Top-quality Embryos Research Articles

Identify high-risk DOR women ≤ 35 years old following assisted reproduction technology through cutoffs of anti-mullerian hormone and antral follicle counts

BackgroundFemales with diminished ovarian reserve (DOR) have significantly lower cumulative live birth rates (CLBRs) than females with normal ovarian reserve. A subset of young infertile patients, whose ovarian reserve is declining but has not yet met the POSEIDON criteria for DOR, has not received the attention it merited. These individuals have not been identified in a timely manner prior to the initiation of assisted reproductive technology (ART), leading to suboptimal clinical pregnancy outcomes. We categorized this overlooked cohort as the “high-risk DOR” group.ObjectiveThe primary aim of this study was to identify high-risk DOR patients through anti-Mullerian hormone (AMH) and antral follicle counts (AFCs).MethodsA total of 10037 young women (≤ 35 years old) who underwent their first initial oocyte aspiration cycle at a single reproductive medicine center were included and further classified into three groups, based on the thresholds for AMH and AFC established through receiver operating characteristic (ROC) analysis and in alignment with the POSEIDON criteria. Two ROC analyses were performed to identify the cutoff values of AMH and AFC to obtain one viable embryo (one top-quality embryo or one viable blastocyst). The cutoffs of ROC were measured by sensitivity and specificity. The primary outcome was the cumulative live birth rate (CLBR) per oocyte aspiration cycle. The secondary outcomes included the number of oocytes retrieved and the number of viable embryos formed. Pearson’s chi-square tests were conducted to compare the clinical outcomes among the three groups. Furthermore, univariate logistic regression analyses were performed to investigate the associations between ovarian reserve and clinical outcomes. All of the above comparisons between the high-risk DOR and NOR were further confirmed by propensity score matching (PSM) (1:1 nearest-neighbor matching, with a caliper width of 0.02).ResultsAccording to the ROC analyses and POSEIDON criteria, the present study identified a population of high-risk DOR patients (1.20 ng/mL < AMH values < 2.50 ng/mL, with 6 ≤ AFC ≤ 10; n = 682), and their outcomes were further compared to those of DOR patients (positive control, AMH values ≤ 1.2 ng/mL, and/or AFC ≤ 5; n = 1153) and of NOR patients (negative control, 2.5 ng/mL ≤ AMH values ≤ 5.5 ng/mL, and 11 ≤ AFC ≤ 20; n = 2649). Patients in the high-risk DOR group had significantly lower CLBRs than those in the NOR group (p < 0.001) but higher CLBRs than those in the DOR group (p < 0.001). Logistic regression further demonstrated that high-risk DOR was associated with a lower likelihood of cumulative live birth chance (OR 0.401, 95% CI: 0.332–0.486, p < 0.001) than NOR was, with a greater likelihood of cumulative live birth chance (OR 1.911, 95% CI:1.558–2.344, p < 0.001) than DOR was. To investigate the effects of embryo development stage, the outcomes of D3 embryos and blastocysts were analyzed separately. Significant differences in pregnancy outcomes were detected only in D3 embryo ET cycles among the three groups (high-risk DOR vs. NOR, all p < 0.05; DOR vs. NOR, all p < 0.05). DOR/high-risk DOR did not influence the pregnancy loss rates or pregnancy outcomes (clinical pregnancy rates and ongoing pregnancy rates) per positive HCG cycle (all p > 0.05). After PSM, the differences in ovarian response and pregnancy outcomes between the high-risk DOR and NOR groups were consistent with the results before PSM.Conclusion(s)Our study revealed that the CLBR of the high-risk DOR patients was significantly lower than that of females with normal ovarian reserve and greater than that of females with DOR. The values of AMH ranging from 1.2 to 2.5 and AFC ranging from 6 to 10 appeared to constitute meaningful thresholds in females with mildly reduced ovarian reserve.

Dual trigger or hCG alone: A retrospective analysis on patients with diminished ovarian reserve under in vitro fertilization and embryo transfer (IVF-ET) treatment

ObjectiveWith remarkable deficiency in both oocyte stock and competence, the prognosis of IVF-ET in diminished ovarian reserve (DOR) is obstinately poor, underscoring warranted optimization to current procedures. We compared the efficacy of dual-trigger (hCG plus GnRH-a) and hCG alone on the outcomes for DOR patients. Study designA total of 381 couples and 857 controlled ovarian stimulation (COS) cycles, and 222 couples and 366 frozen embryo transfer (FET) ones were included. The intermediate outcomes during oocyte retrieval and in vitro culture were compared based on COS dataset, while outcomes after embryo transfer analyzed based on FET dataset. The marginal effect of all study factors and covariates were evaluated with a cluster-weighted GEE model. Results and conclusionNeither the intermediate nor implantation outcomes were improved by dual-trigger. The OR values were 1.08 (95 % CI: 0.41–2.78) for retrieval cancellation, 1.33 (95 % CI: 0.89–2.00) for oocyte harvest, 1.04(95 %CI: 0.94–1.15) for viable embryo and 1.03(95 %CI: 0.88–1.19) for top-quality embryo. Similarly, the ORs were 0.90 (95 %CI: 0.62–1.30) for implantation and 0.97 (95 %CI: 0.56–1.69) for clinical pregnancy. This equivalence remained unchanged after adjusting for the covariates such as age, BMI, controlled ovarian stimulation protocols, etc. Thus, dual-trigger cannot provide significant advantage over hCG in related to immediate or clinical outcomes of IVF-ET treatments in DOR patients.

Assisted oocyte activation significantly improves zygote formation, cleavage, and implantation rates in patients with a history of fertilization failures.

Fertilization check performed at the 18th hour following classic in vitro fertilization procedure (IVF) or intracytoplasmic sperm injection (ICSI) is a critical stage in assisted reproduction. The success of the treatment is significantly reliant on the quantity of zygotes exhibiting two pronuclei. Consequently, low fertilization rates or complete fertilization failure are highly undesirable outcomes for both patients and reproductive specialists. Applying additional calcium ionophore for oocyte activation subsequent to ICSI may offer benefits and potentially enhance treatment outcomes, particularly for patients who have experienced low or absent fertilization rates (FR) in previous treatment cycles. The aim of the study is to evaluate the efficacy of Ca2+ ionophore application for oocyte activation. A retrospective analysis of 924 oocytes obtained from 120 patients who underwent ICSI cycles with a history of low or no fertilization as a result of previous unsuccessful treatment rounds. The next ART cycle followed with additional oocyte Ca2+ ionophore activation applied in 57 of the cases in order to optimize the treatment process (Group 1), and 63 patients were included and their outcomes followed as a control group (Group 2).We conducted a comparative analysis of results in both groups. The study's primary outcomes encompassed fertilization, cleavage embryo quality, blastocyst rate, and established clinical pregnancies. At day 1fertilization check we had 274/386 zygotes (71%FR) in group 1 and 132/410 in group 2 (32.2%FR), (P<0.0001). Twenty-two (34.9%) cycles in group 2 resulted in total fertilization failure (TFF). At the cleavage stage top-quality embryos from group 1 were significantly higher (P = 0.0021) in comparison to group 2. Forty-eight embryo transfers (ET) were performed in group 1 resulting in 41.67% clinical pregnancies versus 33 ET and only 4 pregnancies (12.12%) for group 2 (P = 0.0044). The results confirm the appropriateness of assisted oocyte activation as an additional method in cases of previous fertilization failure cycles.

The efficacy of orally administered L-carnitine in alleviating ovarian dysfunctions has laid the foundation for targeted in vivo use: a study employing self-control and propensity score matching.

This study aimed to evaluate the effectiveness of oral L-carnitine administration in patients after treatment failure to lay the groundwork for targeted in vivo use. A total of 515 In Vitro Fertilization (IVF) patients undergoing subsequent cycles were included after applying exclusion criteria. They were divided into a control group of 362 patients and a study group of 153 patients who received oral L-carnitine until oocyte retrieval.140 patients were matched according to maternal age, infertility duration, body mass index (BMI), day three top-quality embryos rate, by propensity score matching (PSM). The study investigated the relationship between L-carnitine treatment and in vivo oocyte maturation, normal fertilization, and subsequent embryo development. Following PSM, initial differences in BMI and Day3 top-quality embryo rate between groups were nullified, we created two comparable cohorts with highly similar characteristics. In the subsequent cycles, the study group showed significant improvements in in vivo oocyte maturation rate at retrieval (p=0.002), normal in vitro fertilization rate (p=0.003), blastocyst formation rate (p=0.003), and usable blastocyst rate compared to controls. Although there was no significant difference in the top-quality embryo rate on Day 3, the study group showed a 10% increase in the upper quartile (55.35% vs. 66.67%). The cumulative clinical pregnancy and live birth rates showed a significant improvement (59.82% vs. 68.42%,p=0.004, 47.41% vs. 59.80%, p=0.002). Furthermore, self-control analysis revealed substantial enhancements (p<0.001) in all outcome measures following L-carnitine administration, resulting in the birth of 74 healthy neonates without congenital anomalies. We theorized that daily oral intake of L-carnitine before oocyte retrieval could boost oocyte quality and embryonic development, thus improving IVF outcomes. Ongoing investigations hold the potential to offer valuable insights into the applications and mechanisms underlying the therapeutic effectiveness of L-carnitine.

