Preimplantation Genetic Diagnosis Research Articles

The frontier of precision medicine: application of single-cell multi-omics in preimplantation genetic diagnosis.

The advent of single-cell multi-omics technologies has revolutionized the landscape of preimplantation genetic diagnosis (PGD), offering unprecedented insights into the genetic, transcriptomic, and proteomic profiles of individual cells in early-stage embryos. This breakthrough holds the promise of enhancing the accuracy, efficiency, and scope of PGD, thereby significantly improving outcomes in assisted reproductive technologies (ARTs) and genetic disease prevention. This review provides a comprehensive overview of the importance of PGD in the context of precision medicine and elucidates how single-cell multi-omics technologies have transformed this field. We begin with a brief history of PGD, highlighting its evolution and application in detecting genetic disorders and facilitating ART. Subsequently, we delve into the principles, methodologies, and applications of single-cell genomics, transcriptomics, and proteomics in PGD, emphasizing their role in improving diagnostic precision and efficiency. Furthermore, we review significant recent advances within this domain, including key experimental designs, findings, and their implications for PGD practices. The advantages and limitations of these studies are analyzed to assess their potential impact on the future development of PGD technologies. Looking forward, we discuss the emerging research directions and challenges, focusing on technological advancements, new application areas, and strategies to overcome existing limitations. In conclusion, this review underscores the pivotal role of single-cell multi-omics in PGD, highlighting its potential to drive the progress of precision medicine and personalized treatment strategies, thereby marking a new era in reproductive genetics and healthcare.

Pregnancy and inherited aortopathies: insights from specialized care

Abstract Background Genetic counselling, pregnancy follow-up and delivery planning in women with aortopathy should be conducted by a multidisciplinary team in an expert centre. Purpose Our objective was to analyze the impact of pregnancy on aortopathy outcomes in a centre specialized in inherited cardiovascular conditions in the United Kingdom. Methods All women with aortopathy referred to our centre due to pregnancy between 2016 and 2023 were retrospectively analyzed. Demographic, clinical, obstetric, genetic, electrocardiographic, and multimodal imaging variables were collected. Results Between 2016 and 2023, 22 women with aortopathy were referred to our centre for pregnancy follow-up and delivery planning. Among them, 14 (63.6%) had Marfan syndrome (FBN1+), 4 (18.2%) family history of aortic dissection without known genetic mutations, 2 (9.1%) bicuspid aortic valve with aortic dilation and 2 (9.1%) Loeys-Dietz syndrome (SMAD3+). The average number of births per patient was 1.55. Among those with an identified mutation, 4 (25%) underwent preimplantation diagnosis. In total, there were 40 pregnancies resulting in 34 live births. 15 women (68.2%) had an aortic root dimension <40mm and 7 (31.8%) ≥40mm. There were 4 (26.7%) and 5 (71.4%) patients in each group under beta-blockers. Out of the total 40 pregnancies, there were 6 (15%) miscarriages, all in the first trimester, 2 (5%) cases of preeclampsia and 1 (2.5%) case of gestational diabetes. There were no statistically significant differences between these complications and the aortic root dimension (8 (20%) <40mm vs 1 (2.5%) ≥40mm; p = 0.160) or the absence or presence of treatment (3 (7.5%) vs 6 (15%); p = 0.769). Regarding delivery, 16 patients (72.7%) underwent vaginal delivery and 6 (27.3%) has a caesarean section. After delivery, there were 6 cases of major bleeding, 4 (10%) after vaginal delivery and 2 (5%) after caesarean (p=0.376). One patient had a subacute type A aortic dissection in the context of Marfan syndrome with an aortic root of 40mm before pregnancy. No significant differences were observed in the occurrence of these complications based on the aortic root dimension (5 (12.5%) <40mm vs 2 (5%) ≥40mm; p = 0.928) or the absence or presence of treatment (2 (5%) vs 5 (12.5%) respectively; p = 0.591). 100% of the patients survived pregnancy and the postpartum period. Conclusions Our findings underscore the complexity of managing aortopathy during pregnancy. This highlights the importance of a comprehensive risk assessment for precise pregnancy and delivery planning.

Steady morphokinetic progression is an independent predictor of live birth: a descriptive reference for euploid embryos.

