Multicenter Study Research Articles

P-510 The impact of the severe male factor in the egg donation cycles using preimplantation genetic test for aneuploidy

Abstract Study question Does severe male factor affect the euploidy rate and IVF outcomes in donor egg ICSI cycles using PGT-A? Summary answer Severe male factor affects the euploidy rate in egg donation cycles, however, no statistical differences were reported in live birth and cumulative live birth rate. What is known already Around 50% of couples seeking fertility treatment have male-factor infertility. Intracytoplasmic sperm injection (ICSI) relies on evaluating sperm morphology, but the direct relationship between semen quality and embryonic potential remains unclear. Evidence indicates that poor semen characteristics correlate with an increase in aneuploidy rate of spermatozoa. Therefore, pregenetic testing for aneuploidies (PGT-A) is recommended in cases of male factor infertility. The relationship between aneuploidy rates and abnormal sperm remains a debated topic. The use of the egg donation model offers a valuable chance to isolate the male-related factor and evaluate its influence on the occurrence of embryonic aneuploidies. Study design, size, duration This is a retrospective, observational, cohort study including all European IVI clinics. The study included egg donation cycles using pre-implantation genetic testing for aneuploidies (PGT-A) generated in IVI clinics from January 2017 until December 2023. 690 egg donation cycles were included in the study, N = 202 in the group with sperm < 5mil/ml, N = 102 in the sperm group with 5-15 mil/ml and N = 386 in the group with sperm >15 mil/ml. Participants/materials, setting, methods The study included women between 30-49 years undergoing egg-donor cycles, using fresh/frozen sperm Sperm samples were derived after 2-5 days of abstinence. The cycles of egg donation were divided into three groups according to sperm concentration, severe oligozoospermia (SO, < 5mil/ml), moderate oligozoospermia (MO, between 5-15 mil/ml), and control group(>15 mil/ml). Trophectoderm biopsy was performed on day 5 or 6 and subsequent analysis used the next sequencing generation (NGS). Single euploid blastocyst transfers were included. Main results and the role of chance The fertilization rate was decreased in SO (73.33 ± 16.19) than in the MO group (79.07 ±12.45) and control group (77.80 ±14.72) p = 0.0047. The number of blastocysts was higher in the control (5.80 ± 2.72) and MO (5.97± 2.78) than in the SO group (4.99 ± 2.40), p = <0.001. The number of euploid blastocysts was decreased in SO (3.21 ± 2.07) than in the control (4.09 ± 2.41) and MO group (4.05 ± 2.15), p = <0.001. The euploidy rate of SO decreased (63.32 ±26.39) compared to the control (69.67 ± 25.08) and MO group (68.55 ± 20.37), p = 0.012. The logistic regression model included possible confounders (male age, age donor, male BMI, sperm motility, sperm morphology and sperm concentration) that showed no effect on the euploidy rate. The pregnancy rate was similar across the groups 69.77% vs 66.05% vs 66.23%, p = 0.54. No statistical differences were reported for the clinical pregnancy rate 62.79% vs 61.73% vs 58.40%, p = 0.44, and the live birth rate 62.79% vs 61.73% vs 58.40%, p = 0.44. 48.05 vs 46.38% vs 42.64%, p = 0.37. No differences were reported in the miscarriage rate (p = 0.37) and CLBR (p = 0.26). Limitations, reasons for caution The retrospective nature of the study and the sample size represent the principal limitations of the study. The clinical reason for severe male factors was not considered in the study. Wider implications of the findings SO affects the quality affect the early embryonic potential through an increase in aneuploidy, despite no clinical effects.The use of PGT-A in case of severe male factor remains not justified in the egg donation.Future prospective multi-center studies are needed to highlight the effect of sperm quality on early embryonic potential. Trial registration number not applicable

P-383 Development and validation of a visualized prediction model for early miscarriage risk in patients undergoing IVF/ICSI procedures: a real-world multi-center study

