A Method to Study the Meiotic Recombination Map in Human Preimplantation Blastocysts.

Homologous recombination plays pivotal roles in physical attachments and genetic diversity. In the past, it was studied among individuals from different populations. However, only few gametes from individual could generate offspring, which limits its exploration in nature selection. In the last few years, preimplantation blastocysts based on trio SNP-chip data were available in individuals for preimplantation genetic testing (PGT). In this protocol, we demonstrate how to detect meiotic recombination events and construct the genetic map based on trio SNP-chip data, obtained from biopsied blastocysts and their related individuals in PGT cycles, which may allow better understanding of recombination events in nature selection.

Factors affecting biochemical pregnancy loss (BPL) in preimplantation genetic testing for aneuploidy (PGT-A) cycles: machine learning-assisted identification.

PURPOSE: To determine the factors influencing the likelihood of biochemical pregnancy loss (BPL) after transfer of a euploid embryo from preimplantation genetic testing for aneuploidy (PGT-A) cycles. METHODS: The study employed an observational, retrospective cohort design, encompassing 6020 embryos from 2879 PGT-A cycles conducted between February 2013 and September 2021. Trophectoderm biopsies in day 5 (D5) or day 6 (D6) blastocysts were analyzed by next generation sequencing (NGS). Only single embryo transfers (SET) were considered, totaling 1161 transfers. Of these, 49.9% resulted in positive pregnancy tests, with 18.3% experiencing BPL. To establish a predictive model for BPL, both classical statistical methods and five different supervised classification machine learning algorithms were used. A total of forty-seven factors were incorporated as predictor variables in the machine learning models. RESULTS: Throughout the optimization process for each model, various performance metrics were computed. Random Forest model emerged as the best model, boasting the highest area under the ROC curve (AUC) value of 0.913, alongside an accuracy of 0.830, positive predictive value of 0.857, and negative predictive value of 0.807. For the selected model, SHAP (SHapley Additive exPlanations) values were determined for each of the variables to establish which had the best predictive ability. Notably, variables pertaining to embryo biopsy demonstrated the greatest predictive capacity, followed by factors associated with ovarian stimulation (COS), maternal age, and paternal age. CONCLUSIONS: The Random Forest model had a higher predictive power for identifying BPL occurrences in PGT-A cycles. Specifically, variables associated with the embryo biopsy procedure (biopsy day, number of biopsied embryos, and number of biopsied cells) and ovarian stimulation (number of oocytes retrieved and duration of stimulation), exhibited the strongest predictive power.

Preimplantation Genetic Testing of Spinocerebellar Ataxia Type 3/Machado-Joseph Disease-Robust Tools for Direct and Indirect Detection of the ATXN3 (CAG)n Repeat Expansion.

Spinocerebellar ataxia type 3/Machado-Joseph disease (SCA3/MJD) is a neurodegenerative disorder caused by the ATXN3 CAG repeat expansion. Preimplantation genetic testing for monogenic disorders (PGT-M) of SCA3/MJD should include reliable repeat expansion detection coupled with high-risk allele determination using informative linked markers. One couple underwent SCA3/MJD PGT-M combining ATXN3 (CAG)n triplet-primed PCR (TP-PCR) with customized linkage-based risk allele genotyping on whole-genome-amplified trophectoderm cells. Microsatellites closely linked to ATXN3 were identified and 16 markers were genotyped on 187 anonymous DNAs to verify their polymorphic information content. In the SCA3/MJD PGT-M case, the ATXN3 (CAG)n TP-PCR and linked marker analysis results concurred completely. Among the three unaffected embryos, a single embryo was transferred and successfully resulted in an unaffected live birth. A total of 139 microsatellites within 1 Mb upstream and downstream of the ATXN3 CAG repeat were identified and 8 polymorphic markers from each side were successfully co-amplified in a single-tube reaction. A PGT-M assay involving ATXN3 (CAG)n TP-PCR and linkage-based risk allele identification has been developed for SCA3/MJD. A hexadecaplex panel of highly polymorphic microsatellites tightly linked to ATXN3 has been developed for the rapid identification of informative markers in at-risk couples for use in the PGT-M of SCA3/MJD.