Can modelling the longitudinal morphokinetic pattern of euploid embryos during time-lapse monitoring (TLM) be helpful for selecting embryos with the highest live birth potential? Longitudinal reference ranges of morphokinetic development of euploid embryos have been identified, and embryos with steadier progression during TLM are associated with higher chances of live birth. TLM imaging is increasingly adopted by fertility clinics as an attempt to improve the ability of selecting embryos with the highest potential for implantation. Many markers of embryonic morphokinetics have been incorporated into decision algorithms for embryo (de)selection. However, longitudinal changes during this temporal process, and the impact of such changes on embryonic competence remain unknown. Aiming to model the reference ranges of morphokinetic development of euploid embryos and using it as a single longitudinal trajectory might provide an additive value to the blastocyst morphological grade in identifying highly competent embryos. This observational, retrospective cohort study was performed in a single IVF clinic between October 2017 and June 2021 and included only autologous single euploid frozen embryo transfers (seFET). Reference ranges were developed from [hours post-insemination (hpi)] of the standard morphokinetic parameters of euploid embryos assessed as tPB2, tPNa, tPNf, t2-t9, tSC, tM, tSB, and tB. Variance in morphokinetic patterns was measured and reported as morphokinetic variance score (MVS). Nuclear errors (micronucleation, binucleation, and multinucleation) were annotated when present in at least one blastomere at the two- or four-cell stages. The blastocyst grade of expansion, trophectoderm (TE), and inner cell mass (ICM) were assessed immediately before biopsy using Gardner's criteria. Pre-implantation genetic diagnosis for aneuploidy (PGT-A) was performed by next-generation sequencing. All euploid embryos were singly transferred in a frozen transferred cycle and outcomes were assessed as live birth, pregnancy loss, or not pregnant. Association of MVS with live birth was investigated with regression analyses. TLM data from 340 seFET blastocysts were included in the study, of which 189 (55.6%) resulted in a live birth. The median time for euploid embryos to reach blastulation was 109.9 hpi (95% CI: 98.8-121.0 hpi). The MVS was calculated from the variance in time taken for the embryo to reach all morphokinetic points and reflects the total morphokinetic variability it exhibits during its development. Embryos with more erratic kinetics, i.e. higher morphokinetic variance, had higher rates of pregnancy loss (P = 0.004) and no pregnancy (P < 0.001) compared to embryos with steadier morphokinetic patterns. In the multivariable analysis adjusting for ICM, TE grade, presence of nuclear errors, and time of blastulation, MVS was independently associated with live birth (odds ratio [OR]: 0.62, 95% CI: 0.46-0.84, P = 0.002) along with ICM quality. Live birth rate of embryos with the same ICM grading but different morphokinetic variance patterns differed significantly. Live birth rates of embryos exhibiting low MVS with ICM grades A, B, and C were 85%, 76%, and 67%, respectively. However, ICM grades A, B, and C embryos with high MVS had live birth rates of 65%, 48%, and 21% (P < 0.001). The addition of the MVS to embryo morphology score (ICM and TE grading) significantly improved the model's AUC value (0.67 vs 0.62, P = 0.015) and this finding persisted through repeat cross-validation (0.64 ± 0.08 vs 0.60 ± 0.07, P < 0.001). The exclusion of IVF cases limits, for now, the utility of the model to only ICSI-derived embryos. The utility of these reference ranges and the association of MVS with various clinical outcomes should be further investigated. We have developed reference ranges for morphokinetic development of euploid embryos and a marker for measuring total morphokinetic variability exhibited by developed blastocysts. Longitudinal assessment of embryonic morphokinetics rather than static time points may provide more insight about which embryos have higher live birth potential. The developed reference ranges and MVS show an association with live birth that is independent of known morphological factors and could emerge as a valuable tool in prioritizing embryos for transfer. This study received no external funding. The authors declare no conflicting interests. N/A.

9234 A Case of Gigantism Due to An AIP Gene Mutation: Implications for Genetic Counseling and Preimplantation Genetic Diagnosis

Abstract Disclosure: M. Serebro: None. M. Yacobi Bach: None. N. Even Zohar: None. A. Reches: None. Y. Greenman: None. From the age of 17 the patient experienced accelerated growth, facial changes, and enlargement of hands and feet. Despite having a sister already diagnosed with acromegaly, the patient did not undergo endocrine evaluation until the age of 25, when he was diagnosed with gigantism. Magnetic resonance imaging (MRI) revealed the presence of a macroadenoma with suprasellar extension reaching the optic chiasm. The patient underwent transsphenoidal surgery and excision of a sparsely granulated somatotroph adenoma that was Pit1 positive and exhibited weak immunostaining for growth hormone (GH). Insulin-like growth factor 1 (IGF1) levels normalized after surgery, and a fasting GH level was 0.6 ng/ml. Given the family history and the early onset of the disease, genetic counseling was recommended. Genetic testing revealed a heterozygote likely pathogenic variant in the AIP gene (c.713G&amp;gt;A). This finding prompted the recommendation for genetic testing of all first-degree relatives. The patient and his wife, who desired to have children, were referred to obstetrics and gynecology for pregestational diagnosis. Family segregation testing indicated that the patient's mother also carried the same AIP variant. Preimplantation genetic diagnosis (PGD) was conducted, revealing that 11 out of 20 embryos were carriers of the AIP variant. The non-carrier embryos were selected for implantation, and pregnancy was achieved. Efforts to inform first-degree relatives about the importance of genetic testing were undertaken, and one sister was tested and found to be healthy. Other siblings have not yet been tested. The patient's mother, identified as an AIP variant carrier, underwent endocrine laboratory tests that showed GH, IGF1, and prolactin levels within the normal range. Brain MRI did not reveal a pituitary lesion.This case highlights the significance of genetic counseling and testing in patients with familial endocrine disorders, the potential implications of a molecular diagnosis of a pathogenic AIP variant, and the challenges faced in reproductive decision-making for affected individuals and their families. Presentation: 6/3/2024

Feasibility of preimplantation genetic testing for aneuploidy on frozen-thawed embryos following conventional IVF insemination.

