Cohort Of Blastocysts Research Articles

P-583 The ploidy status in blastocysts generated from progestin primed ovarian stimulation and GnRH antagonist protocols: A comparison of 14.000 trophectoderm biopsies

Abstract Study question Is there any difference in the euploidy rate in a large cohort of blastocysts when produced after a progestin-primed or GnRH antagonist protocol? Summary answer Euploidy rates in trophectoderm biopsies when tested with next-generation sequencing (NGS) were similar after progestin-primed or GnRH antagonist protocol. What is known already Preimplantation genetic testing (PGT) might be employed in patients with advanced female age to improve efficiency of embryo transfer. Given the fact that there is no fresh embryo transfer in that policy, progestin primed ovarian stimulation (PPOS) protocol might be preferred over GnRH analogs due to its lower cost and advantage of oral administration instead of s.c injections. However, due to small sample sizes within the limited number of studies, there is lack of consensus whether ploidy status might be affected when an embryo is generated after a PPOS or GnRH analogs protocol. Study design, size, duration A retrospective cohort study was designed among patients that has undergone in-vitro fertilization (IVF) treatment between January-2018 and June-2023 in Bahceci Health Group. For the aim of PGT, only patients treated with PPOS or GnRH antagonists were included. PPOS protocol referred to utilization of either medroxyprogesterone acetate (10 mg/day) or dydrogesterone (20-30 mg/day) to avoid premature ovulation during ovarian stimulation. In GnRH antagonist protocol, flexible or fixed approach was preferred for pituitary suppression. Participants/materials, setting, methods During the study period, 11.616 and 2.480 trophectoderm biopsies had been performed from blastocysts (D5/6) generated after GnRH antagonist or PPOS protocol, respectively. Of them, 9.390 and 1.801 of them were analyzed with NGS. A total of 303 (3.2%) and 17 (0.9%) biopsies were excluded due to failure or non-informative results after testing. For the final analysis, 9.087 and 1.784 biopsies were included in the antagonist and PPOS arms for the current study. Main results and the role of chance The mean age of women treated with GnRH antagonist and PPOS was comparable (36.5±5.3 vs. 36.7±5.4 years, p = 0.17). The respective numbers of euploid embryos were 3.912 and 1.784. Per biopsied embryos, the rate of euploidy was 33.7% and 34.8% (p = 0.30). The rate of euploidy per metaphase II oocyte and 2-PN were 10.6% and 13.5% in the GnRH antagonist arm and they were 11.3% and 14.4% in the PPOS arm (p = 0.08 and p = 0.07) without any statistical significance. When only patients included in whom all available blastocysts were biopsied, the linear regression model revealed only female age (RR: -0.32 CI%95: -0.38 to – 0.26, p < 0.001) as the independent factor for the prediction of ploidy rate (euploid/euploid + aneuploid embryos) when body mass index, duration of infertility, AMH, number of oocytes, number of metaphase-II oocytes, and the type of ovarian stimulation protocol were entered into the model. This model represented female age as the sole independent factor (RR: -0.14 CI%95: -0.18 to – 0.11, p < 0.001) when euploidy rate per 2PN was tested with the same variables. Limitations, reasons for caution The inherent risks cannot be excluded due to the retrospective nature of the current study. Wider implications of the findings The current study includes the largest number of trophectoderm biopsies generated from PPOS and GnRH antagonist protocol, giving the opportunity to compare them with each other. Due to lack of any superiority of one protocol regarding ploidy status, financial cost and easiness might favor PPOS when PGT is planned. Trial registration number None

The Blastulation Rate Is Negatively Associated With Euploid Rate

ObjectivesTo study the association between the blastulation rate, the presence of 1 pronucleus (1PN) zygotes, and the ploidy of the cohort of blastocysts. MethodsA 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. ResultsA 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. ConclusionsThis 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.