Efficacy of non-invasive chromosome screening, preimplantation genetic testing for aneuploidy, and morphological grading in selecting embryos of patients with advanced maternal age: a three-armed prospective cohort study.

BACKGROUND: Non-invasive chromosome screening (NICS) and trophectoderm biopsy preimplantation genetic testing for aneuploidy (TE-PGT) were both applied for embryo ploidy detection, However, the cumulative live birth rates (CLBR) of NICS and TE-PGT in older age groups have yet to be reported. This study aimed to ascertain whether NICS and TE-PGT could enhance the cumulative live birth rates among patients of advanced maternal age. METHODS: A total of 384 couples aged 35-40 years were recruited. The patients were assigned to three groups: NICS, TE-PGT, and intracytoplasmic sperm injection (ICSI). All patients received frozen single blastocyst transfer. Patients in the NICS and TE-PGT groups underwent aneuploidy screening. RESULTS: When compared to the ICSI group, the CLBR was significantly higher in the NICS and TE-PGT groups (27.9% vs. 44.9% vs. 51.0%, p = 0.003 for NICS vs. ICSI, p < 0.001 for TE-PGT vs. ICSI). There were no significant differences in the clinical outcomes between the NICS and TE-PGT groups. Adjusting for confounding factors, the NICS and TE-PGT groups still showed a higher CLBR than the ICSI group (adjusted odds ratio (OR) 3.847, 95% confidence interval (CI) 1.939 to 7.634; adjusted OR 3.795, 95% CI 1.981 to 7.270). Additionally, the cumulative pregnancy loss rates of the NICS and TE-PGT groups were significantly lower than that of the ICSI group (adjusted OR 0.277, 95% CI 0.087 to 0.885; adjusted OR 0.182, 95% CI 0.048 to 0.693). There was no significant difference in the birth weights of the three groups (p = 0.108). CONCLUSIONS: In women 35-40 years old, the CLBR can be increased by selecting euploid embryos using NICS and TE-PGT. For elderly women at high risk of embryonic aneuploidy, NICS, characterized by its safety and non-invasive nature, may emerge as an alternative option for preimplantation genetic testing.

[Ethical Reflections on Preimplantation Genetic Diagnoses].

With fertility rates at an all-time low, children have become even more the 'treasures' of their families. Progress in genetic selection technology has made preimplantation genetic diagnosis an increasingly common practice in clinics. However, the practice of purposively selecting genes for future children remains controversial. In this article, the process of preimplantation genetic diagnosis is introduced and related philosophical and social perspectives are reviewed. Finally, the ethics related to this practice are discussed in the contexts of obligation theory, utility theory, and four ethical principles. The authors hope this article sheds light on the diverse perspectives used to consider and discuss the ethical issues surrounding gene selection and, importantly, helps nurses provide care grounded in ethics and humanity in ethically uncertain circumstances.

Investigating developmental characteristics of biopsied blastocysts stratified by mitochondrial copy numbers using time-lapse monitoring.