Association of ambient PM2.5 and its components with in vitro fertilization outcomes: The modifying role of maternal dietary patterns

Despite the associations of dietary patterns and air pollution with human reproductive health have been demonstrated, the interaction of maternal preconception diet and PM2.5 and its components exposure on in vitro fertilization (IVF) treatment outcomes has not been investigated. A total of 2688 couples from an ongoing prospective cohort were included. Principle component analysis with varimax rotation was performed to determine dietary patterns. One-year and 85-day average PM2.5 and its components exposure levels before oocyte retrieval were estimated. Generalized linear regression models were conducted to assess the association of dietary patterns and PM2.5 and its components exposure with IVF outcomes. Interactive effects of dietary patterns on the association between PM2.5 and its components and IVF outcomes were evaluated by stratified analyses based on different dietary patterns. A positive association between the “Fruits-Vegetables-Dairy” pattern and normal fertilization (p-trend = 0.009), Day 3 available embryos (p-trend = 0.048), and top-quality embryos (p-trend = 0.041) was detected. Conversely, women with higher adherence to the “Puffed food-Bakery-Candy” pattern were less likely to achieve Day 3 available embryos (p-trend = 0.042) and top-quality embryos (p-trend = 0.030), clinical pregnancy (p-trend = 0.049), and live birth (p-trend = 0.020). Additionally, increased intake of animal organs and seafood improved the odds of live birth (p-trend = 0.048). Exposure to PM2.5, SO42-, organic matter (OM), and black carbon (BC) had adverse effects on embryo development and pregnancy outcomes. Furthermore, our findings indicated that the effects of PM2.5 components exposure on normal fertilization and embryo quality were modified by the “Grains-Tubers-Legumes”. Moreover, moderate intake of animal organs and seafood appeared to attenuate the effect of NO3- and NH4+ on the risk of early abortion. Our findings provide human evidence of the interaction between dietary patterns and PM2.5 exposure on IVF outcomes during preconception, implicating the potential for dietary interventions in infertile women to improve reproductive outcomes under conditions of unavoidable ambient air-pollutant exposure.

P-771 Does follicle size impact effect of follicular flushing on oocyte retrieval rate and embryology outcomes?

Abstract Study question Is the effect of follicular flushing on oocyte retrieval rate (RR) and embryology outcome influenced by follicular size? Summary answer Flushing improved RR in discordant growth with smaller(8-12 mm) follicles and larger(&amp;gt;22mm) follicle. Flushing did not impact embryological outcomes adversely. It increased top quality embryo What is known already Follicular flushing is done in poor responder for improving retrieval rate. There are very few studies in normal or hyper responders. Studies show controversial results of flushing, some showing benefits, while others, a detrimental effect because of increased intra-follicular pressure, increased procedure time, and change in paracrine milieu of oocytes by dilution with flushing medium No study has addressed whether increased retrieval rates with flushing are dependent on follicular size and indirectly maturity of oocyte. Most studies flushed with fixed volume of 1-2ml as size of follicle was not taken into account, making volume inadequate for completely flushing larger follicles. Study design, size, duration Prospective cohort study conducted from September 2022 to August 2023. Group 1: Study group- follicles were flushed after aspiration. Total patients were 120. They were divided into 3 groups according to follicular size and fluid volume aspirated with sample size calculated as A:306 small follicles 8-12 mm(&amp;lt;1.0ml), B:467 medium follicles 13-22 mm (1.0-6.0ml) C:102 large follicle &amp;gt; 22 mm (&amp;gt;6.0 ml). Group 2- Control group where follicles aspirated without flushing in 280 patients, 2572 follicles. Participants/materials, setting, methods Ethical approval taken. Inclusion - patients 21-40years of age undergoing oocyte retrieval for IVF/ICSI cycle with antagonist protocol Exclusion of oocyte of indeterminate origin ie one aspirate have two oocytes Group1 After aspiration of follicle with double lumen oocyte retrieval needle, follicle flushed with media volume equivalent to follicular volume. Group2 only follicular aspiration and volume measured Analysis done using SPSS-25 version. Primary outcome retrieval rate(RR) of oocytes. Secondary outcome: fertilization rate, percentage of top quality embryos(TQE)on day3&amp;5, blastocyst rate. Main results and the role of chance Flushing follicle significantly increased RR in smaller follicles(8-12mm) compared to controls 159/311 (51.12%)vs 176/780 (22.56%) p = &amp;lt;0.001 and also in larger follicles(&amp;gt;6ml) 76/109 (69.72%)vs 116/219 (52.96%) p = &amp;lt;0.01 No difference seen in optimum size follicles (13-22mm) 337/470 (71.7%)vs 1099/1573(69.86%) p = 0.44. Overall, there was statistically significant increase in RR with flushing 567/890 (63.7%)vs 1391/2572 (54.08%) p = &amp;lt;0.001. Significantly more time was taken for flushing vs control groups 16.50vs 12.15 minutes p = &amp;lt;0.001. Secondary outcomes like fertilization rate was not different with flushing 476/567 (83.95±15.01%)vs 1125/1391(80.88±19.01 %) p = 0.21. Percentage of TQE (day 3) were significantly more in flushing group 321/476 (67.43±25.41%)vs control 642/1125 (57.07±25.94%)p = &amp;lt;0.01. TQE (day 5) were significantly more in flushing group 142/476 (29.83±11.02)vs control 251/1125 (22.31±12.11%)p=0.04. Blastocyst rate remained similar in both groups 196/476 (41.18±16.73 %) vs 403/1125 ( 35.82±13.90 %) p = 0.12. Subgroup analysis showed that younger women (&amp;lt;35yrs)had a significantly higher RR with flushing 66.33±16.29%vs controls 57.88±13.68% (p = &amp;lt;0.01), compared to older women &amp;gt;35 years 60.29±16.37%vs 56.35±17.30% (p = 0.38). Women with higher AMH had better RR with flushing 63.71±15.11%vs controls 56.50±14.33 (p = 0.01) compared to women with lower AMH (&amp;lt;1 ng/ml) where flushing made no difference 63.47±19.59%vs 60.62±14.93(p = 0.79). Normal and hyper responders had significantly higher RR with flushing 66.16±15.11vs controls 57.53±13.85%(p = &amp;lt;0.001) compared to poor responders (&amp;lt;5 oocytes) 51.07±18.07vs 56.24±27.47(p = 0.55). Limitations, reasons for caution Although RR was better with flushing in small and large follicles we could not translate the advantage to pregnancy rate(PR) PR could not be calculated as some patients wanted 2 embryos transferred. Since patient had one each embryos from small and optimum follicles both were transferred. Further RCT required. Wider implications of the findings Practice of follicular flushing needs re-evaluation Women with discordant growth should be taken for flushing as it increases RR in small and large follicles Role of flushing should be widened beyond poor responders to younger women, with normal or hyper response increasing oocyte yield which may increase cumulative pregnancy rate Trial registration number not applicable

P-683 Progestins vs flexible GnRH antagonist for LH suppression during controlled ovarian stimulation in POSEIDON groups 3 and 4 patients: Analysis of effectiveness and cycle outcomes