Abstract Study question What are the key predictors of early miscarriage in patients undergoing IVF/ICSI treatment? Summary answer This study identified risk factors for early miscarriage in IVF/ICSI patients, and developed a validated predictive nomogram, highlighting age-dependent factor variations. What is known already Early miscarriage, occurring in 15-20% of confirmed pregnancies, can lead to complications, psychological distress, and fertility impairment. It is especially prevalent during the first trimester, with up to 80% of all miscarriages happening during this phase. Women who conceive via Assisted Reproductive Technology (ART) face higher stress levels and increased risk of miscarriage. Although many contributing factors to early miscarriage have been identified, there is a scarcity of integrated predictive models for IVF/ICSI patients. This paper aims to develop a validated, user-friendly nomogram to predict first-trimester miscarriage risk in IVF/ICSI patients and help clinicians and patients make informed decisions. Study design, size, duration A retrospective analysis was conducted on 20,322 first cycles out of 31,307 for IVF/ICSI at a reproductive centre in Southern China, After excluding ineligible cycles, 6,724 first fresh cycles were included and randomly divided into a training dataset (n = 4,516) and an internal validation dataset (n = 2,208). An external validation dataset (n = 1,179) was obtained from a reproductive center in Northern China, further ensuring the robustness and generalizability of the findings. Participants/materials, setting, methods This retrospective study analysed 6,724 out of 31,307 IVF/ICSI treatments from 2011 to 2020 at a Southern China reproductive center and 1,179 cycles from a Northern China center (2013-2018) for validation. Early miscarriage was defined as a loss before 12 weeks’ gestation. Patients were divided into two age groups: ≥35 and <35 years. The study followed the TRIPOD statement and received Institutional Review Board approval. Main results and the role of chance The study found statistically significant differences across the training, internal validation, and external validation sets in terms of patient age, BMI, number of good quality embryos, antral follicle count (AFC), basal testosterone (T), luteinizing hormone (LH) levels, among other variables. The early miscarriage rates were 9.43%, 10.55%, and 13.50% across these sets. Elder women (age ≥35 years) had a higher duration of infertility and early miscarriage rate, but lower AFC and numbers of retrieved oocytes, 2PN, and MII oocytes compared to younger women (age <35 years). A nomogram prediction model was developed using variables such as female age, BMI, number of spontaneous abortions, number of induced abortions, and basal FSH, which were found to be independent prognostic factors for early miscarriage. The model’s performance was validated using AUC, calibration plots, and DCA, indicating acceptable performance and good agreement between prediction and observation. The model was more beneficial for predicting miscarriage risk than the treat-all or treat-none schemes. Subgroup analysis for elder and younger women showed different risk factors for miscarriage, notably female age, basal FSH for elder women, and female age, BMI, and the number of spontaneous abortions for younger women. The subgroup models also showed consistent results and good clinical relevance. Limitations, reasons for caution The retrospective design of our study may have introduced certain limitations and potential biases. To overcome these limitations, it is advisable that future research endeavors adopt a prospective clinical study design. Wider implications of the findings This study uses a diverse dataset and rigorous methods to create a user-friendly prediction model for early miscarriage in ART. By considering regional differences, it ensures the model’s broader applicability and accuracy, enabling personalized treatment strategies and improving infertility treatment outcomes. Trial registration number The study was a retrospective study, and it was approved by the Institutional Review Boards of Sun Yat-Sen Memorial Hospital (SYSEC-KY-KS-2021-121) and the First Affiliated Hospital of Xinjiang Medical University (K202106-17) and is reported in accordance with the TRIPOD statement

Determining the Minimal Clinically Important Difference (MCID) and Responsiveness of the Chinese Version of Zurich Chronic Middle Ear Inventory (ZCMEI-21-Chn): A Prospective Multicenter Study.

This study aimed to establish the minimal clinically important difference (MCID) and assess the responsiveness of the Chinese version of Zurich Chronic Middle Ear Inventory (ZCMEI-21-Chn). Prospective multicenter study. Four Chinese tertiary referral centers admitting patients nationwide. 230 adult patients with chronic otitis media (COM) undergoing tympanoplasty. Patients were required to complete the ZCMEI-21-Chn to measure health-related quality of life both preoperatively and postoperatively. An anchor-based method was used to determine the MCID of the derivative cohort by including the Global Rating of Change Questionnaire as an anchor. The generalizability and consistency with functional outcomes of the MCID estimates were externally examined in a validation cohort using a receiver operating characteristic curve analysis. A total of 161 and 69 patients were included in the derivative and validation cohort. The mean preoperative and postoperative ZCMEI-21-Chn total scores were 28.4 (standard deviation [SD] 14.5) and 17.5 (SD 12.6). The mean change in ZCMEI-21-Chn score was 10.9 (SD 14.3, p < 0.001). The MCIDs of the ZCMEI-21-Chn for improvement and deterioration were estimated at 13 (SD 13.0) and -7 (SD 12.9), accordingly. For patients who have reported an improved health-related quality of life, a cutoff value of 15.6 dB HL for elevation of the air-conducted hearing threshold was noticed. However, change of clinical importance judged according to MCID and Japan Otological Society criteria disagreed with each other, notably with a Cohen's kappa (κ) of 0.14 (p = 0.21) in the validation cohort. This study is the first to establish the MCID of a COM-specific questionnaire in Chinese. For the COM population undergoing surgical intervention, MCID values of 13 for improvement and -7 for deterioration are recommended. The results were externally validated to be generalizable to nationwide usage, yet distinguishable from the audiological criteria. The availability of the MCID greatly adds to the clinical utility of the ZCMEI-21-Chn by enabling a clinically meaningful interpretation of its score changes.