BACKGROUND: For in vitro fertilization (IVF), mitochondrial DNA (mtDNA) levels in the trophectodermal (TE) cells of biopsied blastocysts have been suggested to be associated with the cells' developmental potential. However, scholars have reached differing opinions regarding the use of mtDNA levels as a reliable biomarker for predicting IVF outcomes. Therefore, this study aims to assess the association of mitochondrial copy number measured by mitoscore associated with embryonic developmental characteristics and ploidy. METHODS: This retrospective study analyzed the developmental characteristics of embryos and mtDNA levels in biopsied trophectodermal cells. The analysis was carried out using time-lapse monitoring and next-generation sequencing from September 2021 to September 2022. Five hundred and fifteen blastocysts were biopsied from 88 patients undergoing IVF who met the inclusion criteria. Embryonic morphokinetics and morphology were evaluated at 118 h after insemination using all recorded images. Blastocysts with appropriate morphology on day 5 or 6 underwent TE biopsy and preimplantation genetic testing for aneuploidy (PGT-A). Statistical analysis involved generalized estimating equations, Pearson's chi-squared test, Fisher's exact test, and Kruskal-Wallis test, with a significance level set at P < 0.05. RESULTS: To examine differences in embryonic characteristics between blastocysts with low versus high mitoscores, the blastocysts were divided into quartiles based on their mitoscore. Regarding morphokinetic characteristics, no significant differences in most developmental kinetics and observed cleavage dysmorphisms were discovered. However, blastocysts in mitoscore group 1 had a longer time for reaching 3-cell stage after tPNf (t3; median: 14.4 h) than did those in mitoscore group 2 (median: 13.8 h) and a longer second cell cycle (CC2; median: 11.7 h) than did blastocysts in mitoscore groups 2 (median: 11.3 h) and 4 (median: 11.4 h; P < 0.05). Moreover, blastocysts in mitoscore group 4 had a lower euploid rate (22.6%) and a higher aneuploid rate (59.1%) than did those in the other mitoscore groups (39.6-49.3% and 30.3-43.2%; P < 0.05). The rate of whole-chromosomal alterations in mitoscore group 4 (63.4%) was higher than that in mitoscore groups 1 (47.3%) and 2 (40.1%; P < 0.05). A multivariate logistic regression model was used to analyze associations between the mitoscore and euploidy of elective blastocysts. After accounting for factors that could potentially affect the outcome, the mitoscore still exhibited a negative association with the likelihood of euploidy (adjusted OR = 0.581, 95% CI: 0.396-0.854; P = 0.006). CONCLUSIONS: Blastocysts with varying levels of mitochondrial DNA, identified through biopsies, displayed similar characteristics in their early preimplantation development as observed through time-lapse imaging. However, the mitochondrial DNA level determined by the mitoscore can be used as a standalone predictor of euploidy.

Artificial intelligence system for outcome evaluations of human in vitro fertilization-derived embryos.

BACKGROUND: In vitro fertilization (IVF) has emerged as a transformative solution for infertility. However, achieving favorable live-birth outcomes remains challenging. Current clinical IVF practices in IVF involve the collection of heterogeneous embryo data through diverse methods, including static images and temporal videos. However, traditional embryo selection methods, primarily reliant on visual inspection of morphology, exhibit variability and are contingent on the experience of practitioners. Therefore, an automated system that can evaluate heterogeneous embryo data to predict the final outcomes of live births is highly desirable. METHODS: We employed artificial intelligence (AI) for embryo morphological grading, blastocyst embryo selection, aneuploidy prediction, and final live-birth outcome prediction. We developed and validated the AI models using multitask learning for embryo morphological assessment, including pronucleus type on day 1 and the number of blastomeres, asymmetry, and fragmentation of blastomeres on day 3, using 19,201 embryo photographs from 8271 patients. A neural network was trained on embryo and clinical metadata to identify good-quality embryos for implantation on day 3 or day 5, and predict live-birth outcomes. Additionally, a 3D convolutional neural network was trained on 418 time-lapse videos of preimplantation genetic testing (PGT)-based ploidy outcomes for the prediction of aneuploidy and consequent live-birth outcomes. RESULTS: These two approaches enabled us to automatically assess the implantation potential. By combining embryo and maternal metrics in an ensemble AI model, we evaluated live-birth outcomes in a prospective cohort that achieved higher accuracy than experienced embryologists (46.1% vs. 30.7% on day 3, 55.0% vs. 40.7% on day 5). Our results demonstrate the potential for AI-based selection of embryos based on characteristics beyond the observational abilities of human clinicians (area under the curve: 0.769, 95% confidence interval: 0.709-0.820). These findings could potentially provide a noninvasive, high-throughput, and low-cost screening tool to facilitate embryo selection and achieve better outcomes. CONCLUSIONS: Our study underscores the AI model's ability to provide interpretable evidence for clinicians in assisted reproduction, highlighting its potential as a noninvasive, efficient, and cost-effective tool for improved embryo selection and enhanced IVF outcomes. The convergence of cutting-edge technology and reproductive medicine has opened new avenues for addressing infertility challenges and optimizing IVF success rates.