Intracytoplasmic sperm injection (ICSI) is commonly employed in preimplantation genetic testing (PGT) to minimize the risk of foreign sperm DNA contamination. Cryopreserved embryos from patients with recurrent miscarriage or repeated implantation failure, who have undergone conventional in vitro fertilization (IVF), can be thawed and biopsied for PGT. Therefore, we aimed to assess the accuracy and effectiveness of preimplantation genetic testing for aneuploidy (PGT-A) on frozen embryos using conventional IVF (c-IVF) insemination methods. From January 2021 to November 2023, our center conducted 107 thawed cryopreserved embryo biopsy cycles to screen for PGT-A. Among them, 58 cycles used c-IVF insemination, and 49 used ICSI insemination. Basic patient clinical information, laboratory data, PGT test results, and clinical outcome data were collected. To minimize the confounding effects of patient characteristics and embryo quality on PGT-A outcomes, clinical outcomes, and contamination assessment, these variables were included in the analysis. We then evaluated the blastocyst euploidy rate, clinical outcomes, and accuracy of PGT-A results between the two groups and analyzed potential contamination in the c-IVF insemination group. A total of 320 blastocysts underwent PGT-A testing, with 179 blastocysts from c-IVF insemination and 141 from ICSI insemination. Considering participants' baseline characteristics and embryological outcomes, no significant differences were found between the two groups regarding infertility type, average age, body mass index, percentage of PGT-A indications, or quality of embryonic development. Regarding PGT-A results, all 320 biopsy samples were successfully analyzed, showing no statistical variance in chromosomal euploidy, abnormality, or mosaicism rates between the two insemination methods. No parental contamination was detected in the c-IVF insemination group. When assessing clinical outcomes, parameters such as biochemical pregnancy, clinical pregnancy, and miscarriage rates did not exhibit significant discrepancies between the two groups, and no misdiagnoses were reported during the study period. Embryo transfer and PGT-A results are not affected by potential parental contamination in frozen-thawed embryos conceived via c-IVF. PGT-A guided embryo transfer in thawed embryos conceived by c-IVF is a viable and clinically effective approach.

Single Nucleotide Polymorphism array analysis for fetuses from balanced translocation carriers at the second trimester

Prenatal diagnosis is crucial for pregnancies from couples with a carrier of a balanced translocation. We retrospectively reviewed 195 pregnancies from 189 couples with a balanced translocation carrier. Of these, 126 were from natural conception, while 69 were conceived through assisted reproductive technology (ART) with preimplantation genetic diagnosis (PGD). Both single nucleotide polymorphism (SNP) array analysis and conventional karyotyping were conducted on all pregnancies, and karyotype-visible imbalances and pathogenic/likely pathogenic copy number variations (CNVs) were categorized as clinically significant abnormalities. In natural conception group, couples with a female carrier experiencing more than two miscarriages accounted for 30.2%, significantly higher than the 14.0% in male carrier couples (p<0.05). In the PGD group, similar difference was observed between female and male carrier couples (p<0.05). In the natural pregnancies, SNP array analysis yielded additional 12 cases of CNVs, including two cases of pathogenic (P)/likely pathogenic (LP) aberrations, four variants with uncertain significance (VUS), and six likely benign variants. Only two CNVs were found to be associated with translocation breakpoints, which were finally confirmed to be of parental inheritance. In the PGD pregnancies, two cases of VUS unrelated to the translocation breakpoints were revealed. Taken together, repeated miscarriage was more frequently observed in couples where the carrier was female than male. Conventional SNP array analysis in prenatal diagnosis indicated insufficient evidence to support the notion that balanced translocations increase the likelihood of fetuses having clinically significant CNVs.

Molecular autopsy for fetal structural anomaly: diagnostic and clinical utility of multidisciplinary team approach.

In the West Midlands regional genetics service, cases of perinatal death with a possible genetic diagnosis are evaluated by the perinatal pathology genetic multidisciplinary team (MDT). The MDT assesses autopsy findings and suggests appropriate genomic assessment. The objective of this retrospective service evaluation was to determine the clinical utility of the MDT in assessing perinatal deaths associated with structural anomaly. This is the first evaluation since the introduction of whole-genome and whole-exome sequencing in routine clinical care. This was a retrospective service evaluation including all cases of perinatal death with an associated structural anomaly and suspected genetic etiology that underwent perinatal MDT assessment between January and December 2021. All cases received a full or partial postmortem examination and at least a chromosomal microarray analysis. Demographic characteristics, phenotype, genotype, MDT recommendations, diagnoses, outcomes and impact of postmortem analysis and genetic testing data were collected from patient case notes. Overall, 123 cases were discussed at the MDT meetings in 2021. Genetic evaluation was recommended in 84 cases and accepted in 64 cases. A range of genetic tests were requested according to indication and availability. Thirty diagnoses were made in 29 cases from 26 unrelated families. The diagnostic yield was 24% (29/123) in all cases or 45% (29/64) in cases with a suspected genetic diagnosis who underwent genetic testing. Postmortem examination provided clinically actionable phenotypic data in 79% of cases. A genetic diagnosis enabled accurate recurrence risk counseling and provision of appropriate follow-up, including prenatal testing and preimplantation diagnosis for patients with inherited conditions. Genomic testing was a clinically useful addition to (but not a substitute for) postmortem examination in cases of perinatal death associated with structural anomaly. The MDT approach helped assess cases and plan appropriate follow-up. Expedited whole-genome sequencing or panel-agnostic analysis were most appropriate for heterogeneous presentations. This broad approach can also expand knowledge of prenatal phenotypes and detect novel disease genes, and should be a priority in future research. © 2024 International Society of Ultrasound in Obstetrics and Gynecology.