Abstract Study question How do cycle outcomes differ between controlled ovarian stimulations using progestins vs flexible GnRH antagonists for LH suppression in women with diminished ovarian reserve (DOR)? Summary answer While progestin use is associated with significantly higher blastocyst rate than GnRH antagonists in POSEIDON group 4, the latter gives better results in group 3. What is known already Controlled ovarian stimulation (COS) in patients with DOR carries the additional risk of premature luteinizing hormone (LH) surge and ovulation.GnRH antagonists are popular for LH suppression in COS because of their convenient, quicker onset of action, flexibility to use GnRH agonist as a trigger and continue as double stimulation, allowing rapid pooling of embryos. Ease of oral administration, combined with cost-effectiveness and retention of all the benefits of GnRH antagonists makes progestins an attractive alternative to GnRH antagonists for COS in patients with DOR who often need multiple stimulation cycles to enable the pooling of embryos. Study design, size, duration An open-label, multicentric, randomized controlled study was conducted over 1 year from January 2023 to December 2023. 140 women aged 21- 42 years and with AMH &amp;lt;1.2 ng/dl were randomized into 2 groups in a 1:1 ratio, using either oral progestins or GnRH antagonists for LH suppression. The two groups were further comparative analyzed under POSEIDON 3 and 4 subgroups. Those with BMI &amp;gt;40kg/m2 and with partners having severely compromised semen parameters were excluded. Participants/materials, setting, methods In Group A (PPOS): Medroxyprogesterone acetate 10 mg orally was given daily with gonadotropins (hMG 300- 375 IU) from cycle day 2 until trigger. In Group B (Antagonist), patients had daily GnRH antagonist, cetrotide 0.25 mg, subcutaneous administration started when the dominant follicles reached 12-14 mm size during controlled ovarian stimulation. This was continued until the trigger. Following oocyte retrieval and ICSI, embryos were cultured until D5 and vitrified fortransfer in subsequent HRT cycles. Main results and the role of chance Patients in Group A, using PPOS, were noted to have significantly better fertilization rates than those in Group B, using flexible GnRH antagonists (0.82 + 0.2 vs 0.66 + 0.4, p-value &amp;lt;0.01). Though the number of top-quality embryos (TQE) frozen was not significantly different between groups A and B (p-value = 0.44), a significant difference in the outcome was noted when comparatively analyzed under the subgroups of POSEIDON groups 3 and 4. While patients in POSEIDON group 4 got significantly more top-quality embryos (TQE) when progestins were used for LH suppression (1 + 0.11 vs 0.77 + 0.41, p-value &amp;lt;0.01), those in POSEIDON group 3 got significantly more top-quality embryos when GnRH antagonist was used (0.94 vs 2, p-value &amp;lt;0.01). Patients in Group A and B were found to have no significant difference in the following outcomes: incidence of premature LH surge (4.3% vs 7.1%, p value= 0.71), number of days of stimulation (10.26 + 2.18 vs 10.26 +2.94, p value= 0.99), total gonadotropin dosage used (3448.64+ 1007.98 vs 3087.41 + 1260.56, p value=0.06), total number of oocytes retrieved (5.84 + 3.3 vs 4.9 + 3.4, p value=0.09) and total cycle cancellation rate (30.1% vs 35.7%, p value= 0.11). Limitations, reasons for caution The small sample size of the cases limits the current statistical power of the study. The results being encouraging need to be confirmed in a larger patient population and followed for a longer duration to enable calculation of implantation and clinical pregnancy rates for all patients. Wider implications of the findings Oral progestins are an effective and cheaper, patient-friendly alternative to injectable GnRH antagonists during COS in women with DOR. Considering the significantly higher TQEs formed, PPOS can be preferred in older, POSEIDON group 4 patients, whereas flexible GnRH antagonist protocol can be our choice in the younger, POSEIDON group 3. Trial registration number Not applicable

Co-administration of GnRH-agonist and hCG (double trigger) for final oocyte maturation increases the number of top-quality embryos in patients undergoing IVF/ICSI cycles

BackgroundThe utilization of a double trigger, involving the co-administration of gonadotropin-releasing hormone agonist (GnRH-a) and human chorionic gonadotropin (hCG) for final oocyte maturation, is emerging as a novel approach in gonadotropin-releasing hormone antagonist (GnRH-ant) protocols during controlled ovarian hyperstimulation (COH). This protocol involves administering GnRH-a and hCG 40 and 34 h prior to ovum pick-up (OPU), respectively. This treatment modality has been implemented in patients with low/poor oocytes yield. This study aimed to determine whether the double trigger could improve the number of top-quality embryos (TQEs) in patients with fewer than three TQEs.MethodsThe stimulation characteristics of 35 in vitro fertilization (IVF) cycles were analyzed. These cycles were triggered by the combination of hCG and GnRHa (double trigger cycles) and compared to the same patients’ previous IVF attempt, which utilized the hCG trigger (hCG trigger control cycles). The analysis involved cases who were admitted to our reproductive center between January 2018 and December 2022. In the hCG trigger control cycles, all 35 patients had fewer than three TQEs.ResultsPatients who received the double trigger cycles yielded a significantly higher number of 2PN cleavage embryos (3.54 ± 3.37 vs. 2.11 ± 2.15, P = 0.025), TQEs ( 2.23 ± 2.05 vs. 0.89 ± 0.99, P < 0.001), and a simultaneously higher proportion of the number of cleavage stage embryos (53.87% ± 31.38% vs. 39.80% ± 29.60%, P = 0.043), 2PN cleavage stage embryos (43.89% ± 33.01% vs. 27.22% ± 27.13%, P = 0.014), and TQEs (27.05% ± 26.26% vs. 14.19% ± 19.76%, P = 0.019) to the number of oocytes retrieved compared with the hCG trigger control cycles, respectively. The double trigger cycles achieved higher rates of cumulative clinical pregnancy (20.00% vs. 2.86%, P = 0.031), cumulative persistent pregnancy (14.29% vs. 0%, P < 0.001), and cumulative live birth (14.29% vs. 0%, P < 0.001) per stimulation cycle compared with the hCG trigger control cycles.ConclusionCo-administration of GnRH-agonist and hCG for final oocyte maturation, 40 and 34 h prior to OPU, respectively (double trigger) may be suggested as a valuable new regimen for treating patients with low TQE yield in previous hCG trigger IVF/intracytoplasmic sperm injection (ICSI) cycles.

P-763 Twin births in Belgium: a national cohort analysis of 35 523 double fresh embryo transfers between 2012 and 2021

Abstract Study question Should a national embryo transfer (ET) policy designed in the era of cleavage stage transfers be adapted for blastocyst transfers to avoid twin pregnancies? Summary answer Double embryo transfer policy results in unacceptable high twin rates at all ages and treatment ranks which is significantly more pronounced after double blastocyst transfer. What is known already In 2003, authorities in Belgium implemented a law to reduce multiple pregnancies after ART by tailoring ET policies based on age, embryo quality, and cycle rank. A choice between SET or Double Embryo Transfer (DET) depending on embryo quality is allowed in the second cycle for patients under 36y with SET being mandatory with top quality embryos. DET is allowed as of the first cycle for patients aged 36y or more regardless of embryo quality. Blastocyst transfer results in significantly higher live birth rates, implying an increased risk of multiple pregnancies after multiple blastocyst transfers. Study design, size, duration A analysis of 35,523 double embryo fresh transfer cycles over a period of 10 years from 2012 to 2021 from the Belgian national BELRAP (Belgian Registry on Assisted Procreation) registry was performed. The study included 26,930 double cleavage stage transfers (DET-CL) and 8,593 double blastocyst transfers (DET-BL). Participants/materials, setting, methods The primary outcomes were delivery rate (DR) per transfer and twin delivery rate (TDR). DR includes all deliveries with a least one live birth per transfer; the TDR includes all twin deliveries as a proportion of all deliveries. For analysis, the data were categorized into the following age groups: 18-25 years, 26-29 years, 30-35 years, 36-39 years and 40-45 years. Cycle ranks were categorized as follows: rank 1, rank 2 and cycle 3- ≥ 6. Main results and the role of chance For all age groups and cycle ranks, delivery rate per transfer and twin delivery rate per delivery were significantly higher after DET-Bl compared to DET-Cl (26.0% versus 19.9%; p &amp;lt; 0.0001 and 28.0% versus 20.6%; p &amp;lt; 0.0001, respectively). In the age groups 26-29 years, 30-35 years and 36-39 years, a significantly higher TDR was observed in DET-Bl compared to DET-Cl, respectively (34.4% versus 24.2%, p = 0.0005; 33.5% versus 24.9%, p &amp;lt; 0.0001 and 24.3% versus 17.3%, p = 0.0002). TDR after DET-Bl was significantly lower in women aged 40-45 years compared to all other age groups, 18-25 years, 26-29 years, 30-35 years and 36-39 years, respectively (11.3% versus 32.8%, p &amp;lt; 0.0001; 11.3% versus 34.4%; p &amp;lt; 0.0001; 11.3% versus 33.5%; p &amp;lt; 0.0001 and 11.3% versus 24.3%; p &amp;lt; 0.0001), respectively). DR was significantly lower in cycle rank 1 compared to cycle rank 2 (22.5% versus 25.5%; p &amp;lt; 0.0001) and compared to cycle rank 3 - &amp;gt; 6 rank 1 and (22.5% versus 27.1%; p &amp;lt; 0.0001). After DET-Bl, a significantly lower TDR was observed in cycle rank 1 compared to cycle rank 2 (15.6% versus 30.0%; p &amp;lt; 0.0001) and compared to cycle rank 3 - &amp;gt; 6 (15.6% versus 29.7%; p &amp;lt; 0.0001). Limitations, reasons for caution The retrospective design is a limitation. Outcomes might be influenced by clinical and laboratory improvements over the years. The registry does not collect embryo quality data making it impossible to assess the effect of embryo quality on the ET policy and outcome. Wider implications of the findings This data indicates that a double blastocyst transfer should only be considered from the age of 40 onwards. In the first cycle, regardless of age, we observe a significantly lower twin delivery rate, suggesting that a double ET is performed when only poor-quality blastocysts are available. Trial registration number Not applicable

