons, of which 196 (77.2%) self-identified as transgender women (TW), 14 (5.5%) as cross-dressers and 44 (17.3%) as gender non-binary (GNB) people. Fifty-five (21.6%) respondents had a current/future parental desire, parental desire was already fulfilled in 81 (31.9%) and not present in 57 people (22.4%) (other: 19.2%). TW were more likely to express a parental desire, compared to GNB people and cross-dressers (P = 0.004). In total, 196 AMAB people previously sought medical assistance, of which 30 (15.3%) considered the loss of fertility due to the transitioning process undesirable. The majority (68.2%) did not want fertility preservation (FP). Fourteen people (9.8%) had proceeded with FP. The main reasons not to proceed with FP included not feeling the need (70; 68.0%), not desiring a genetic link with (future) child(ren) (20; 19.4%) and having to postpone hormone treatment (15; 14.6%). TW were more likely to have a parental desire and to have cryopreserved or to consider cryopreserving gametes, compared to GNB people. Conclusion: Parental desire and FP use were lower in the current non-clinical sample than in previous research on clinical samples. This can possibly be explained by the barriers transgender people face when considering fertility options, including postponing hormone therapy. Also, GNB persons have different needs for gender affirming treatment and FP.

A methodological validation of an easy one-step swimout semen preparation procedure for selecting DNA fragmentation-free spermatozoa for ICSI.

The main purpose of this methodological paper was to describe a recently designed one-step ICSI semen preparation swim-out method (called swim-ICSI) and to compare its efficacy with our conventional two-step swim-out method for the selection of motile spermatozoa for ICSI with minimal DNA damage. In this observational cohort study, 42 fresh ejaculate sperm samples for ICSI were included to compare the new swim-ICSI with the conventional swim-out. In a sub-analysis (n = 20), both in-house designed ICSI preparation methods were compared with a commercial magnetic-activated cell sorting test (MACS® ). Sperm DNA fragmentation (SDF), using Halosperm® , was determined at different time points during sperm preparation: on the native sample (a), after density gradient centrifugation (DG) (b), on the motile (A + B) spermatozoa selected with conventional swim-out post-DG (c) and selected with swim-ICSI method post-DG (d). For a subgroup (n = 20), SDF was also calculated after MACS (e). The mean SDF significantly reduced after EACH preparation step and reduced to almost zero in the recovered A + B spermatozoa when the semen prepared with DG was further processed for ICSI (swim-ICSI vs. swim-out, p = .001). In conclusion, the optimised one-step and fine-tuned swim-ICSI technique shows the possibility to select a population of spermatozoa with almost zero SDF to be used in ICSI treatments.