Perspectives of preimplantation genetic testing patients in Belgium on the ethics of polygenic embryo screening.

RESEARCH QUESTION: What are the perspectives of preimplantation genetic testing (PGT) patients in Belgium on the ethics of PGT for polygenic risk scoring (PGT-P)? DESIGN: In-depth interviews (18 in total, 10 couples, 8 women, n = 28) were performed with patients who had undergone treatment with PGT for monogenic/single-gene defects (PGT-M) or chromosomal structural rearrangements (PGT-SR) between 2017 and 2019 in Belgium. Participants were asked about their own experiences with PGT-M/SR and about their viewpoints on PGT-P, including their own interest and their ideas on its desirability, scope and consequences. Inductive content analysis was used to analyse the interviews. RESULTS: Participants stated that their experiences with PGT-M/SR had been physically, psychologically and practically difficult. Most participants stated that, partly because of these difficulties, they did not see the added value of knowing the risk scores of embryos via PGT-P. Many participants worried that PGT-P could lead to additional anxieties, responsibilities and complex choices in reproduction and parenthood. They argued that not everything should be controlled and felt that PGT-P, especially non-medical and broad screening, was going too far. With regards to the clinical implementation of PGT-P, participants in general preferred PGT-P to be limited to people with a serious polygenic family history and wanted embryo selection decisions to be made by healthcare professionals. CONCLUSIONS: This study shows that individuals with experience of PGT-M/SR saw PGT-P as different from PGT-M/SR. They had various ethical concerns with regards to PGT-P, especially regarding broadly offering PGT-P. These stakeholder viewpoints need to be considered regarding potential PGT-P implementation and guidelines.

Effect of freeze-thawing, cell collection, and laser irradiation cycles on mosaicism occurrence in preimplantation genetic testing for aneuploidy.

OBJECTIVE: In preimplantation genetic testing for aneuploidy, opinions regarding the handling of mosaic embryos vary. In this study, we aimed to investigate the effects of freeze-thawing, the number of cells obtained, and the number of laser irradiation cycles on the degree of embryonic mosaicism. STUDY DESIGN: This study was conducted in three parts. First, we classified specimens into the normal biopsy (control) (119 patients, 304 blastocysts) and thawed-biopsy (TB group) (26 patients, 72 blastocysts)) groups. The control and TB groups were then classified into three categories (euploidy, mosaic and aneuploidy) according to next-generation sequencing (NGS) results, and the number of cells collected and laser irradiation cycles were compared for each category. Subsequently, the effects of differences in the number of cells collected and laser irradiation cycles on NGS results were investigated in the control and TB groups. Finally, data on cell collection and laser irradiation cycles and NGS analysis results for the groups were compared. RESULTS: The TB group had a significantly higher incidence of chromosomal mosaicism than the control group. Neither the number of cells collected nor the laser irradiation cycles affected the percentage of chromosomal mosaicism. However, the freeze-thaw process increased the occurrence of mosaicism. CONCLUSIONS: This study showed that repeated freeze-thaw cycles increase the incidence of mosaicism, but the embryos are not aneuploid and are therefore suitable for transfer.

Multiple collapses of blastocysts after full blastocyst formation is an independent risk factor for aneuploidy - a study based on AI and manual validation.