Devenir clinique après transfert de blastocystes vitrifiés selon l’indication de congélation

ObjectivesIn France, embryo thawing concern 45.8% of attempts at assisted reproductive technologies excluding artificial inseminations. This proportion is constantly increasing for various reasons. The main objective of this study is to compare the live birth rate following frozen blastocyst transfer (FBT) according to the initial indication for freezing. MethodsThis is a retrospective study including patients who underwent FBT between 01/01/2020 and 06/30/2022 at the Regional University Hospital Center of Tours. The results were compared (univariate and multivariate analyses) between the three main indications for freezing: freezing of the complete cohort of blastocysts for risk of ovarian hyperstimulation (=OHS), freezing of supernumerary blastocysts after fresh blastocyst transfer (BT) with pregnancy (=second request) or without pregnancy (=BT failure). Results have also been described for other indications. ResultsAmong the 963 FBT cycles selected, 28% of live births by thawing were obtained, all indications of freezing combined. A significantly lower rate was identified in the FBT failure group compared to the OHS group. However, after adjustment, the results remained significant for the age of the patient on the freezing cycle but not for the indication for freezing. ConclusionsThe outcome of a FBT does not seem significantly impacted by the indication of freezing considering the confounding factors. The prospective analysis of more data from a multicenter study would be necessary to confirm these results.

P-743 Semen samples quality and blastocysts diversity after PGT-A

Abstract Study question Does semen samples quality contribute to blastocyst diversity by correlating with the presence of DNA in the blastocoelic fluid (BF)? Summary answer Semen samples quality is associated with the detection of DNA in BFs from both euploid and aneuploid blastocysts What is known already DNA was detected by whole genome amplification (WGA) in BFs from expanded blastocysts dependently on the blastocyst chromosome condition (assessed by PGT-A on trophectoderm (TE) cells), being more frequent in aneuploid blastocysts. TE-euploid blastocysts with positive BF-WGA also seem to have less chances to implant compared to those with failed BF-WGA. The extrusion of abnormal cells from the embryo proper was proposed as a mechanism triggered by the mosaic embryo to eliminate abnormal cells. Severe male factor samples are at risk of causing embryo mosaicism. Possible association between semen quality and DNA in BFs has not yet been investigated. Study design, size, duration This retrospective cohort study included 115 patients (maternal age 37.9±4.1), which underwent PGT-A in the last three years. The aim of the study was to evaluate whether the detection of DNA in BFs was correlated with semen samples indices. Both TE-euploid and TE-aneuploid blastocysts were considered for the analysis and stratified according to the semen samples quality as normozoospermic (N), moderate oligoasthenoteratozoospermic (m-OAT) and severe OAT (s-OAT) following the 2021 WHO criteria. Participants/materials, setting, methods BF and trophectoderm (TE) biopsies were collected from high grade expanded blastocysts before vitrification, and submitted to WGA. Amplification after WGA was evaluated by loading an aliquot of the amplified product onto a 1.5% agarose gel. 24-chromosome analysis was performed on TE biopsies. To assess the impact of semen samples quality on the detection of DNA in BFs, a multiple logistic regression analysis was performed correcting for maternal age as a confounding factor. Main results and the role of chance The BF from 590 blastocysts (265 TE-euploid and 325 TE-aneuploid) were submitted to WGA. In TE-euploid blastocysts, 125 derived from N semen samples, 52 from m-OAT, and 88 from s-OAT. The incidence of positive BF-WGA was proportional to the severity of the male factor condition and occurred in 48.0% of blastocysts originated from N samples, in 55.8% of blastocysts derived from m-OAT, and in 60.2% of blastocysts from s-OAT. In TE-aneuploid blastocysts, 178 originated from N semen samples, 80 from m-OAT, and 67 from s-OAT. The incidence of positive BF-WGA showed an opposite trend compared with the TE-euploid group being detected in 64.0% of blastocysts originated from N samples, in 66.3% of blastocysts derived from m-OAT, and in 52.2% of blastocysts obtained from s-OAT. The relationship between semen quality and positive BF-DNA amplification was confirmed by a multiple logistic regression model setting the BF-WGA as the outcome variable. In the TE-euploid blastocyst cohort, s-OAT showed an odd ratio (OR) of 1.78 (95% CI: 1.02-3.14; p=0.045), whereas in the TE-aneuploid blastocyst group, s-OAT displayed an OR of 0.55 (95% CI: 0.31-0.98; p=0.043). Limitations, reasons for caution This is a retrospective cohort study including a limited number of cases where the different categories of sperm samples were not equally represented. Although the results of BF-WGA were evaluated per single blastocyst, they cannot be considered independent variables. Wider implications of the findings In s-OAT samples, abnormal cell elimination is especially active in TE-euploid blastocysts, whereas in TE-aneuploid blastocysts the efficiency of this process is reduced suggesting the difficulty of eliminating abnormal cells in an aneuploid background. BF-DNA amplification is a minimally invasive tool contributing additional knowledge to elucidate blastocyst diversity. Trial registration number None