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

PurposeTo 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.MethodsThe 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.ResultsThroughout 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.ConclusionsThe 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.

Development of a PCR-based method to identify fetal sex during IVF cycles.

One of the most recognizable cases of preimplantation genetic diagnosis (PGD) is X-linked diseases. Diagnosis of fetal sex is essential for couples who are known to be at risk of some X-linked disorders. The objective of this study was to discriminate between female (XX) and male (XY) embryos by detecting sex chromosomes-specific sequences in spent culture medium and comparing these results to PGD/CGH array results. It may open new window for the development of a non-invasive PGD method. 120 Embryo's spent media from Day 3 and Day 5 embryos were collected. Modified phenol-chloroform solution was used for DNA extraction from spent media. Sex determination was performed using SRY, TSPY and AMELOGENIN evaluation through quantitative polymerase chain reaction (q-PCR) method. IBM SPSS and MedCalc were used for statistical analyses to compare sex determination of embryos by spent medium with PGD/CGH array results. Culture time was demonstrated to increase the DNA amount among day 5 embryos culture medium samples. Non-invasive PGD by means of spent culture medium gave a sensitivity, specificity, positive predictive value and negative predictive value of 100% for sex determination. Results of sex determination using spent medium by q-PCR were consistent with the results of PGD/CGH array. Improvements in cell-free DNA extraction and PCR amplification procedures provide us an effective method to perform a PGD test without biopsy in the future, especially about X-linked diseases.

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.

Family Planning in Genetic Optic Atrophies in Israel, a Case Series and a Discussion of Ethical Considerations.

Patients with genetic optic atrophies must navigate all stages of life with their visual impairment, including the important milestone of family planning. Advances in genetic testing now allows physicians and affected families to consider medical help with the aim of preventing blindness through preconception, preimplantation, and perinatal methods. This case series presents 4 patients with different genetic optic atrophies (Leber hereditary optic neuropathy [LHON], autosomal dominant optic atrophy, Wolfram syndrome, and papillorenal syndrome) who were followed by the Neuro-Ophthalmology Unit at a tertiary medical center between 2010 and 2023 and were of child-bearing age. The aim of this study was to increase understanding in family planning options for patients with optic atrophies, raise awareness of the solutions available, and provide guidance for clinicians to support their patients. Advances in medicine, genetics, and medical technology allow multidisciplinary teams to assist patients in fulfilling their desire for a genetically healthy offspring. Customized solutions can be designed to meet the specific challenges posed by each type of genetic optic atrophy. The solutions proposed in this series are based on genetic testing done in the parents, which then allows to plan medical and genetic intervention individually. The solutions opted for in this series range from the decision to not have another child until PGD (Preimplantation genetic diagnosis). We describe how genetic advancements have made it possible for patients with the 4 most common hereditary optic atrophies to fulfill their wish to have children without visually threatening genetic mutations. We also review the recent literature on the penetrance of optic atrophy in OA-mutation carriers and raise 2 significant ethical considerations: the reduction of a future life to a non-life-threatening impairment and that of public expenditure for non-life-threatening conditions.

À propos du diagnostic préimplantatoire des aneuploïdies (DPI-A). Les inconvénients de mots inappropriés et d’une réglementation inadaptée

Finding biological markers to assess the ability of embryos to develop is a major objective of research being carried out to improve the results of in vitro fertilization (IVF). Among these markers, the chromosomal content of embryonic cells has been the subject of numerous studies to identify euploid embryos which will be transferred as a priority into the uterus. In France, a clinical research program funded by the Ministry of Health was undertaken to evaluate the benefit of this strategy when IVF is carried out in women aged 35 to 41. This program was interrupted because its authorization was canceled by a court on the grounds that the search for aneuploidy in the embryo is a preimplantation genetic diagnosis (PGD) and that this type of analysis is not provided for in the governing law. The law says in fact that a PGD can only be carried out if “has been previously and precisely identified, in one of the parents or one of their immediate ascendants in the case of a seriously disabling illness, with late revelation and prematurely endangering the vital prognosis”. But the chromosomal analysis provided for in the research protocol is not a diagnosis, it is a test which identifies the embryos most capable of developing. Like all other techniques with the same goal, this test is not the responsibility of the law but of the good practice guide. Studies seeking to improve the results of IVF for the benefit of couples who use it should not be prevented.

Maternal Placental Growth Factor (PlGF) levels, sonographic placental parameters, and outcomes of IVF pregnancies with and without embryo trophectoderm biopsy.