BACKGROUND: The occurrence of blastocyst collapse may become an indicator of preimplantation embryo quality assessment. It has been reported that collapsing blastocysts can lead to higher rates of aneuploidy and poorer clinical outcomes, but more large-scale studies are needed to explore this relationship. This study explored the characteristics of blastocyst collapse identified and quantified by artificial intelligence and explored the associations between blastocyst collapse and embryo ploidy, morphological quality, and clinical outcomes. METHODS: This observational study included data from 3288 biopsied blastocysts in 1071 time-lapse preimplantation genetic testing cycles performed between January 2019 and February 2023 at a single academic fertility center. All transferred blastocysts are euploid blastocysts. The artificial intelligence recognized blastocyst collapse in time-lapse microscopy videos and then registered the collapsing times, and the start time, the recovery duration, the shrinkage percentage of each collapse. The effects of blastocyst collapse and embryo ploidy, pregnancy, live birth, miscarriage, and embryo quality were studied using available data from 1196 euploid embryos and 1300 aneuploid embryos. RESULTS: 5.6% of blastocysts collapsed at least once only before the full blastocyst formation (tB), 19.4% collapsed at least once only after tB, and 3.1% collapsed both before and after tB. Multiple collapses of blastocysts after tB (times ≥ 2) are associated with higher aneuploid rates (54.6%, P > 0.05; 70.5%, P < 0.001; 72.5%, P = 0.004; and 71.4%, P = 0.049 in blastocysts collapsed 1, 2, 3 or ≥ 4 times), which remained significant after adjustment for confounders (OR = 2.597, 95% CI 1.464-4.607, P = 0.001). Analysis of the aneuploid embryos showed a higher ratio of collapses and multiple collapses after tB in monosomies and embryos with subchromosomal deletion of segmental nature (P < 0.001). Blastocyst collapse was associated with delayed embryonic development and declined blastocyst quality. There is no significant difference in pregnancy and live birth rates between collapsing and non-collapsing blastocysts. CONCLUSIONS: Blastocyst collapse is common during blastocyst development. This study underlined that multiple blastocyst collapses after tB may be an independent risk factor for aneuploidy which should be taken into account by clinicians and embryologists when selecting blastocysts for transfer.

[Reproductive Health-Oriented Development in Assisted Reproductive Technologies]. / ä»¥ç”Ÿæ®–å¥åº·ä¸ºå¯¼å‘çš„è¾ åŠ©ç”Ÿæ®–æŠ€æœ¯å‘å±•.

Assisted reproductive technologies (ARTs) are core components of the field of reproductive medicine, encompassing multiple pivotal stages of early development from gamete maturation and fertilization to embryo development. Against the backdrop of a deteriorating trend of global decline in fertility rates, patients with infertility problems increasingly turn to ARTs to realize their dreams of parenthood. However, concomitant with this trend is a growing apprehension regarding the potential adverse effects of ARTs. Herein, we endeavor to discuss several common ARTs procedures utilized in clinical settings and the relevant cutting-edge advancements. The ARTs discussed in the article include in vitro fertilization (IVF), intracytoplasmic sperm injection (ICSI), biphasic in vitro maturation (biphasic IVM), frozen embryo transfer (FET), preimplantation genetic testing (PGT), non-invasive PGT (niPGT), etc. In addition, we reevaluated their roles within the broader context of assisted reproduction aimed at promoting reproductive health. Additionally, we will delve into the impact of ARTs on the reproductive health of the offspring. By prioritizing the reproductive well-being of both patients and their offspring, the ongoing development and improvement of ARTs to enhance their efficacy and safety will contribute significantly to the advancement of human reproductive health.

Challenges of Preimplantation Genetic Counselling in the Context of Cystic Fibrosis and Other CFTR-Related Disorders: A Monocentric Experience in a Cohort of 92 Couples.