Identifying parental and cell-division origins of aneuploidy in the human blastocyst

Preimplantation genetic testing commonly employs simplistic copy-number analyses to screen for aneuploidy in blastocyst trophectoderm biopsies. Interpreting intermediate copy number alone as evidence of mosaicism has led to suboptimal estimation of its prevalence. Because mosaicism originates from mitotic nondisjunction, utilizing SNP microarray technology to identify the cell-division origins of aneuploidy might provide a more accurate estimation of its prevalence. The present study develops and validates a method of determining the cell-division origin of aneuploidy in the human blastocyst by using both genotyping and copy-number data in parallel. The concordance of predicted origins with expected results was demonstrated in a series of truth models (99%-100%). This included determination of X chromosome origins from a subset of normal male embryos, determination of the origins of translocation chromosome-related imbalances via embryos from couples with structural rearrangements, and prediction of either mitotic or meiotic origins via multiple rebiopsies of embryos with aneuploidy. In a cohort of blastocysts with parental DNA (n= 2,277), 71% were euploid, 27% were meiotic aneuploid, and 2% were mitotic aneuploid, indicating a low frequency of bona fide mosaicism in the human blastocyst (mean maternal age: 34.4). Chromosome-specific trisomies in the blastocyst were also consistent with observations previously established in products of conception. The ability to accurately identify mitotic-origin aneuploidy in the blastocyst could benefit and better inform individuals whose IVF cycle results in all aneuploid embryos. Clinical trials with this methodology might also help provide a definitive answer regarding the reproductive potential of bona fide mosaic embryos.

HIGH FOLLICLE STIMULATING HORMONE (FSH) LEVEL ON THE DAY OF TRIGGER IMPACTS NEGATIVELY THE EUPLOIDY RATE OF BLASTOCYSTS

To evaluate, in ovarian stimulation cycles for IVF/ICSI, the impact of the systemic FSH level on the Day of Trigger (DoT) on the ploidy rate of the blastocyst cohort.

CONCORDANCE RATES BETWEEN EMBRYONIC CELL-FREE DNA AND TROPHECTODERM BIOPSY FROM FRESH AND FROZEN-THAWED BLASTOCYSTS

CONCORDANCE RATES BETWEEN EMBRYONIC CELL-FREE DNA AND TROPHECTODERM BIOPSY FROM FRESH AND FROZEN-THAWED BLASTOCYSTS

P–167 Assuring quality in embryology decision making: blastocyst grading agreement assessed via a smartphone application