P-532 Preimplantation genetic testing versus morphology as selection criteria for single embryo transfer in women aged 35-42: results of a pilot randomized controlled trial

Abstract Study question To assess the feasibility of running a large randomised controlled trial (RCT) evaluating the benefit of preimplantation genetic testing for aneuploidy (PGT-A) for embryo selection. Summary answer Running a PGT-A RCT is feasible. Considerations include participant recruitment, seven-day biopsy-embryologist availability and logistical complexities. What is known already Critics question the effectiveness of PGT-A leading to concerns about false positives and negatives, unnecessary embryo discarding and clinical effectiveness. Past studies have been criticised for only performing biopsy on top quality embryos, their criteria for randomization and the age of the population being assessed. Given these uncertainties there is a need for a rigorous RCT evaluating the role of PGT-A using current methodologies and biopsy techniques in women most affected by aneuploidy. Our pilot study is the first study where all embryos generated will undergo biopsy and ranked in terms in implantation potential prior to transfer. Study design, size, duration In this prospective, concealed allocation two-arm parallel RCT, our goal was to randomise 100 women with ≥3 good quality embryos on day 3 post egg collection into two groups: embryos screened using next-generation sequencing (NGS)-based PGT-A or selection-based on morphology alone. PGT-A patients underwent a trophectoderm biopsy on day 5/6/7 following egg collection. Biopsied embryos were frozen and subsequently transferred. Euploid and embryos described as being low mosaic (&amp;lt;30% abnormal cells) were prioritised for transfer. Participants/materials, setting, methods Women aged between 35–42 years undergoing IVF±ICSI were included, with those undergoing PGT for inherited genetic disorders, gamete donation cycles, untreated hydrosalpinges or untreated uterine abnormalities excluded. Primary objective during the pilot phase was to assess the ability to recruit, randomise, adherence to the study protocol and plan for a full study. Secondary outcomes were clinical pregnancy rate per embryo transferred, clinical pregnancy rate per couple randomised, miscarriage rate and time to pregnancy. Main results and the role of chance Out of 247 eligible women,100 were randomised, and 94 received the assigned treatment. The recruitment rate was 70.18%(173/247, 95%CI 61.65%-78.70%). An average of 6 participants were randomised each month. All randomised participants completed the follow-up. Our main recruitment challenges included 19% of eligible patients choosing to have PGT-A privately and 6% withdrawing their consent choosing to have a fresh transfer. Lack of biopsy-embryologist also meant patients whose potential biopsy would have been on a weekend had to be excluded in the first 4 months of our recruitment process. Baseline characteristics including age, BMI and cause of infertility were comparable in both arms and a total of 56 euploid, 53 aneuploid and 3 low mosaic embryos were identified. There was no statistically difference in CPR (23/50 vs 21/50, RR 1.09 95%CI:0.83-1.15, p-value&amp;gt;0.23) and miscarriage rate (3/50 vs 2/50, RR 1.5 95%CI:0.84-1.75, p-value: 0.15) per single embryo transfer or per woman randomised (23/46 vs 21/48, RR 1.14 95%CI 0.78-1.27, p-value:0.24 and 3/46 vs 2/48, RR 0.72 95%CI 0.58-1.17 p-value: 0.17) in either arm of the study. We cannot yet confirm the clinical benefit of PGT-A owing to the insufficient sample size in this pilot study. No low mosaic embryos were transferred. Limitations, reasons for caution The study was conducted in a single centre. If future definitive trials are conducted in multiple centres, the criteria for recruitment, randomisation, intervention, and follow-up still need to be unified. Many women aged above 40 failed to reach randomization due to challenges in producing three good-quality embryos. Wider implications of the findings The pilot study found that the eligibility rate is adequate however researchers should consider reviewing the conditions for randomisation in main study such as reducing the number of good embryos required to two. In addition, stratified randomization should be used in order to better balance female age within each group. Trial registration number NCT05009745

P-254 Optimizing clinical outcomes in failed ICSI Donor cycles: evaluating the efficacy of Assisted Oocyte Activation

Abstract Study question Does Assisted Oocyte Activation (AOA) confer benefits to couples undergoing donor ICSI cycles with a history of poor clinical outcomes and poor sperm quality? Summary answer AOA demonstrates efficacy in enhancing reproductive outcomes in donor ICSI cycles, in cases where oocyte activation deficiency (OAD) is attributed to a sperm factor. What is known already Oocyte activation is a spatial-temporal process caused by a series of intracellular calcium oscillations (Ca2+)triggered by the entry of sperm into the oocyte. Insufficient Ca2+ oscillations, primarily attributed to the absence of sperm-specific phospholipase C zeta (PLCζ) can result in OAD that leads to unsuccessful fertilization and poor embryo growth. Ionomycin and calcimycin, are the most common activating agents used to restart meiosis by raising intercellular Ca2+. As far as our knowledge extends, there aren’t studies reporting clinical outcomes on the use of AOA in patients who undergo cycles both without and with this treatment in donor oocytes. Study design, size, duration This study includes 38 couples undergoing ICSI cycles with donor oocytes from January 2020 to December 2023. After a previous cycle with a fertilization rate of &amp;lt; 50% and poor clinical outcomes, these couples underwent a subsequent attempt with AOA. Maternal age of recipients was 42±4.7 and 42±5 years in cycles without and with AOA (P=NS). Fertilization, blastocyst quality, implantation, clinical pregnancy rates (CPR) and pregnancy loss were compared between the cycles with and without AOA. Participants/materials, setting, methods Oocytes were exposed to a ready-to-use solution of Ca2+-ionophore A23187 (GM508 Cult-Active, Gynemed, Germany) for 15 min after sperm injection. Embryos were cultured in a time-lapse technology (Geri®) andevaluated according to the Gardner blastocyst grading system. Embryo transfer was performed on Day 5 or Day 6. Vitrified and fresh ejaculated sperm samples were used. T-test and Fisher’s Exact test were used for statistical analysis. P-value was considered statistically significant at a threshold of &amp;lt; 0.05. Main results and the role of chance A total of 38 men underwent ICSI cycles alongside their female partners, utilizing donor oocytes. The men’s average semen parameters were as follows: concentration of 15.3±25 x 106/mL, motility of 20.1±15%, and 2.0±1% morphology. In the initial 41 cycles without the AOA procedure, an average and standard deviation of 5.8±3 donor oocytes were injected, resulting in a fertilization rate of 33.9% (81/239) and a blastocyst rate of 30.8% (25/81). Among these cryopreserved embryos, 72% (18/25) were identified as top-quality, characterized by A and/or B quality of both inner cell mass and trophectoderm. Couples (N = 13) who underwent a transfer had an implantation rate of 19% (4/21), with CPR of 28.5% (4/14), which resulted in a pregnancy loss of 75% (3/4). Subsequently, these couples underwent 47 ICSI cycles with AOA, injecting an average of 5±1.7 donor oocytes (P=NS). This led to a significantly higher fertilization rate of 74.2% (176/237, P &amp;lt; 0.0001) and a blastocyst rate of 51.7% (91/176, P &amp;lt; 0.01), with 61% (56/91) identified as top-quality embryos (P=NS). Couples (N = 31) who underwent a transfer had a notable increase in implantation rate to 52% (25/48, P &amp;lt; 0.05), accompanied by a significant rise in CPR, reaching 71.4% (25/35, P &amp;lt; 0.01). Importantly, pregnancy loss decreased to 26% (6/23, P=NS). Limitations, reasons for caution The number of patients included in this study is limited. Although we excluded that OAD could be oocyte-related by using donor gamete, PLCζ or mouse oocyte activation tests were not performed to confirm sperm-related OAD. All patients, diagnosed with teratozoospermia, likely exhibit abnormalities causing PLCζlocalization issues or its absence. Wider implications of the findings In our study, sperm’s inability to generate accurate calcium signaling pathways correlates with poor clinical outcomes, even with donor oocytes. Consequently, for couples facing Oocyte Activation Deficiency attributed to sperm factors, implementing the AOA procedure significantly improves fertilization, blastocyst formation, implantation, and clinical pregnancy rates. Trial registration number na