In vitro fertilization (IVF) is associated with abnormal trophoblast invasion and resultant decreased levels of circulating placental biomarkers such as placental growth factor (PlGF).Our objective was to evaluate maternal serum levels of second/third trimester PlGF, sonographic placental parameters, and clinical outcomes among IVF frozen embryo transfer (FET) pregnancies with and without embryo trophectoderm biopsy. This was a retrospective study of pregnant patients who conceived using a single frozen embryo transfer (FET) and gave birth between 30 January 2018 and 31 May 2021. We compared PlGF levels, sonographic placental parameters, and clinical outcomes between FET with biopsy and FET without biopsy groups. The median PlGF level was 614.5 pg/mL (IQR 406-1020) for FET pregnancies with biopsy, and 717.0 pg/mL (IQR 552-1215) for FET pregnancies without biopsy. The adjusted mean difference was 190.9 pg/mL lower in the FET biopsy group (95% CI, -410.6, 28.8; p = 0.088). There were no statistically significant differences in placental parameters or clinical pregnancy outcomes. This exploratory study demonstrated a possible trend toward lower maternal serum PlGF in the pregnancies conceived with FET using a biopsied embryo. Further investigation is warranted into the potential placental health effects of trophectoderm biopsy.

The sufficiency of genetic diagnosis in managing patients with inborn errors of immunity during prenatal care and childbearing.

Individuals with inborn errors of immunity face challenges in fertility, pregnancy, and genetic disorder transmission. Prenatal genetic counseling is crucial, especially in tribal societies with consanguineous unions. Ten families with confirmed inborn errors of immunity were studied, revealing diverse pregnancy decisions: An architect with autosomal dominant STAT-1 gain of function underwent prenatal diagnosis despite initial plans for preimplantation genetic diagnosis. In a consanguineous family, two children died from leukocyte adhesion deficiency type 1 because the father refused prenatal diagnosis. First cousins opted against terminating the second pregnancy, resulting in two children affected by Bruton disease. Another consanguineous couple, with two children afflicted by ataxia-telangiectasia, chose oocyte donation for their third child, ensuring a healthy birth. Recurrent pregnancy loss was observed in a mother subsequently diagnosed with ZAP70 deficiency. A mother with Wiskott-Aldrich syndrome child opted for in vitro fertilization, leading to a healthy birth post-prenatal diagnosis. A misdiagnosis of anaplastic anemia occurred in a family with multiple instances of Wiskott-Aldrich syndrome. A leukocyte adhesion deficiency type 1 case led to parental dissolution due to the father's refusal to acknowledge the condition. In a non-consanguineous couple, the father's diagnosis of TACI deficiency influenced the mother's decision to discontinue pregnancy post-prenatal diagnosis. Genetic diagnosis alone cannot optimize prenatal care for immune dysregulation disorders. Various factors, including patient education, societal norms, ethics, and economics, impact pregnancy decisions. Clinical immunologists must integrate these elements into guidance strategies to enhance patient outcomes.

New Variants Identified by Next-Generation Sequencing in Polycystic Kidney Disease Patients.

Polycystic kidney disease (PKD) is a common inherited disease characterized by multiple cysts in kidneys and various extra renal manifestations. Molecular diagnosis plays a crucial role in confirming both the clinical diagnosis and preimplantation genetic diagnosis furthermore, selecting appropriate treatment options. This study aimed to expand the understanding of genetic mutations in patients with polycystic kidney disease and to improve the management of patients. The study included 92 patients with a clinical diagnosis of PKD based on renal ultrasound criteria. Targeted next-generation sequencing was performed using a custom panel kit. Of the 92 patients included in the study, pathogenic/likely pathogenic variants of the PKD1, PKD2 genes were detected in 37 patients (40.2%), while 8 patients (8.6%) had variants with uncertain clinical significance. After the additional assessment of pathogenic/likely pathogenic variants, it was found that 15 of the variants in PKD1 and 2 of the variants in PKD2 have not been reported in the literature previously. Additionally, pathogenic variants, 5 of which were novel, have been identified in different genes in 8 patients. This study presented the largest patient cohort conducted in Turkey. These findings were significant in expanding our understanding of the genetic variations associated with polycystic kidney disease. The study contrıbuted the literature data on polycystic kidney disease by reporting important findings that could pave the way for further investigations in the diagnosis, treatment, and management of the affected patients.