Abstract Study question Given the subjectivity of blastocyst grading and the challenge of performing routine competency assessments, how consistently do embryologists grade blastocysts when using an easily accessible phone application? Summary answer Grading agreement was fair to moderate for inner cell mass (ICM) and trophectoderm (TE), evidence that a mobile application can be used for quality assurance. What is known already Embryologists routinely perform external quality assessments (EQA), though the utility of EQA for quality improvement is limited, and more active, user-friendly tools are needed to improve quality assurance in embryology. Blastocyst grading is one of the most important and subjective tasks in clinical embryology, important for both blastocyst ranking and decision to freeze. Inter-user agreement is only fair for ICM (kappa 0.349) and TE grade (kappa 0.397; Storr et al., 2017). Fair agreement has also been reported for decision to freeze for a cohort of blastocysts that exhibited borderline morphology (kappa 0.301; Hammond et al., 2020). Study design, size, duration A prospective study of blastocyst grading consistency using ARTCompass, a mobile phone application designed to assess clinical decision making of laboratory staff for andrology and embryology competency. Two assessments, each with 100 images of expanded blastocysts in three planes, were performed by 42 embryologists from 9 clinics in 2 countries between April to July 2020. Survey 1 assessed ICM grading and survey 2 assessed TE grading using the same set of images for consistency. Participants/materials, setting, methods Blastocysts were of proportionally mixed grades (ranging from grade A to X) using a modified Gardner system that included “X” for non-viable ICM/TE. Embryologists were advised to complete the tests individually in one sitting. The Fleiss kappa coefficient (k) measured inter-rater agreement among embryologists when assigning blastocyst grade. Kappa value interpretation is as follows <0.20: poor; 0.21–0.40: fair; 0.41–0.60: moderate; 0.61–0.80: good and 0.81–1.00: very good. Main results and the role of chance Overall, agreement for ICM and TE grades was moderate among embryologists (kappa 0.47, 0.52, respectively). ICM grade B and C had the lowest agreement (0.37, 0.39), while ICM grade X (no, or degenerate ICM), and TE grade A showed the highest agreement (0.68, 0.62). These results illustrate that embryologists had difficultly classifying ICM grade when it was of moderate to poor quality (grade b or c), likely due to subjectivity in grading size and compaction level, but were good at classifying ICM grade when there was no apparent ICM (grade x). For TE grade, embryologists consistently identified a top-quality TE (grade a), which is reassuring as TE grade is the primary morphological feature used for blastocyst ranking. In general, this QA platform offers ease of use and shows agreement values for ICM and TE that are similar to other studies, suggesting that blastocyst grading with a mobile phone application is a viable option for quality assurance. Limitations, reasons for caution Only blastocyst grading was assessed, therefore additional competency assessments using a mobile device should be assessed for accuracy. Further studies are needed to determine if mobile applications can improve competency. Wider implications of the findings: Ease of use by 42 embryologists indicates mobile applications may provide a user-friendly and accessible platform for QA. Since effective and efficient assessment of competency and KPIs is an ongoing challenge for laboratories, a mobile application is a novel and effective tool to monitor QA parameters in the IVF laboratory Trial registration number Not applicable

P–551 Blastocyst cohort size is not associated with embryo aneuploidy: comprehensive multi-centre data from current preimplantation genetic testing cycles

Abstract Study question Does blastocyst cohort size impact aneuploidy rates, evaluated by next generation sequencing (NGS)? Summary answer Embryo aneuploidy rates were independent of blastocyst cohort size across all patient ages. What is known already The effects of ovarian response on oocyte and embryo quality remain controversial. Several studies have proposed that a high response to ovarian stimulation may negatively impact oocyte competence. Alternatively, irrespective of maternal age, a poor ovarian response may potentially compromise embryo quality. Using blastocyst cohort size as an indirect measure of ovarian response, previous studies applying array comparative genomic hybridisation (aCGH) have demonstrated that the number of embryos available for biopsy does not impact embryo aneuploidy rates. Nevertheless, these findings remain to be confirmed in a comprehensive cohort, using current approaches for preimplantation genetic testing for aneuploidies (PGT-A). Study design, size, duration Retrospective, international, cohort study of 3998 patients from 16 clinics undergoing PGT-A from 2016–2020. We evaluated 11665 blastocysts, tested using trophectoderm (TE) biopsy and next generation sequencing (NGS). To eliminate bias of multiple treatments, we considered only the first PGT-A cycle for all patients. Both autologous and donation cycles were included in the analysis. Cycles were excluded if they utilised preimplantation genetic testing for monogenic disorders (PGT-M) or preimplantation genetic testing for structural rearrangements (PGT-SR). Participants/materials, setting, methods We evaluated aneuploidy and mosaicism rates, as well as the proportion of patients who had at least one euploid embryo suitable for transfer. Findings were stratified according to SART-defined maternal age groups, <35 (n = 698/2622 patients/blastocysts), 35–37 (n = 988/3141 patients/blastoycsts), 38–40 (n = 1447/3939 patients/blastocysts), 41–42 (653/1562 patients/blastocysts) and >42 (212/401 patients/blastocysts) and blastoycst cohort size (1–2, 3–5, 6–9 and 10 or more biopsied blastocysts). Main results and the role of chance The mean maternal age was 37.0±3.7. The overall embryo aneuploidy rate was 50.6% (5904/11665), while mosaicism was established in 4.0% (469/11665) of blastocysts. As expected, the proportion of aneuploid embryos increased steadily with advancing maternal age (31.8%, 41.5%, 58.4%, 71.2%, 87.8%; p < 0.0001), while mosaicism rates did not vary significantly (p = 0.2). Within each age group, we observed no association between the number of blastocysts biopsied and aneuploidy or mosaicism rates. However, as previously suggested, the chance of having at least one euploid embryo increased linearly with the number of embryos biopsied. We observed that young patients (<35) with 1–2 blastocysts had a 70.4% of having at least one embryo suitable for transfer, which increased to 96.4% and 99.2% with 3–5 and 6–9 blastocysts, respectively. Similar trends were observed in the 36–38 and 39–40 age groups. Patients in the 40–41 age group had a significantly lower chance of having a suitable embryo for transfer. Nevertheless, the chance increased from 27.2% with 1–2 embryos to 61.2% with 3–5 blastocysts. Patients with >10 embryos had at least one euploid embryo in 100% of cases, across all ages. Albeit, the numbers of patients within this category was low, and decreased significantly with advancing maternal age. Limitations, reasons for caution While blastocyst cohort size is considered to be an indirect measure of ovarian reserve, the number of oocytes retrieved was not evaluated. Our study only included the first PGT-A cycle for all patients. Subsequent, alterations in stimulation protocols may have resulted in an improved response in some patients. Wider implications of the findings: The comprehensive nature of the study, based on current PGT-A approaches and a large number of cycles across 16 centres increases clinical confidence in the notion that ovarian response is independent of embryo aneuploidy. Importantly, our findings may serve as a valuable clinical resource to guide patient counselling strategies. Trial registration number NA