P-359 Ten years’ experience using cryopreserved oocytes from In Vitro Maturation (IVM) or Controlled Ovarian Hyperstimulation (COH) cycles for fertility preservation in oncologic indications

Abstract Study question Is IVM a relevant option for fertility preservation in cancer patients, considering biological/clinical outcomes post-thawing, compared to COH? Summary answer This study shows that despite a significantly altered embryo morphology within the IVM group, the clinical outcomes remain comparable to those of COH. What is known already Fertility Preservation (FP) techniques are proposed to patients with cancer to provide the possibility of childbearing using their own gametes. Currently, oocyte vitrification after COH is the standard option. IVM serves as an alternative when COH is contraindicated, urgent cancer therapy is required, or in conjunction with ovarian cortex cryopreservation. Limited live births have been reported from frozen-thawed oocytes in IVM cycles of cancer survivors. Scarce data are available for comparing biological/clinical outcomes of frozen-thawed oocytes from IVM versus COH cycles in cancer survivors. Study design, size, duration This bicentric retrospective cohort study aimed to analyze the outcomes of all oocyte warming cycles in 73 cancer survivors having undergone oocyte vitrification for FP after COH and/or IVM. All of them had oocyte retrieval before the administration of gonadotoxic treatment and returned after being cured, with their oncologist agreement, for assisted reproduction treatments, between November 2012 and January 2024. Participants/materials, setting, methods Seventy-seven warming attempts followed by ICSI respectively from 37 COH and 40 IVM cycles were analyzed. Four patients benefited from an IVM and COH cycle. Survival, fertilization, top and good-quality embryos, defined at day-2 respectively as 4 and 3-5 adequate-sized blastomeres, without multinucleation with &amp;lt; 20% of cytoplasmic fragments, implantation, biochemical (hCG&amp;gt;100 UI/mL), clinical (intrauterine sac with fetal heartbeat) and live birth rates were compared using appropriate statistical tests. Main results and the role of chance The mean age and antral follicle count at the time of FP was similar in both groups. The indications for FP were breast cancer (n = 61), hematologic malignancies (n = 7), ovarian cancer (n = 3), other (n = 2). The number of cryopreserved oocytes tended to be higher in the COH group, when compared to the IVM group (8.5±7.3 vs 5.9±4.04; p = 0.16). The mean oocyte maturation rate after IVM was 68.6±23.1%. Oocyte survival, and fertilization rates were similar in COH and IVM groups, respectively (76.4±27.0% vs 76.1±22.0%; p = 0.76) and (67.1±34.3% vs 65.3±25.4%; p = 0.36). We reported a significantly better embryo quality at day-2 in the COH group when compared to the IVM group, with respectively top-quality embryos (44.0±34.2% vs 26.1±33.4%, p = 0,04) and good-quality embryos (62.2±38.0% vs 42.8±40.8%, p = 0.049), and also a significantly higher number of useful embryo in the COH group (2.7±2.6 vs 1.5±1.2, p = 0.03). Regarding clinical outcomes, the results were similar in COH and IVM groups, in terms of implantation rates (12.2±30.4% vs 15.5±35.6%; p = 0.86), biochemical (32.3% (10/31) vs 17.9% (6/28); p = 0.39), clinical pregnancies (25.8% (8/31) vs 17.9% (4/28); p = 0.54), live birth rates ( 25.8% (8/31) vs 14.3% (4/28); p = 0.34). Limitations, reasons for caution Statistical power to compare IVF outcomes after COH and IVM is limited by the few women who returned to use their frozen oocytes. The subjective nature of embryo morphology assessment. Wider implications of the findings This study is the largest evaluating frozen-warmed oocytes from IVM cycles in cancer patients, with 5 live births newly reported. A higher oocyte-yield may be necessary in IVM as the number of useful embryos is significantly lower. Novel approaches are being developed to improve maturation rates following IVM. Trial registration number Not applicable

P-530 Precise quantification of DNA in spent culture media sheds light on associations between DNA quantity, embryonic ploidy status and morphological quality

Abstract Study question Is it possible to accurately measure cell-free DNA (cfDNA) quantity in embryo spent culture media, and are amounts correlated with embryo morphological and/or chromosomal status? Summary answer A sensitive DNA quantification method was created, demonstrating that cfDNA quantity is not associated with embryo ploidy, but increases when embryos are of poor morphology. What is known already Preimplantation genetic testing involves embryo biopsy, necessitating specialist equipment and highly trained staff, increasing costs. Additionally, there have been concerns over potential embryo damage to the embryo resulting from biopsy. The discovery of cfDNA in medium in which embryos were cultured, led to suggestions that non-invasive PGT (niPGT) might be possible. However, the amount of cfDNA in media samples, and potential correlations with other aspects of embryo biology, require clarification. We aimed to provide the first accurate measurement of the amount of DNA in spent culture medium, providing insights into whether niPGT is likely to provide acceptable accuracy rates. Study design, size, duration This prospective analysis involved media samples from 151 fresh embryos, which were produced at a single IVF clinic over the course of one year. Embryos were cultured individually, with a medium change on day-3 of development. Trophectoderm biopsy was undertaken on day-5/-6, followed by PGT-A. Embryo morphology was assessed according to ASEBIR criteria (ranked A-C). Participants/materials, setting, methods The culture and biopsy protocols used were standard for the clinic. Media was collected prior to biopsy and underwent a novel quantitative whole genome amplification method, allowing precise measurement of DNA with respect to a standard curve generated using samples containing known amounts of DNA. PGT-A involved the use of a highly validated method, based upon next generation sequencing. Correlation between DNA quantity and embryo morphological quality was also studied. Data were analyzed using ANOVA. Main results and the role of chance There was no difference in the quantity of DNA in media samples associated with euploid and aneuploid embryos (mean DNA quantity and standard deviations were 1.72 ± 3.23 pg and 1.54 ± 3.66 pg for aneuploid and euploid embryos, respectively; p = 0.75). Overall, 91% of media samples contained less than the equivalent of one cell (&amp;lt;5pg of DNA) and almost two-thirds of samples (65%) contained less than 1pg of DNA. Given such low quantities of embryonic DNA, achieving accurate genetic testing is likely to be challenging. Drops that held embryos until day-6 had significantly higher levels of DNA than those associated with culture to day-5 (P &amp;lt; 0.0001), suggesting cfDNA accumulates with time. Thus, longer culture periods are likely to yield more DNA for analysis. Comparing the amount of cfDNA with morphological status revealed a significant association. Embryos of fair quality (C) tended to have higher quantities of DNA in spent culture medium, whereas top-quality embryos (classified as A or B) typically had less (mean and standard deviation of DNA quantity: 1.23 ± 3.14 pg for class A and B versus 2.60 ± 3.85 pg for class C; p &amp;lt; 0.05). This may reflect increased cell death and DNA release in poorer quality embryos. Limitations, reasons for caution This project is limited by its single-centre nature and relatively small sample size. A large-scale, multicentre study would be necessary to confirm these results. Wider implications of the findings It has been hypothesised that cfDNA quantity in culture medium may be predictive of embryo chromosomal status. We found no evidence to support this. Although DNA quantities were higher after culture to day-6, levels remained low, calling into question the ability of niPGT to deliver high accuracy diagnosis of embryos. Trial registration number not applicable

P-108 Automated sperm selection software (SiD) provides similar results as human selection for ICSI