Extrauterine growth restriction in very-low-birthweight infants: prevalence and concordance according to Fenton, Olsen, and INTERGROWTH-21st growth charts in a multicenter Spanish cohort.

Multiple criteria and growth references have been proposed for extrauterine growth restriction (EUGR). We hypothesized that these may impact the diagnosis of EUGR. The objective was to evaluate the prevalence of EUGR with its different definitions and the concordance according to Fenton, Olsen, and INTERGROWTH-21st in very-low-birthweight (VLBW) infants. This is an observational, retrospective, and multicenter study including VLBW infants from the Spanish SEN1500 Network from 2011 to 2020. Patients with major congenital anomalies, embryopathies, and gestational age less than 24weeks were excluded. EUGR prevalence was calculated at discharge with cross-sectional, longitudinal, "true" cross-sectional, and "true" longitudinal definitions. Concordance was assessed with Fleiss' kappa coefficient. 23582 VLBW infants from 77 NICUs were included. In total, 50.4% were men with a median of gestational age of 29 (4) weeks. The prevalence of EUGR (cross-sectional, longitudinal, and "true") was variable for weight, length, and head circumference. Overall, the prevalence was higher with Fenton and lower with Olsen (cross-sectional and "true" cross-sectional) and INTERGROWTH-21st (longitudinal and "true" longitudinal). Agreement among the charts by weight was good only for cross-sectional EUGR and moderate for longitudinal, "true" cross-sectional, and "true" longitudinal. Concordance was good or very good for EUGR by length and head circumference.Conclusions: The prevalence of EUGR with the most commonly used definitions was variable in the cohort. Agreement among growth charts was moderate for all the definitions of EUGR by weight except cross-sectional and good or very good for length and head circumference. The choice of reference chart can impact the establishment of the diagnosis of EUGR. What is known: • EUGR has been defined in the literature and daily practice considering weight, length and head circumferencewith multiple criteria (cross-sectional, longitudinal, and "true" definition) • Different growth charts have been used for EUGR diagnosis What is new: • Prevalence of EUGR is variable depending on the definition and growth chart used in our cohort of VLBW infants • For the most frequently EUGR criteria used, traditionally considering weight, concordance among Fenton, Olsen and INTERGROWTH-21st growth charts is only moderate for all the definitions of EUGR by weight except cross-sectional definition. Concordance among the charts is good or very good for the different criteria of EUGR by head circumference and length.

P-552 Unveiling associated factors to in vitro embryonic ploidy: Insights from 8194 blastocysts by Next Generation Sequencing