Tobacco consumption is associated with slow-growing day-6 blastocysts

ObjectiveTo investigate if there are any obvious clinical factors associated with delayed blastulation at day 6 (D6) compared with day 5 (D5).DesignMonocentric observational cohort study from November 2012 to December 2018.SettingTertiary-care academic medical center.Patient(s)A total of 941 women with an entire cohort of exclusively D5 blastocysts compared with 162 patients with a cohort of exclusively D6 blastocysts.Intervention(s)None.Main Outcome Measure(s)Clinical characteristics and data related to the ovarian stimulation protocols.Result(s)After univariate analysis, a significantly higher proportion of women who were active smokers was found in the D6 group compared with the D5 group (n = 22/162 [13.6%] vs. n = 82/941 [8.7%]). In addition, the women in the D6 group had a higher rank number of assisted reproductive technology (ART; total no. of ART cycles performed: 2.1 ± 1.4 vs. 1.6 ± 1.1) and a lower antral follicle count (AFC; 18.7 ± 11.3 vs. 22.2 ± 12.8). Moreover, fertilization with the use of intracytoplasmic sperm injection was used more frequently in the D6 group compared with the D5 group. Logistic regression analysis adjusted for confounders highlighted several independent predictors for reaching blastocyst stage at D6 rather than D5: being an active smoker, previous ART cycles, and a lower AFC.Conclusion(s)Obtaining an exclusively D6 blastocyst cohort is independently associated with women who are active smokers, previous ART cycles, and a lower AFC. These findings provide evidence, to be confirmed by further studies, that women who are active smokers could greatly benefit from smoking cessation before undergoing ART.

Should we freeze it? Agreement on fate of borderline blastocysts is poor and does not improve with a modified blastocyst grading system.