Cystic fibrosis is a highly prevalent genetic disorder caused by biallelic pathogenic variants in the CFTR gene, causing an altered function of the exocrine glands and a subsequent spectrum of hypofunctional and degenerative manifestations. The increasing availability of carrier screening programmes, the enhanced life expectancy of patients due to improved treatment and care strategies and the development of more precise and affordable molecular diagnostic tools have prompted a rise in demand of prenatal diagnosis procedures for at-risk couples, including Preimplantation Genetic Testing (PGT). However, challenges remain: heterogeneity among screening programmes, nuances of variant interpretation and availability of novel treatments demand a considerate and knowledgeable approach to genetic counselling. In this work, we retrospectively evaluated the molecular data of 92 unselected couples who received a diagnosis of CFTR-related status and were referred to the genetics clinic at the University Hospital of Padua for genetic counselling on eligibility for PGT. A total of 50 couples were considered eligible for the procedure based on risk of transmitting biallelic pathogenic variants. We report and discuss our experience with this case series in the context of the Italian medical care system and present an overview of the most relevant issues regarding genetic counselling for PGT in CFTR-related disorders.

A healthy live birth after mosaic blastocyst transfer in preimplantation genetic testing for GATA1-related cytopenia combined with HLA matching.

BACKGROUND: GATA1-related cytopenia (GRC) is characterized by thrombocytopaenia and/or anaemia ranging from mild to severe. Haematopoietic stem cell transplantation (HSCT) is a healing therapeutic choice for GRC patients. We identified a novel pathogenic variant (GATA1: c.1019delG) in a boy with GATA1-related cytopenia. Then we performed preimplantation genetic testing (PGT) in this GRC family. After a mosaic embryo transfered, a healthy and HLA-compatible with the proband baby was delivered. CASE PRESENTATION: The proband is a 6-year-old boy who was diagnosed to have transfusion-dependent anaemia since 3 year old. Whole-exome sequencing (WES) showed that the proband has a hemizygous variant c.1019delG in GATA1, which is inherited from his mother. His parents decided to undergo PGT to have a health and HLA-compatible offspring. After whole genome amplification (WGA) of biopsied trophectoderm (TE) cells, next generation sequencing (NGS)-based PGT was preformed to analyse embryos on chromosomal aneuploidy, target mutation and HLA typing. There were 3 embryos HLA-matched to the proband. The genotypes of the 3 embryos were heterozygous variant, hemizygous variant, normal respectively. After a heterozygous, mosaic partial trisomy (chr)16, and HLA-matched embryo transfer, a healthy baby was delivered and whose HSCT is compatible with the proband. CONCLUSIONS: NGS-based PGT-HLA is a valuable procedure for the treatment of GATA1-related cytopenia caused by GATA1 variants, or other haematological disorders, oncological and immunological diseases. Furthermore, our study reconfirms that mosaic embryos transfer would bring healthy offspring.

Development of pre-implantation genetic testing protocol for monogenic disorders (PGT-M) of Hb H disease.

Hb H disease is the most severe form of α-thalassemia compatible with post-natal life. Compound heterozygous α0-thalassemia- SEA deletion/α+-thalassemia- 3.7kb deletion is the commonest cause of Hb H disease in Thailand. Preimplantation genetics testing for monogenic disorders (PGT-M) is an alternative for couples at risk of the disorder to begin a pregnancy with a healthy baby. This study aims to develop a novel PCR protocol for PGT-M of Hb H disease- SEA/-3.7kb using multiplex fluorescent PCR. A novel set of primers for α+-thalassemia- 3.7kb deletion was developed and tested. The PCR protocol for α0-thalassemia- SEA deletion was combined for Hb H disease- SEA/-3.7kb genotyping. The PCR protocols were applied to genomic DNA extracted from subjects with different thalassemia genotypes and on whole genome amplification (WGA) products from clinical PGT-M cycles of the families at risk of Hb Bart's. The results were compared and discussed. The results showed three PCR products from α+-thalassemia- 3.7kb primer set, and three from α0thalassemiaSEA primer set. The results were consistent with the known thalassemia genotypes. The novel -α3.7 primers protocol was also tested on 37 WGA products from clinical PGT-M cycles giving accurate genotyping results and a satisfying amplification efficiency with the ADO rates of 2.7%, 0%, and 0% for HBA2, HBA1, and internal control fragments, respectively. This novel PCR protocol can precisely distinguish Hb H disease- SEA/-3.7kb from other genotypes. Additionally, this is the first PCR protocol for Hb H disease- SEA/-3.7kb which is optimal for PGT-M.