P-124 Machine learning scores reveal mosaic embryo quality as the key predictor for live births, outperforming mosaicism characteristics in ART success

Abstract Study question Is it possible to score the mosaic embryos in order to prioritise them according to their pregnancy potencial? Summary answer The machine-learning score underscores the crucial role of embryo quality over mosaicism, prioritizing high-quality, day-5 biopsied embryos for enhanced live birth outcomes in ART. What is known already After embryo biopsy and PGT-A analysis, embryos can be classified as euploid, aneuploid, or mosaic, wherein a combination of chromosomally normal and altered cells is detected. The transfer of mosaic embryos has been controversial, but currently, the recommendations of international scientific societies, including ESHRE, advocate for their transfer. In case of pregnancy, a prenatal test is advised to rule out any chromosomal aberration. The non-transfer of these embryos reduces ART success chances, as chromosomally normal children may be born from them. Study design, size, duration The study design is observational and retrospective. A total of 8346 embryos from 2583 PGT-A cycles were considered (January-2017 to January-2023). Only transferred mosaic embryos (n = 263) were included in the study. The trophectoderm biopsies of D5, D6, or D7 blastocysts were analyzed by NGS using the Illumina platform (VeriSeq Illumina®, San Diego, CA, USA). The biopsied embryos were vitrified and transferred in a subsequent cycle. Participants/materials, setting, methods The primary indications for PGT-A included advanced maternal age, abnormal sperm FISH, and recurrent miscarriage or implantation failure. Clinical outcomes, along with variables related to progenitors, embryos, biopsy, ovarian stimulation, and adjuvant treatments, were recorded in a database. Machine learning models were conducted using R statistical software (v.4.3.1), and the ‘Caret’ library was utilized to implement different algorithms. ‘Autoscore,’ as an interpretable scoring tool, combined machine learning and regression modeling. Main results and the role of chance The mean maternal and paternal ages were 34.95±6.38 and 39.13±6.74 years. Biopsies were predominantly performed on day 5 (61%). Additionally, 44% of embryos were of A quality, and 49% were of B quality. Regarding clinical outcomes for mosaic embryos, biochemical, clinical pregnancy, and live birth rates were 51%, 41%, and 36%, respectively. The features used to generate the score included those related to embryo quality, biopsy, and mosaicism. Through 10-fold cross-validation, the study identified top-ranking predictors maximizing the AUC of the predictive model. The highest weight in the score was assigned to embryo quality (42%) and day of biopsy (34%), with lower weights for factors related to mosaicism: monosomy and/or trisomy (18%) and mosaicism level (6%). The machine-learning score indicates that, in transferring embryos with mosaicism, priority should be given to high-quality embryos biopsied on day 5. The type and level of mosaicism have a minor impact on the embryo’s gestational capacity, becoming relevant only when prioritizing embryos of identical quality. In such cases, embryos with lower mosaicism degrees and no monosomy demonstrate higher implantation potential. Limitations, reasons for caution The inherent limitations of a retrospective analysis highlight the need for prospective randomized studies to confirm the validity and usefulness of the mosaic embryo transfer score. Wider implications of the findings A scoring system has been developed to prioritize the transfer of mosaic embryos. The crucial variables are associated with embryo quality and the day of biopsy, reflecting the progression of embryonic development. Conversely, variables related to mosaicism play a secondary role. Trial registration number Not applicable

P-233 Embryo transfer timing and clinical pregnancy success: a retrospective study

Abstract Study question The aim of this study is to compare the clinical outcome of day5 versus day6 genetically normal blastocyst transfers. Summary answer 1000 oocytes were analyzed showing a highly significant difference in the number of normal embryos and clinical pregnancy rates in day5 versus day6 blastocysts What is known already studies have shown that day 5 embryos have a higher pregnancy rate compared to day 6 embryos, some studies had only euploid embryos transferred and others did not have any genetic testing done on embryos. No study had a sample size as large as 1760 and the number of embryos studied was a limiting factor in their results. Study design, size, duration The study is a retrospective cross-sectional chart review with multivariate analysis, data from January 2019 up to February 2023 were extracted. 1760 patients were recruited, 1000 in the day5 group and 760 in the day6 group. The data recorded for each patient included maternal age, body mass index (BMI), anti-Müllerian hormone (AMH) level, number, prenatal genetic diagnostic testing, frozen embryo transfer, sperm parameters, and pregnancy status. Participants/materials, setting, methods All embryos were genetically tested for aneuploidy before transfer, using next-generation sequencing technology to rule out 24-chromosome aneuploidies (1-22, X, Y). All patients who have single embryo transfer, frozen embryos and pre-implantation genetic diagnosis (PGD) were included in the study. Continuous variables were expressed as mean with standard deviation and categorical variables as numbers and frequencies (percentages). Statistical differences between groups were estimated using the chi-square or Fisher’s test. All analyses were performed using SAS studio. Main results and the role of chance 1,000 cycles were analyzed for Day 5 and 760 for Day 6 embryo. The number of blastocysts formed, and genetically diagnosed embryos significantly favor the Day 5 group. There’s a significant difference in the number of normal embryos, with the Day 5 group having more. There is a highly significant difference in clinical pregnancy rates. On the other hand, the differences in biochemical pregnancies are not statistically significant. The occurrence of ectopic pregnancies shows no significant difference. Additionally, there is no statistically significant difference in the rates of miscarriage. There is a highly significant difference in blastocyst formation rates, with Day 5 embryos. This indicates that Day 5 embryos have a significantly higher likelihood of forming blastocysts compared to Day 6 embryos. Implantation: Day 5 embryos have an implantation rate of 62.0% compared to 49.7% for Day 6 embryos. Day 5 embryos exhibit a significantly higher likelihood of successful implantation. Euploid: The difference in euploid rates is statistically significant, albeit with a lower p-value of 0.0236. Day 5 embryos have a rate of 59.6%, while Day 6 embryos have a rate of 56.0%. Day 5 embryos are associated with a lower likelihood of being aneuploid Limitations, reasons for caution Limitation of this study is that we do not have data available regarding complications during pregnancy in the day 5 versus day 6 groups and we had no data regarding the the live birth rate between the two groups. Wider implications of the findings This study has the potential to revolutionize worldwide discussions during IVF consultations. With the large sample size we can provide for the first time a statistically significant difference in the percentage of success when transferring a day5 (61%) versus day6 (49%) euploid embryo. Trial registration number not applicable

P-735 Is Artificial Intelligence (AI) currently able to provide evidence-based scientific responses on methods that can improve the outcomes of embryo transfers?