What is the inter-observer agreement among embryologists for decision to freeze blastocysts of borderline morphology and can it be improved with a modified grading system? The inter-observer agreement among embryologists deciding whether to freeze blastocysts of marginal morphology was low and was not improved by a modified grading system. While previous research on inter-observer variability on the decision of which embryo to transfer from a cohort of blastocysts is good, the impact of grading variability regarding decision to freeze borderline blastocysts has not been investigated. Agreement for inner cell mass (ICM) and trophectoderm (TE) grade is only fair, factors which contribute to the grade that influences decision to freeze. This was a prospective study involving 18 embryologists working at four different IVF clinics within a single organisation between January 2019 and July 2019. All embryologists currently practicing blastocyst grading at a multi-site organisation were invited to participate. The survey was comprised of blastocyst images in three planes and asked (i) the likelihood of freezing and (ii) whether the blastocyst would be frozen based on visual assessment. Blastocysts varied by quality and were categorised as either top (n= 20), borderline (n= 60) or non-viable/degenerate quality (n= 20). A total of 1800 freeze decisions were assessed. To assess the impact of grading criteria on inter-observer agreement for decision to freeze, the survey was taken once when the embryologists used the Gardner criteria and again 6months after transitioning to a modified Gardner criterion with four grades for ICM and TE. The fourth grade was introduced with the aim to promote higher levels of agreement for the clinical usability decision when the blastocyst was of marginal quality. The inter-observer agreement for decision to freeze was near perfect (kappa 1.0) for top and non-viable/degenerate quality blastocysts, and this was not affected by the blastocysts grading criteria used (top quality; P= 0.330 and non-viable/degenerate quality; P= 0.18). In contrast, the cohort of borderline blastocysts received a mixed freeze rate (average 52.7%) during the first survey, indicative of blastocysts that showed uncertain viability and promoting significant disagreement for decision to freeze among the embryologists (kappa 0.304). After transitioning to a modified Gardner criteria with an additional grading tier, the average freeze rate increased (64.8%; P< 0.0001); however, the inter-observer agreement for decision to freeze was unchanged (kappa 0.301). Therefore, significant disagreement for decision to freeze among embryologists is an ongoing issue not resolved by the two grading criteria assessed here. Blastocyst assessment was performed from time-lapse images in three planes, rather than with a microscope in the laboratory. The inter-observer agreement for decision to freeze may be lower for embryologists working in different clinics with different grading protocols. The decision to freeze a blastocyst with borderline morphology is a common clinical issue that has the potential to arise for any patient during blastocyst culture. Disagreement for decision to freeze these blastocysts, and therefore clinical usability in frozen embryo transfer cycles, affects consistency in patient care due to a potential impact on cumulative live birth rates, as well as financial, emotional and time costs associated with the frozen embryo transfer cycles. We demonstrate significant disagreement for decision to freeze borderline blastocysts among embryologists using the same grading scheme within a large multisite organisation, a phenomenon which was not improved with a modified grading system. Decision-making around borderline embryos is an area requiring further research, especially as studies continue to demonstrate the reduced but modest live birth rates for low quality blastocysts (Grade C). These results provide support for emerging technology for embryo assessment, such as artificial intelligence. None declared. Not applicable.

Euploidy in relation to blastocyst sex and morphology.

The objective of our study is to assess the relationship of embryo ploidy status in relation to embryo sex, morphological characteristics, and transfer parameters. This is a retrospective cohort study at an academic medical center of patients who underwent in vitro fertilization with preimplantation genetic screening (PGS) from 2010 to 2015. Embryos were screened with 24-chromosome preimplantation genetic screening with day 5/6 trophectoderm biopsy. We investigated embryo euploidy in relation to morphology (expansion, inner cell mass, trophectoderm), embryo sex, biopsy day, and blastocyst cohort size. We used multivariate logistic regression to calculate odds ratios of euploidy in relation to these parameters. A total of 1559 embryos from 316 cycles and 233 patients (mean maternal age = 37.8 ± 4.2years) were included in the analysis. Six hundred and twenty-eight blastocysts (40.3%) were found to be euploid. Expansion (p < 0.001), inner cell mass (ICM) (p < 0.01), and trophectoderm grade (p < 0.001) were significantly associated with embryo ploidy in bivariate models controlling for maternal age, while embryo sex, biopsy day, and blastocyst cohort size were not associated with embryo ploidy. In a multivariate model, we found that maternal age (p < 0.001), higher grade of expansion (p < 0.01), and better quality trophectoderm (p < 0.001 for A compared to C grade) remained significantly associated with increased embryo euploidy, but ICM grade was no longer significant. Embryo sex was not associated with ploidy status, though male embryos were found to be associated with higher trophectoderm scores (p < 0.02). This is the largest study to date to investigate PGS-tested embryo sex and ploidy status. While maternal age and some morphological parameters (expansion, trophectoderm grade) are associated with euploidy in our cohort, other parameters such as embryo sex, biopsy day, and cohort size are not. Though embryo sex was not associated with euploidy, male embryos were found to be associated with higher trophectoderm grades. Additional investigation in larger studies is warranted.

Estimation of age-dependent decrease in blastocyst euploidy by next generation sequencing: development of a novel prediction model.