Abstract Study question Automated sperm selection or human eye-based sperm selection, does it make a difference on IVF lab outcomes and clinical outcomes? Summary answer Single sperm selection by the AI SiD was as proficient as highly experienced embryologists and embryo development may have improved for inexperienced embryologists What is known already The implementation of artificial intelligence in reproductive medicine has proven effective for a variety of applications. One application for this technology is single sperm selection during ICSI. The computer assisted program SiD, was designed to grade sperm, in real-time, based on motility parameters transforming qualitative sperm assessments into a quantitative score. SiD’s assistance can be used to select the most optimal sperm for ICSI. The application of sperm selection by AI is expected to bypass user bias, fatigue, limited training, while also optimising time. However, there are limited studies correlating the usage of AI for sperm selection with ICSI outcomes. Study design, size, duration 1 year period 608 sibling MII oocytes were randomly divided in two groups; 1-ICSI group (n = 301): ICSI performed with sperm selected by the embryologist and 2- SID-ICSI group (n = 307): ICSI performed with sperm selected using SiD software. Priority was given to sperm classified as “Best” by SID software for ICSI.. Morphology selection was performed before ICSI in both groups. Embryos were cultured during 6 days in the same conditions. Participants/materials, setting, methods Sperm preparation was performed using a microfluidic chamber (Zymot™). Sperm selection was performed as previously described, and ICSI was performed following routine procedures. Biological and clinical outcomes were analysed in both groups and compared including fertilization rate, cleavage rate, embryo and blastocyst development rate, top quality embryo rate, blastocyst development rate and pregnancy rates. Biological outcomes in both groups were also compared based on embryologist seniority. Embryo morphokinetics parameters were analysed using time-lapse system. Main results and the role of chance There was a non significant trend towards better outcomes in the SID-ICSI group for all biological outcomes including fertilization rate (83,1% vs 82,4%), cleavage rate (97,6% vs 97,2%), day 3 embryo development rate (72,9% vs 70,6%), blastocyst development rate at day 5 (49,0% vs 44,8 %), good quality blastocyst development rate at day 5 (45,1% vs 41,5%), and top quality blastocyst development rate at day 5 (25,9% vs 22,2%). Similarly, a non-significant increase was observed in all these outcomes when sperm selection is performed by a junior embryologist. Embryo development was monitored using a timelapse system. Our data show that 2nd polar body extrusion (tPB2) (4,0h vs 4,8 h [p = 0,005]) and early cleavage (t2) (27,3h vs 28,7h [p = 0,05]) happen significantly earlier when SiD is used for ICSI. No significant difference was observed for all other fertilization events (cytoplasmic wave,tPN1,tPN2,presence of cytoplasmic halo,tPNf,and cytoplasmic halo disappearance), and other cleavage timings (t3-&amp;gt;t10,tM,tSC,tEC,tSB), and embryonic cell cycles (ECC1,ECC2,ECC3,s2,s3). Euploid rate obtained with in the 2 groups was comparable (53,3% in SID-ICSI vs 44,4% in ICSI). Although there was a trend toward higher cumulative early and clinical pregnancy rate after ICSI-SID, the difference did not reach significance (43,1%vs34,0% and 27,5%vs21,2%, respectively). Limitations, reasons for caution The ZymotTM microfluidic chamber was used for sperm preparation. This device optimises sperm selection, giving rise to an already optimised population of spermatozoa. Future investigations evaluating the effect of SiD automated sperm selection software on biological outcomes in samples prepared with a less effective technique would potentially yield conversing results. Wider implications of the findings This pilot study demonstrated that the usage of the automated sperm selection software SiD gives rise to similar biological outcomes and embryo morphokinetics, indicating its effectiveness in sperm selection. We expect this program would be useful in the presence of less experienced laboratory members helping by standardizing sperm selection. Trial registration number NA

P-402 Comparison of uterine contractions in artificial hormone treatment versus aromatase inhibitor stimulated frozen embryo transfer cycles

Abstract Study question does uterine contractions differ between aromatase inhibitor stimulated (AIS) or artificial hormone treatment (AHT) Frozen Embryo transfer (FET) cycles Summary answer uterine contraction activity did not show any difference between artificial hormone treatment and aromatase inhibitor stimulated FET cycles What is known already FET cycles are prepared by different protocols. aromatase inhibitor stimulated (AIS) or artificial hormone treatment (AHT) protocols are two common protocols among others. These protocols have different cycle dynamics and might differ in pregnancy rates and different effects on perinatal outcomes. Although embryo related factors are most important for implantation endometrial receptivity and therefore uterine contractions might have a role in implantation. English literature provides scarce data on uterine contractions and pregnancy outcomes. Presence of prominent contractions were related to poor pregnancy outcomes in some studies whereas other few studies failed to show any effect in this sense Study design, size, duration this study is an observational cohort study of successive frozen embryo transfer (FET) cycles of eligible patients between 1st of july 2023 to 31th of October 2023 in a tertiary university hospital in vitro fertilization clinic with ultrasonographic video recordings at transfer day. During study period of four months, 147 out of 265 consecutive FET cycles were included in this study and study is approved by local ethical committee of the affiliated university (KA23/388). Participants/materials, setting, methods Total study group was composed of 99 artificial hormone treatment cycles and 48 aromatase inhibitor stimulated FET cycles after exclusion of patients with uterine congenital malformations, uterine fibroids,adenomyosis patients and patient with previous uterine surgery. Implantation rates, pregnancy rates and clinical pregnancy rates were recorded. Transvaginal ultrasonographic sagittal uterine video image recordings were obtained at transfer day one hour before embryo transfer. Recordings were reviewed by two researchers and uterine contractions were documented by agreement. Main results and the role of chance This is the first study comparing different FET cycles and uterine contraction activities. Patient characteristics were similar between two groups. Artificial hormone treatment and aromatase inhibitor stimulated cycle groups showed similar endometrial thicknesses (11.47±1.92 and 11.24±1.02, respectively p = 0.42) and top quality embryo transfer rates (22.6 % and 26.8% p = 0.82). Although AIH group showed a statistically insignificant advantage, clinical and ongoing pregnancy rates were similar between protocols (supplementary data). Uterine peristaltic activity was not statistically different between aromataze inhibitor stimulated and artificial hormone treatment FET cycle (1.54±1.75 versus 1.63±2.15 p = 0.81 recpectively). Furthermore uterine persistalsizm were similar in pregnant and non-pregnant patients /1.52±2.07 vs 1.72±1.93 p = 0.57). However as far as Aromatase inhibitor stimulated FET cycles were considered; pregnant patients had less uterine peristaltic activity when compared ton non-pregnant patient (45.7% and 15.4%, respectively p = 0.054 ) although this difference was at the border of statistical significance. Limitations, reasons for caution This is an observational study of relatively short duration with limited number of patients. Our study in AIS cycles have a tendency to fewer contractions in cycles ended up in pregnancy. Larger series of FET cycles might prove differences in uterine contraction frequencies and pregnancy outcomes in AIS FET cycles. Wider implications of the findings Our study supported new information on uterine contractility in different FET cycles protocols. Further large scale studies might provide new insights to uterine factor and its role in implantation as far as uterine contraactions are concerned. Trial registration number (KA23/388) local ethic committee approval

P-585 Vaginal progesterone associated with oral dydrogesterone versus vaginal progesterone for luteal support in letrozole-hcg cycles for thawed embryo transfer: a non-inferiority prospective cohort study