Abstract Study question Could chatbots potentially be used as tools for both patients’ and physicians’ education in an infertility journey? Summary answer Artificial Intelligence (AI) is not yet in a position to give clear, evidence-based recommendations in the field of fertility, particularly concerning embryo transfer. What is known already The rapid development of AI has raised questions about its potential uses in different sectors of everyday life. Individuals from diverse fields have tried to incorporate its usage in their personal but also in their professional lives, with varying levels of success. Specifically in medicine, ChatGPT has been studied intensely with more than 400 related articles having been indexed in PubMed until May 2023, while AI has reportedly also succeeded in several standardized medical tests and passed several Medical Board examinations. Hence the question arises whether chatbots could be used as tools for clinical decision-making or patients’ and physicians’ education. Study design, size, duration We used nine of the most popular free AI chatbots available (ChatGPT, Bard, Writesonic, You, Perplexity, Learnt, Bing, Magickpen, and Rytr) and entered the following command: “Write me a 300-word scientific essay about evidence-based methods that can improve the outcomes of embryo transfer” in May 2023. We collected the responses and extracted the methods each chatbot suggested. When sufficient similarity among answers was present we categorized the answers under one category to facilitate the study. Participants/materials, setting, methods We calculated descriptive statistics and the prevalence of each response. We used as a comparator for widely acceptable practices that are proven to improve embryo transfer outcomes the 2017 ASRM guideline on performing embryo transfer taking also into consideration more current literature. Data was compared using chi-squared tests and a level of p &amp;lt; 0.05 was used as the threshold of statistical significance. Main results and the role of chance The range of the recommendations was from one to nine per chatbot (median = 4, IQR = 2) with an average of 4.78 suggestions per chatbot. Out of a total of 43 recommendations, which could be grouped into 19 similar categories, only 3/19 (15.8%) were evidence-based practices, those being “ultrasound-guided embryo transfer”, which was also the most commonly appearing response, appearing in 7/9 (77.8%) chatbots, “single embryo transfer” appearing in 4/9 (44.4%) chatbots and “use of a soft catheter” in 2/9 (22.2%) chatbots. Some controversial responses appeared even more often than the two latter evidence-based ones with “preimplantation genetic testing (PGT)” being the second most common response with 6/9 chatbots suggesting it (66.7%) and the vague suggestion of “optimal endometrium preparation” being in the third place with 5/9 chatbots (55.6%), both non-evidence-based practices to improve embryo transfer. The majority of the recommendations were unique, with 10 answers appearing only once (1/9; 11.1%), those being “endometrial scratching”, “natural cycle embryo transfer”, “use of GnRH antagonist protocol”, “blastocyst transfer”, “use of specialized catheter”, “preimplantation genetic screening (PGS)”, “mock embryo transfer”, “uninterrupted embryo culture”, “minimizing transfer time”, and “maintaining temperature and pH of the culture media”. Limitations, reasons for caution Firstly, we only used some of the most popular chatbots and not all existing ones. Additionally, chatbots tend to give different answers when repeatedly asked the same questions. However, we believe that our study effectively captures the prevailing practices of chatbot users, who typically pose a question only once. Wider implications of the findings Both patients and physicians should be wary of guiding care based on chatbot recommendations in infertility since the majority of responses consists of scientifically unsupported recommendations. Chatbot results might improve with time especially if trained to obtain information from validated medical databases, however, this will have to be verified scientifically. Trial registration number N/A