We developed a model to estimate the female age-dependent decrease in blastocyst euploidy and the impact of blastocyst cohort size on the likelihood of having at least one euploid blastocyst for transfer. Retrospective analysis of 1296 trophectoderm biopsies by next-generation sequencing analysis from 436 infertile couples undergoing intracytoplasmic sperm injection and preimplantation genetic testing for aneuploidy. A logistic regression model was fit to the data. The dependent and independent variables were embryo genetic status and female age, respectively. The method of fitting was quadratic on age, and the model was validated with cross validation by a data splitting technique. The decrease in the probability of blastocyst euploidy follows an age-dependent binomial distribution, progressing with every year of female age, from 1.2% to 24.5% in 28-44 years-old women (P<0.0001). The minimum number of blastocysts needed to obtain at least one euploid blastocyst for transfer was computed for different probabilities and female ages. At the age of 28 years, a total of three blastocysts is required to obtain at least one euploid blastocyst with 90% probability, whereas it is 4, 5, 6, 9, 16 and 29 for ages 35, 37, 39, 41, 43, and 45, respectively. A novel prediction model estimates the probability of blastocyst euploidy and the number of blastocysts required to obtain at least one euploid embryo for transfer. This new resource based on f emale age and blastocyst cohort size will aid clinicians counsel and plan treatment of infertile couples undergoing IVF/ICSI.

Estimation of mosaicism in a blastocyst cohort

Estimation of mosaicism in a blastocyst cohort

Does euploid embryo ranking by trophectoderm cell mitochondrial dna content correspond with ranking by blastocyst morphology within an individual patient’s cohort of blastocysts?

Does euploid embryo ranking by trophectoderm cell mitochondrial dna content correspond with ranking by blastocyst morphology within an individual patient’s cohort of blastocysts?

Do donor oocyte cycles comply with ASRM/SART embryo transfer guidelines? An analysis of 13,393 donor cycles from the SART registry

To analyze donor oocyte cycles in the Society for Assisted Reproductive Technology (SART) registry to determine: 1) how many cycles complied with the 2009 American Society for Reproductive Medicine/SART embryo transfer guidelines; and 2) cycle outcomes according to the number of embryos transferred. For donor oocyte IVF with donor age <35years, the consideration of single-embryo transfer was strongly recommended. Retrospective cohort study of United States national registry information. Not applicable. A total of 13,393 donor-recipient cycles from 2011 to2012. Embryos transferred in donor IVF cycles. Percentage of compliant cycles, multiple pregnancy rate. There were 3,157 donor cleavage-stage transfers and 10,236 donor blastocyst transfers. In the cleavage-stage cycles, 88% met compliance criteria. The multiple pregnancy rate (MPR) was significantly higher in the noncompliant cycles. In a subanalysis of compliant cleavage-stage cycles, 91% transferred two embryos and only 9% single embryos. In those patients transferring two embryos, the MPR was significantly higher (33% vs. 1%). In blastocyst transfers, only 28% of the cycles met compliance criteria. The MPR was significantly higher in the noncompliant blastocyst cohort at 53% (compared with 2% in compliant cycles). The majority of donor cleavage-stage transfers are compliant with current guidelines, but the transfer of two embryos results in a significantly higher MPR compared with single-embryo transfer. The majority of donor blastocyst cycles are noncompliant, which appears to be driving an unacceptably high MPR in these cycles.