A study on methods for preimplantation genetic testing (PGT) on in vivo- and in vitro-produced equine embryos, with emphasis on embryonic sex determination.

Two methods for preimplantation genetic testing (PGT) have been described for equine embryos: trophoblast cell biopsy (TCB) or blastocoele fluid aspiration (BFA). While TCB is widely applied for both in vivo- and in vitro-produced embryos, BFA has been mostly utilized for in vivo-produced embryos. Alternative methods for PGT, including analysis of cell-free DNA (CFD) in the medium where in vitro-produced embryos are cultured, have been reported in humans but not for equine embryos. In Experiment 1, in vivo- (n = 10) and in vitro-produced (n = 13) equine embryos were subjected to BFA, cultured for 24 h, then subjected to TCB, and cultured for additional 24 h. No detrimental effect on embryonic diameter or re-expansion rates was observed for either embryo group (P > 0.05). In Experiment 2, the concordance (i.e., agreement on detecting the same embryonic sex using two techniques) among BFA, TCB, and the whole embryo (Whole) was studied by detecting the sex-determining region Y (SRY) or testis-specific y-encoded protein 1 (TSPY) (Y-chromosome), and androgen receptor (AR; X-chromosome) genes using PCR. Overall, a higher concordance for detecting embryonic sex was observed among techniques for in vivo-produced embryos (67-100 %; n = 14 embryos) than for in vitro-produced embryos (31-92 %; n = 13 embryos). The concordance between sample types increased when utilizing TSPY (77-100 %) instead of SRY (31-100 %) as target gene. In Experiment 3, CFD analysis was performed on in vitro-produced embryos to determine embryonic sex via PCR (SRY [Y-chromosome] and amelogenin - AMEL [X- and Y-chromosomes]). Overall, CFD was detected in all medium samples, and the concordance between CFD sample and the whole embryo was 60 % when utilizing SRY and AMEL genes. In conclusion, equine embryos can be subjected to two biopsy procedures (24 h apart) without apparent detrimental effects on embryonic size. For in vivo-, but not for in vitro-produced equine embryos, BFA can be considered a potential alternative to TCB for PGT. Finally, CFD can be further explored as a non-invasive method for PGT in in vitro produced equine embryos.

Concurrent Preimplantation Genetic Testing and Competence Assessment of Human Embryos by Transcriptome Sequencing.

Preimplantation genetic testing (PGT) can minimize the risk of birth defects. However, the accuracy and applicability of routine PGT is confounded by uneven genome coverage and high allele drop-out rate from existing single-cell whole genome amplification methods. Here, a method to diagnose genetic mutations and concurrently evaluate embryo competence by leveraging the abundant mRNA transcript copies present in trophectoderm cells is developed. The feasibility of the method is confirmed with 19 donated blastocysts. Next, the method is applied to 82 embryos from 26 families with monogenic defects for simultaneous mutation detection and competence assessment. The accuracy rate of direct mutation detection is up to 95%, which is significantly higher than DNA-based method. Meanwhile, this approach correctly predicted seven out of eight (87.5%) embryos that failed to implant. Of six embryos that are predicted to implant successfully, four met such expectations (66.7%). Notably, this method is superior at conditions for mutation detection that are challenging when using DNA-based PGT, such as when detecting pathogenic genes with a high de novo rate, multiple pseudogenes, or an abnormal expansion of CAG trinucleotide repeats. Taken together, this study establishes the feasibility of an RNA-based PGT that is also informative for assessing implantation competence.

Attitudes toward genetic testing, family planning and preimplantation genetic testing in families with a germline CDKN2A pathogenic variant.