P-503 Euploidy rate of day 4 fully developed blastocysts

Abstract Study question Do embryos that fully developed to blastocyst at day 4 have high aneuploidy rates as assessed by preimplantation genetic analysis? Summary answer Fully developed blastocysts at day 4 had a high euploidy rate. What is known already In ART, it is considered important to use good quality blastocysts to increase the pregnancy rate after embryo transfer. Some reports have demonstrated a high pregnancy rate for embryos that fully developed to blastocysts on day 4 after IVF (Ozgur et al., 2021). Furthermore, it has been reported that the speed of embryo development is related to the rate of chromosome aneuploidy (Taylor et al., 2014). There are though, limited reports on the euploidy rate of embryos developed to blastocysts at day 4. Study design, size, duration This retrospective study analyzed 763 embryos (from 184 PGT-A cases) that underwent TE biopsy on day 5 between January to December 2022 to determine their ploidy status. Participants/materials, setting, methods Embryos cultured in a time-lapse incubator were divided into two groups: those that developed to blastocyst (Gardner’s classification: ≥ expansion grade 3) at day 4 (96-100 h) (D4(+) group) and those that did not develop to full blastocyst at day 4 (D4(-) group) and the euploidy rates were compared. Statistical analysis was performed using Fisher’s exact test to determine significant differences (P &amp;lt; 0.05). Main results and the role of chance The euploidy rate of the D4(+) group was 36.5% (80/219), which was significantly higher than the D4(-) group, 23.3% (127/544) (p &amp;lt; 0.01). Moreover, when considering the age of the female, for age ≤39 years, the rates were significantly higher in the D4(+) group (52.1% (61/117) and 35.4% (97/274) p &amp;lt; 0.01 respectively ). For age 40≥ years, it tended to be higher but was not significantly different in the D4(+) group vs D4(-) group (18.6% 19/102 and 11.1% 30/270 respectively). Even when limited to embryos that were graded as good quality blastocysts at day 5, the D4(+) group was significantly higher (p &amp;lt; 0.01) than the D4(-) group (55.0% (60/109) vs 36.1% (90/249)). Limitations, reasons for caution This was a retrospective study limited to day 5 embryos with biopsies taken after vitrification-warming. Wider implications of the findings These results demonstrate that embryos that had already developed to blastocyst at day 4 had a higher rate of euploidy compared to day 5 blastocysts. Thus, when performing PGT-A, biopsy of embryos that have already developed to blastocyst at day 4 are expected to be prioritised for transfer. Trial registration number not applicable

P-318 Reproductive ageing: the seed, the soil, or both?

Abstract Study question Is it all about the embryo? Can a single euploid blastocyst transfer sustain reproductive outcomes in women with advanced reproductive age? Summary answer Despite euploid embryo transfer, endometrial senescence appears to play a crucial role in determining reproductive success, particularly in older patients undergoing in vitro fertilization (IVF). What is known already Advanced reproductive age significantly impairs oocyte and embryo quality. Effective crosstalk and synchronization between embryo and endometrium are pivotal for successful pregnancies, both in vivo and in vitro. Implantation failure remains a persistent challenge in IVF, exacerbated by delayed family planning trends. The transfer of euploid blastocysts through preimplantation genetic testing for aneuploidy (PGT-A) is a potential strategy to address implantation issues, decrease miscarriages, and shorten time-to-pregnancy. However, a significant proportion of IVF treatments remain unsuccessful, suggesting critical factors beyond the embryo. Study design, size, duration This retrospective study analyzed 457 single euploid blastocyst transfers performed by two doctors at a private IVF center from January to November 2023. Inclusion criteria encompassed cases with PGT-A indications (advanced maternal age, repeated implantation failure, recurrent pregnancy loss, etc.) or cases where embryo biopsy was patient-requested. The primary outcome was ongoing pregnancy rate (OPR), with secondary outcomes including pregnancy rate (PR) and clinical pregnancy rate (CPR). Participants/materials, setting, methods Participants were categorized into four groups: Group 1 (&amp;lt; 35 years old; N = 100), Group 2 (35-37 years old; N = 102), Group 3 (38-40 years old; N = 118), and Group 4 (&amp;gt; 41 years old; N = 87). A control group (oocyte donation; N = 50) with single euploid embryo transfers showed a mean recipient age of 43 years. Donors were always less than 32 years old. Endometrial preparation protocols were consistent across all groups. Chi-square test was used to compare groups. Main results and the role of chance Among the 457 transfers analyzed, with a mean female age of 37 years, the overall PR was 71.3%, CPR was 64.8%, and OPR was 60.2%. Groups showed no differences in demographic variables and cycle parameters. Group 1 demonstrated higher PR (83.3%), CPR (78.4%), and OPR (73.5%) compared to Group 2 (PR 70.9%, p = 0.034; CPR 66%, p = 0.047; OPR 62.1%, p = 0.081), Group 3 (PR 68.2%, p = 0.009; CPR 61.2%, p = 0.005; OPR 56.6%, p = 0.008), and Group 4 (PR 65%, p = 0.002; CPR 56.1%, p &amp;lt; 0.001; OPR 51.2%, p = 0.001). Furthermore, the control group (oocyte recipients) showed significantly lower PR (68%, p = 0.031), CPR (54%, p = 0.002), and OPR (46%, p &amp;lt; 0.001) compared to Group 1, but not with the other groups. These findings demonstrate that even with euploid embryos originating from young donors, older women exhibited lower pregnancy rates than young women transferring single euploid blastocysts from their own oocytes and similar rates compared to those with advanced reproductive age transferring their own euploid blastocysts, implying that endometrial ageing may disproportionately influence IVF treatment outcomes. Limitations, reasons for caution The retrospective design introduces inherent bias, notably selection bias and unmeasured confounders. This preliminary data did not account for male infertility factors and their impact on embryo quality. Wider implications of the findings Reproductive aging appears to be influenced by factors beyond the embryo itself, especially in women above 35 years. Further research is warranted to elucidate the causes and impact of reproductive aging on implantation potential, aiming to refine clinical practices and enhance the success of IVF treatments. Trial registration number not applicable