310 COBALAMIN SUPPLEMENTATION DURING IN VITRO MATURATION IMPROVES PREIMPLANTATION DEVELOPMENT OF SHEEP EMBRYOS

Oocyte maturation process includes the establishment of proper epigenetics marks, fundamental to ensure successful pregnancy. Epigenetic maturation of the oocyte depends on one-carbon-metabolism (OCM), whose key elements (cobalamin, folate) are crucial cofactors for the transfer of methyl groups onto chromatin. Commercially available IVM-media only partially supports oocyte metabolic requirements, and thus may negatively affect epigenetic maturation. Of relevance, cobalamin, one of the OCM cofactors normally present in follicular fluid, is missing in IVM-media. We investigated if cobalamin supplementation of IVM media affects sheep oocyte developmental competence in term of subsequent embryo development, epigenetic pattern, and fetal survival. Briefly, sheep oocytes were isolated from ovaries and divided into 2 groups: an untreated control (in vitro CTR group) and oocytes in vitro matured in medium containing cobalamin at 200 p.m. (Cobalamin group). Following maturation, MII oocytes were in vitro fertilized and cultured until blastocyst stage. A cohort of blastocysts was surgically transferred to recipient ewes and then conceptuses were collected at 20 days of pregnancy. Naturally mated conceptuses were also collected (in vivo CTR group). In vitro-matured oocytes and -derived blastocysts were analysed (i) by immunofluorescence anti-5-methylcitydine and (ii) by qRT-PCR for the expression of a panel of genes (MATb, ACHY, CBS, MTHFR, DNMT1) involved in OCM pathway. Immunofluorescence results were analysed by Image J software. Decimal variables were analysed using the Mann-Whitney test, while variables were expressed as percentage with the Fisher exact test. Cobalamin exposure during IVM significantly increased (i) cleavage rate (cobalamin 130/220 v. in vitro CTR 134/191, P &lt; 0.02) following in vitro fertilization and (ii) global DNA methylation in blastocyst-stage embryos (P &lt; 0.05). Then, qRT-PCR analysis of a select panel of OCM genes following IVM supplemented with cobalamin revealed a downregulation of MATb, ACHY, and DNMT1 in cobalamin-treated oocytes v. in vitro CTR group (P &lt; 0.05), while no differences were observed at blastocyst stage. Finally, we found that the survival rate at 20 days of pregnancy was comparable in in vitro-produced (IVP) embryos from cobalamin and in vitro CTR oocytes, but reduced compared to in vivo CTR (cobalamin 60%, in vitro CTR 72%, and in vivo CTR 100%). Altogether, our results showed that cobalamin supplementation in IVM medium positively affects competence of oocytes and methylation profile at the blastocyst stage, presumably through the OCM pathway. Moreover, postimplantation survival of IVP conceptus, derived from untreated and treated oocytes, was reduced compared to naturally mated ones. Further investigation will clarify whether cobalamin supplementation influences fetal and placental development of IVP conceptus.

Comparable clinical outcomes and live births after single vitrified–warmed and fresh blastocyst transfer

Selective single-blastocyst transfer (SBT) in fresh cycles has been effective in reducing multiple pregnancies. However, we do not know whether this successful strategy of fresh transfer cycles is suitable for cryopreserved cycles. The present study was undertaken to evaluate the feasibility and value of SBT in vitrified–warmed cycles. Clinical pregnancy rate (CPR) was similar with vitrified and fresh SBT (46.61% versus 52.15% respectively). Of the pregnant patients, monozygotic twin, miscarriage and ectopic pregnancy rates were similar with vitrified and fresh SBT. For the newborns, no significant difference was observed in live birth, low birthweight, premature delivery and birth defects rates between vitrified and fresh SBT. With respect to the quality of transferred blastocysts (from BB to AA), a similar CPR and miscarriage rate was obtained for both vitrified and fresh SBT when a similar blastocyst cohort graded ⩾3BB was transferred. The data show that vitrified SBT is an effective means of reducing multiple pregnancy and that comparable clinical outcomes and live births are achieved if single blastocysts graded ⩾3BB are transferred for both vitrified and fresh SBT. These data should encourage clinics to evaluate their embryo transfer policy and adopt vitrified SBT as everyday practice.Selective single-blastocyst transfer in fresh cycles has been an effective method to reduce the multiple pregnancies. However, due to a lack of adequate studies, we do not know whether this successful strategy in fresh transfer cycles is suitable in cryopreserved cycles. The present study was undertaken to explore the feasibility and value of single-blastocyst transfer in vitrified–warmed cycles. We found that single-blastocyst transfer in vitrified–warmed cycles is an effective means of reducing multiple pregnancy, and comparable clinical outcomes and live births were achieved if single blastocysts graded ⩾3BB were transferred for both vitrified–warmed and fresh blastocyst transfer. These data should encourage clinics to evaluate their embryo transfer policy and adopt single-blastocyst transfer in cryopreserved cycles as their everyday practice.

Predictive Value of Sperm Parameters on Embryo Quality, a 26970 Embryo Cohort Study of an Ovum Donation Program

Predictive Value of Sperm Parameters on Embryo Quality, a 26970 Embryo Cohort Study of an Ovum Donation Program