Individuals with a germline CDKN2A pathogenic variant (PV) have a highly increased life time risk of melanoma and pancreatic cancer. This cross-sectional study assessed the attitudes among toward genetic testing, family planning, and preimplantation genetic testing (PGT) in confirmed CDKN2A PV carriers and individuals with a 50% risk of the PV (at-risk carriers) using of a one-time questionnaire.A total of 537 individuals were screened for eligibility, of whom 208 of 366 (57%) confirmed carriers (56% female, median age 54 years [IQR 46-63]) and 39 of 171 (23%) at-risk carriers (59% female, median age of 26 years [IQR 22-32]) participated in the study. Primary motivations for genetic testing were to gain control over their personal and children's cancer risk, as well as increasing cancer surveillance practices. In contrast, concerns about obtaining a mortgage and life insurance were frequently cited as reasons for postponing genetic testing. Family planning decisions remained largely unaffected in both confirmed and at-risk carriers; however, the majority of confirmed carriers were still unaware of their familial or personal cancer risk when starting a family. More than 60% of the participants were unfamiliar with PGT and only a minority (19% of confirmed carriers and 10% of at-risk carriers) would be open to considering PGT as a reproductive option. This study found different attitudes toward genetic testing, family planning, and PGT among individuals affected by the CDKN2A PV. Understanding these different attitudes can help clinicians to address the complexities surrounding these issues, especially for younger individuals facing difficult decisions about the timing of genetic testing, family planning, and the potential use of assisted reproductive options.

Best quality vs. sex selection - an analysis of embryo selection preferences for patients undergoing preimplantation genetic testing for aneuploidy over a 10-year period.

PURPOSE: Investigate patient preferences in embryo selection for transfer regarding quality versus sex in IVF/ICSI cycles with PGT-A and assess associated clinical implications. METHODS: Retrospective cohort study at a university fertility practice from January 2012 to December 2021. Included were patients undergoing single frozen euploid transfers with at least one embryo of each sex available. Primary outcomes were preference for embryo selection (quality vs. sex) and sex preference (male vs. female). Trends over 10 years were evaluated and clinical outcomes, including clinical pregnancy rate (CPR), sustained implantation rate (SIR), and live birth rate (LBR), were compared. RESULTS: A total of 5,145 embryo transfer cycles were included; 54.5% chose the best-quality embryo, while 45.5% selected based on sex. Among those choosing based on sex, 56.5% chose male embryos and 43.5% chose female. Preference for quality remained consistent over the decade (p = 0.30), while male embryos were consistently favored (p = 0.64). Best-quality embryos had higher grades (p < 0.001). Clinical outcomes were similar between groups (CPR: 74.4% vs. 71.9%, p = 0.05; SIR: 64.9% vs. 63.4%, p = 0.26; LBR: 58.8% vs. 56.7%, p = 0.13), and between male and female embryo selections. CONCLUSIONS: Sex selection remains common, with 45.5% selecting embryos based on sex, predominantly favoring males. This trend persisted over 10 years, with comparable clinical outcomes regardless of selection criteria.

The Blastulation Rate Is Negatively Associated With Euploid Rate.

OBJECTIVES: To study the association between the blastulation rate, the presence of 1 pronucleus (1PN) zygotes, and the ploidy of the cohort of blastocysts. METHODS: A cross-sectional study using the existing databases of 2 university fertility centres in Canada. We included 345 cycles from 235 couples who underwent next-generation sequencing preimplantation genetic testing for the detection of aneuploidy in the study. RESULTS: A total of 1456 blastocysts were biopsied. In multivariate analysis, only female age and the number of 1PN/2PN embryos showed a negative association with euploid ratio. Surprisingly, when the analysis was limited to cycles with no delayed blastulation, the blastulation rate was also negatively associated with the euploid ratio. CONCLUSIONS: This study sheds some light on the stages of early embryo development. Further study on the mechanisms governing embryo development and the different cell cycle checkpoints in embryo development is warranted.