Abstract Study question Does micronized vaginal progesterone (MVP) associated with oral dydrogesterone (DYG) an alternative for luteal support in Letrozole-hcg cycles for thawed embryo transfer? Summary answer The association of MVP and DYG is a non-inferior alternative to only MVP for luteal support in thawed embryo transfer in Letrozole-hcg cycles. What is known already Endometrial preparation is essential for thawed embryo transfer. Several protocols involving different luteal support with progesterones are available. Moreover, the presence of corpus luteum decreases the obstetrical morbidity mainly linked to hypertensive disorders. Letrozole-hcg administration is a good alternative for frozen embryo transfer (FET). However, the literature lacks information about the comparison among the different protocols for luteal support in Letrozole-hcg cycles. Dydrogesterone is an oral progesterone with some unique characteristics which could be utilized with some advantage for luteal support, including oral administration and some immunological activity. Study design, size, duration This is a prospective non-inferiority cohort study performed at an infertility clinic including patients performing the first FET cycle during 2023. Reproductive outcomes were compared between patients receiving 600 mg (3x200 mg) of MPV or 40 mg of DYG plus 400 mg of MVP after Letrozole-hcg administration. Participants/materials, setting, methods All patients (aged 23-39 years) undergoing their first FET cycle during 2023 were included in this study (n = 169), divided into two groups: MVP+DYG (n = 54) or MVP (n = 115) for luteal support after Letrozole-hcg. Letrozole was administered for 5 days (2,5 mg/bid) and hcg was done when a dominant follicle reached 18 mm with a follicular endometrium measuring at least 7 mm. Main results and the role of chance Both groups were equivalent in terms of age, infertility aetiology, BMI and AMH (P &amp;gt; 0.05). Moreover, the proportion of cleavage or blastocyst stage was also similar between both groups (57% blastocyst transfer for MVP+DYG and 50% blastocyst transfer for MVP group, P &amp;gt; 0.05). The number of transferred embryos was (mean ±SD) 1.6±0.5 for both groups, P &amp;gt; 0.05. Furthermore, the number of top-quality embryos (cleavage stage or blastocyst) was also similar, P &amp;gt; 0.05. Moreover, clinical pregnancy (&amp;gt; 12 weeks) was 42% in MVP+DYG group versus 31% in the MVP group (-0.22-1.27, P = 0.156) and the early pregnancy loss per positive hcg test did not differ (18% for MVP+DGY and 22% for MVP, P = 0.687). In addition, we performed a multivariable analysis controlling possible confounders for clinical pregnancy: BMI, age, AMH, embryo quality, infertility aetiology and embryo stage during the transfer, and only age (0.86; 95%CI -0.229 to -0.064, P = 0.001) was related to clinical pregnancy. Limitations, reasons for caution The main limitation is the cohort design of our study, with an inherent risk of bias (selection bias). Moreover, we calculated this study to achieve the power for a non-inferiority comparison, not superiority comparison. Wider implications of the findings This is the first study comparing MVP+DYG to only MVP in patients submitted to FET using the Letrozole-hcg protocol. We demonstrated in this non-inferiority study the usefulness of adding DYG. Moreover, our results were promising showing a trend towards better pregnancy rates after MVP+DYG. Trial registration number not applicable

P-251 Denudation time is crucial to increase the proportion of top-quality embryos in egg donation cycles

Abstract Study question How do different denudation times affect in the efficiency of the egg donation cycles? Summary answer Undetected in vitro matured oocytes, resulting from delayed denudations, contribute to a decreased top-quality blastocyst ratio. What is known already While the timeframe for sperm microinjection in ICSI cycles is widely agreed upon (2-6 hours post-oocyte retrieval), no definitive denudation time preference has been established. Published studies diverge on the advantages of early or late denudation, assessing the impact of trigger-to-denudation time (T-D time) on oocyte maturation. Advocates of early denudation contend it prevents oocyte aging, reducing in vitro culture and enhancing outcomes. These studies, often using data from cycles with own oocytes, exhibit high variability and inconsistent conclusions. The present work introduces an alternative hypothesis on the benefits of early denudation, employing a more stable study model. Study design, size, duration This retrospective study encompasses oocyte donation cycles, employing a short antagonist protocol and ovum pick-up 35.5 hours post-trigger, conducted from January 2019 to July 2023 including 1,559 stimulations. Additionally, 1,275 ICSI cycles without detected sperm pathologies derived from these stimulations were examined to evaluate the influence of T-D time on the reproductive outcomes. Six comparable groups were established by segmenting the sample according to T-D time, annotated in 30-minute intervals (36.5h to 39h). Participants/materials, setting, methods Rates of MII, MI, and nonviable oocytes were examined among 1,559 stimulations resulting in 29,671 oocytes (36.5h:187-stimulations/3,178-oocytes; 37h:226/3,836; 37.5h:614/11,921; 38h:329/6,884; 38.5h:119/2,322; 39h:84/1,530). Additionally, in 1,275 ICSI cycles involving 11,333 MII-oocytes (36.5h:117-ICSI cycles/1,027-MII; 37h:130/1,133; 37.5h:502/4,426; 38h:316/2,925; 38.5h:120/1,041; 39h:90/781), comparisons were made for fertilization, abnormal fertilization, ICSI degeneration, useful day 5/6 blastocyst (transferred or cryopreserved; &amp;gt;3BB-top and ≤3BB-good quality by Gardner score), pregnancy, and live birth rates. Statistics were ANOVA, Chi-square test and Pearson’s correlation. Main results and the role of chance Oocyte maturation profile assessment reveals that groups with later denudation exhibit a higher proportion of MII-oocytes (36.5h:80.71%; 37h:81.04%; 37.5h:81.26%; 38h:80.83%; 38.5h:84.84%; 39h:85.29%; r:0.8475; p &amp;lt; 0.05), reaching significance from 38.5h onward (p &amp;lt; 0.05). This is complemented by a strong negative correlation between T-D time and MI rate (36.5h:5.51%; 37h:5.32%; 37.5h:4.64%; 38h:4.17%; 38.5%h:3.27%; 39h:2.94%; r:-0.9881; p &amp;lt; 0.05), with significant declines at 38h and 38.5h (p &amp;lt; 0.05). No correlation between useless oocytes proportion and T-D time was observed (r:-0.5983; p = 0.21). This illustrates that certain MI-collected oocytes progressively mature during in vitro culture and subsequently, upon denudation, are classified as MII. This overestimates in vivo-matured oocytes. No differences attributed to T-D time were observed regarding fertilization, abnormal fertilization, or ICSI degeneration rates. While time does not affect the rate of useful embryos achieved (36.5h:49.4%; 37h:46.4%; 37.5h:44.6%; 38h:48.0%; 38.5h:50.6%; 39h:47.2%; r:0.1342; p = 0.80), it has an impact on the blastocysts quality. The proportion of top-quality blastocysts (&amp;gt;BB) progressively decreases with T-D time (36.5h:86.2%; 37h:85.2%; 37.5h:85.4%; 38h:81.2%; 38,5h:80.5%; 39h:80.5%; r:-0.925; p &amp;lt; 0.05). Out of 1,794 single embryo transfers, 1,477 have conclusive outcome. No implication of T-D time was detected in pregnancy rate (36.5h:59.88%; 37h:64.07%; 37.5h:61.54%; 38h:65.02%; 38.5h:59.55%; 39h:62.26%; r:0.0489; p = 0.93) or live birth rate (36.5h:40.31%; 37h:51.85%; 37.5h:39.12%; 38h:45.04%; 38.5h:41.83%; 39h:37.96%; r:-0.375; p = 0.46). Limitations, reasons for caution This study has inherent limitations as a retrospective analysis. Additionally, the potential effects of a denudation earlier than 36.5 hours post-trigger, not yet standardized in the laboratory, were not assessed. Wider implications of the findings Delayed denudation may benefit autologous IVF by fostering in vitro oocyte maturation. In egg donation cycles, early denudation is preferable for stringent selection of in vivo matured oocytes, yielding superior results. Additionally, shortening or eliminating incubation periods would enhance the automated sequence of diverse processes during an IVF cycle. Trial registration number not applicable

O-127 Blastocyst expansion speed assay: a putative artificial intelligence-powered tool for embryo selection. Retrospective study involving 2184 blastocysts and 786 PGT-A cycles

Abstract Study question Is blastocyst expansion-speed, examined through Artificial-Intelligence (AI) between time of blastulation (tB) and time of expanding blastocyst (tEB), associated with embryo blastulation and reproductive competence? Summary answer Blastocyst expansion-speed-assay (ESA) was significantly associated with euploidy and live-birth rates, being discordant with clinical embryologists’ ranking in 50% of embryonic cohorts. What is known already Blastocyst expansion is one of the earliest morphogenetic events in mammalian development. It is essential for the establishment of the blastocyst layout and requires trophectoderm integrity. Several studies suggested that expansion is a valuable feature for ranking and prioritizing blastocysts for transfer based on their developmental competence. Time-lapse technology (TLT) implementation in IVF allows a deeper understanding of blastocyst expansion dynamics. In this study, we combined TLT and AI to develop a blastocyst-ESA and investigate its association with embryo blastulation and reproductive competence. Study design, size, duration Retrospective study including 2184 blastocysts cultured in EmbryoScope incubators during 786 PGT-A cycles across 2013-2020. Videos were analyzed through an AI-powered tool (CHLOETM, Fairtility). The expansion-speed was calculated as [(Δ embryo proper area at tEB – embryo proper area at tB) / (Δ tEB - tB)]. ESA was tested for its association with embryo quality, euploidy and live-birth rate (LBR) in 548 vitrified-warmed single euploid transfers. A simulation of ESA putative clinical effectiveness was conducted. Participants/materials, setting, methods ICSI, trophectoderm biopsy of fully-expanded blastocysts without day3 zona-drilling, and qPCR/NGS to assess full-chromosome uniform aneuploidies were performed. tB and tEB, expressed as hours-post-insemination (hpi), embryo proper area (in µm2) at both these stages, and blastocyst quality (EQ-Score from 0 to 1) were all automatically recorded through CHLOE. Possible confounders (e.g., maternal age, male factor, and cause of infertility) were considered. Regression analyses were conducted to adjust the data. Main results and the role of chance The average ESA was 705±458 µm2/hour. Higher ESA was associated with higher EQ-Score. Euploid blastocysts showed higher ESA than aneuploid (761±465 µm2/hour versus 667±449 µm2/hour; p &amp;lt; 0.01). ESA was also associated with LBR per euploid blastocyst transfer (LB: 873±438 µm2/hour versus 736±467 µm2/hour; p &amp;lt; 0.01). Of note, maternal age showed no association with ESA. Multivariate regressions outlined a + 5.1%-increase in the chance of euploidy and a + 6.3%-increase in chance of LB, every +100 µm2/hour-increase in ESA. ESA and embryologists were discordant in grading top-quality embryos in each cohort in 50% of the cases. In 59% of the 352 cohorts where both euploid and aneuploid blastocysts were obtained, ESA would have ranked the former embryos as top-quality. In the 216 cycles with ≥2 euploid blastocysts obtained and ≥1 transferred, ESA would have prioritized (i) a competent embryo in 46% of the cases, (ii) an incompetent embryo in 20%, but (iii) in 33% its value could not be assessed (i.e., the top embryo according to ESA has not been transferred yet). When compared to the embryologists, ESA would have been (i) equally-effective in 49% of the cases, (ii) more effective in 12-24%, and (iii) less effective in 5%-26%. Limitations, reasons for caution Retrospective single-center study. Both continuous and sequential media were used, although they were not associated with the outcomes under investigation. To assess the clinical value of ESA, a prospective study among first transfers is warranted. Wider implications of the findings ESA is an automated, unbiased, and easily-applicable measurement that certainly deserves further appraisal. If validated in prospective studies, AI-based selection algorithms could include this assessment to increase their predictive power. Trial registration number None

P-547 Impact of SARS-COV-2 infection on embryo euploidy rate: a retrospective cohort study of 498 PGT-A cycles in China

Abstract Study question Does SARS-CoV-2 infection before oocyte retrieval have any impacts on embryo euploidy rate? Summary answer SARS-CoV-2 infection before oocyte retrieval had no detrimental impact on embryo euploidy rate, indicating that cycle cancellation due to SARS-CoV-2 infection may not be necessary. What is known already SARS-CoV-2 infection has significant effects on the health of people worldwide and poses a serious threat to reproductive health. According to the previous studies, SARS-CoV-2 infection had negative impacts on the blastocyst formation rate, top-quality embryos rate. However, the association between the infection and the embryo euploidy rate is indeterminate. It is still worth exploring the impact of SARS-CoV-2 infection on embryo euploidy rate. Especially this is the key question to be addressed for providing appropriate counseling to couples seeking for preimplantation genetic testing for aneuploidy (PGT-A) treatments during the ongoing pandemic. Study design, size, duration This was a retrospective cohort study of 498 couples who underwent PGT-A cycles with next-generation sequencing technology from November 1st, 2022 to June 30th, 2023 at a tertiary-care medical center in China. Participants/materials, setting, methods Women tested positive for SARS-CoV-2 before oocyte retrieval were included in the COVID-19 group, while patients who were asymptomatic and tested negative during the same period were included in the non-COVID-19 group. The primary outcome was the euploidy rate. Subgroup analysis was conducted according to the time interval between infection and oocyte retrieval. Univariate and multivariate logistic regression models were applied to adjust for potential confounders. Main results and the role of chance A total of 278 and 220 women were included in the COVID-19 and non-COVID-19 group, respectively. In the COVID-19 group, the maturated oocyte rate (79.5% vs. 83.7%, p＜0.001), number of total viable embryos (2 vs. 3, p=0.038), and number of euploid blastocyst (1 vs. 1, p=0.028) were significantly decreased compared with non-COVID-19 group. The euploidy rate (38.5% vs.40.3%, p = 0.451), aneuploidy rate (45.0% vs. 43.8%, p=0.616), and mosaicism rate (16.5% vs. 15.9%, p=0.746) were all comparable between the two groups. Results of multivariate logistic regressions revealed no association between SARS-CoV-2 infection and euploidy rate, aneuploidy rate, or mosaicism rate. Similarly, no association was detected between the time interval of infection and oocyte retrieval (≤30 days, 31-60days, 61-90 days, 91-120 days, or ≥ 120 days prior to oocyte retrieval) and euploidy rate. Limitations, reasons for caution The retrospective nature of the study and limited sample size for subgroup analysis limits the generalizability of the findings. Other potential confounders, such as the level of SARS-CoV-2 antibody, symptoms of infection, are needed to be taken into consideration as well. Large-scale prospective studies are warranted to confirm the conclusion. Wider implications of the findings Our study demonstrated that SARS-CoV-2 infection before oocyte retrieval had no detrimental impact on embryo euploidy rate, indicating that cycle cancellation due to SARS-CoV-2 infection may not be necessary. Trial registration number Not Applicable

P-668 Progesterone levels on the thawed embryo transfer day in artificial cycles with vaginal progesterone are not a predictor of clinical pregnancy: a prospective study

Abstract Study question Are serum progesterone (P) levels on the thawed embryo transfer day a good predictor of clinical pregnancy with own oocytes in artificial endometrial preparation cycles? Summary answer In artificial endometrial preparation cycles, serum P levels on the thawed embryo transfer day are not a good predictor of clinical pregnancy with own oocytes. What is known already Some studies have shown that low serum P levels on the thawed embryo transfer (ET) day are associated with a lower pregnancy rate after artificial endometrial preparation (AEP) cycles. Other studies have shown that serum P is not a good predictor of pregnancy rates. Furthermore, studies are inconclusive about which cutoff level for P on ET day correlates with better gestational outcomes. Study design, size, duration Prospective cohort study with 235 patients who underwent ET after AEP cycle with estradiol valerate and vaginal micronized P (200 mg/ 8 h). All patients underwent blastocyst-stage or cleavage-stage ET with embryos from their own oocytes, without preimplantation genetic testing for aneuploidies, after AEP cycle, with a triple layer endometrium &amp;gt; 7 mm. The study was carried out at the Human Reproduction Center of Ribeirão Preto Clinical Hospital from November 2021 to June 2023. Participants/materials, setting, methods Blood was collected between 8–10 am on the thawed ET day and serum P concentration was measured. The primary outcome was clinical pregnancy (CP) beyond pregnancy week 8. A ROC curve detected the best P cutoff level that correlated with higher CP rate. Thereafter, patients were categorized into those with serum P above and below the cutoff level. Multivariate logistic regression was performed to verify which variables were predictors of CP and P levels. Main results and the role of chance Serum P measured on the ET day showed no significant difference between patients who had CP and those who did not (12.76 +4.1 versus 11.9 + 5.6, p 0.3318). The cutoff level for serum P measured on the ET day above which correlates with higher CP rate was 10.6 ng/mL (AUC 0.58), with a sensibility of 72% and specificity of 44%. Patients who had serum P on the day of ET above this value had a significantly higher CP rate than those who had it below this value (25.9% versus 14.6%) (p 0.0372). However, when the multivariate logistic regression model was performed, there was no significant difference in the CP between the groups (OR 2, 95%IC: 0.9-5). Predictors of CP in the study were the presence of at least one top-quality embryo transferred (OR 2.6, 95%IC: 1-6.6) and the transfer of embryo at the blastocyst-stage (OR 5.2, 95%IC: 1.9-14). The presence of endometriosis was negatively associated with CP (OR 0.2, 95%IC: 0.09-0.9). Limitations, reasons for caution The small sample size is a limitation. We included the blastocyst-stage and the cleavage-stage embryo transfer. Only women with vaginal micronized P were included. Extrapolation to other P administration forms needs to be evaluated. Wider implications of the findings Monitoring serum P levels on the day of ET in AEP cycles with vaginal micronized P is not justified as serum P levels are not a good predictor of CP in cycles with own oocytes. Trial registration number Not applicable