Abstract Study question What factors are associated with an increased probability of having aneuploid or mosaic embryos in vitro? Summary answer Maternal age and biopsy day significantly influence aneuploidy, while trophectoderm quality impacts both aneuploidy and embryo mosaicism. What is known already Embryonic aneuploidy and mosaicism are critical factors affecting the success of in vitro fertilization. Several studies have demonstrated that advanced maternal age is the most important variable associated with chromosomal abnormalities. However, the influence of other factors, including the day of the blastocyst biopsy and the quality of specific embryonic components, such as the trophectoderm, on the likelihood of aneuploidy and mosaicism is less understood. Study design, size, duration This retrospective study involved the analysis of 8,194 blastocyst embryos derived from 2,180 IVF/ICSI cycles at a specialized assisted reproduction clinic. The study spanned 7 years, from 2017 to 2023, focusing on identifying factors influencing embryonic ploidy Participants/materials, setting, methods Retrospective analysis of 8194 blastocysts from 2180 IVF/ICSI cycles at a single fertility center. Evaluated maternal age, paternal age, day of embryo biopsy, inner cell mass and trophectoderm quality, embryo biopsy operator, fertilization technique, using logistic regression to assess impact on ploidy (euploid, aneuploid, mosaic). Analysis identified significant factors influencing ploidy, adjusting for confounders. Statistical significance at p &amp;lt; 0.05. Main results and the role of chance The analysis of 8,194 blastocyst embryos highlighted significant findings. Maternal age showed a considerable impact on aneuploidy with a p-value &amp;lt;0.001 and an odds ratio (OR) of 1.74, indicating a strong association. Similarly, the day of embryo biopsy was a significant factor (p &amp;lt; 0.001, OR = 1.51) emphasizing its influence on embryo aneuploidy. The quality of the trophectoderm was a crucial factor, with a notable correlation with both aneuploidy (p &amp;lt; 0.001, OR = 1.78) and mosaicism (p &amp;lt; 0.001, OR = 1.28). Further analysis was also performed to find the variables associated with mosaicism, and the only significant variable was quality of trophectoderm increasing the probability of mosaicism by 28%. These statistical values underscore the robustness of the study’s conclusions. The large sample size and the application of logistic regression models further minimized the role of chance, enhancing the reliability of these findings. The research provides significant insights into the factors influencing embryonic ploidy, extending beyond the well-recognized impact of maternal age. It underscores the importance of considering these variables in clinical assessments during fertility treatments, particularly in evaluating the risk of chromosomal abnormalities. Limitations, reasons for caution The study’s retrospective nature limits causal inferences. The findings need validation in prospective, multicenter studies to establish broader applicability and account for potential biases. Wider implications of the findings These results contribute to the understanding of the factors associated with aneuploidy and embryonic mosaicism, beyond maternal age. Integrating the day of embryo biopsy and trophectoderm quality as key factors into clinical evaluation that could improve clinical decision-making in fertility treatments, especially in assessing the risk of chromosomal abnormalities. Trial registration number Non-Clinical trials

PSA doubling time 4.65 months as an optimal cut-off of Japanese nonmetastatic castration-resistant prostate cancer

A multicenter study of nonmetastatic castration-resistant prostate cancer (nmCRPC) was conducted to identify the optimal cut-off value of prostate-specific antigen (PSA) doubling time (PSADT) that correlated with the prognosis in Japanese nmCRPC. Of the 515 patients diagnosed and treated for nmCRPC at 25 participating Japanese Urological Oncology Group centers, 450 patients with complete clinical information were included. The prognostic values of clinical factors were evaluated with respect to prostate specific antigen progression-free (PFS), cancer-specific survival (CSS), and overall survival (OS). The optimal cutoff value of PSADT was identified using survival tree analysis by Python. The Median PSA and PSADT at diagnosis of nmCRPC were 3.3 ng/ml, and 5.2 months, respectively. Patients treated with novel hormonal therapy (NHT) showed significantly longer PFS (HR: hazard ratio 0.38, p < 0.0001) and PFS2 (HR 0.45, p < 0.0001) than those treated with vintage nonsteroidal antiandrogen agent (Vintage). The survival tree identified 4.65 months as the most prognostic PSADT cutoff point. Among the clinical and pathological factors PSADT of < 4.65 months remained an independent prognostic factor for OS (HR 2.96, p = 0.0003) and CSS (HR 3.66, p < 0.0001). Current data represented optimal cut-off of PSADT 4.65 months for a Japanese nmCRPC.

P-662 Effect of rescue-dose progesterone in patients with low serum progesterone levels on the day of embryo transfer in frozen embryo transfer cycles

Abstract Study question What is the effect of rescue-dose progesterone in patients with low serum progesterone on the day of embryo transfer in frozen embryo transfer cycles? Summary answer In patients with low serum progesterone on the day of embryo transfer, rescue dose progesterone results in significantly higher clinical pregnancy rates. What is known already Frozen embryo transfer (FET) has become more common lately with many centers utilizing hormone replacement therapy (HRT) to electively plan embryo transfer with improved compliance rates. Serum progesterone level on the day of embryo transfer is a very simple, useful investigation which can be used as a predictor of clinical outcomes. Patients with lower progesterone levels (&amp;lt;5ng/ml) are associated with lower clinical pregnancy and ongoing pregnancy rates. Progesterone levels of &amp;gt; 10ng/ml are considered optimal by various studies. There is limited data on the efficacy of progesterone supplementation in patients with low progesterone levels. Study design, size, duration This study was conducted to assess the effect of rescue dose progesterone in patients with low serum progesterone levels on the day of embryo transfer in patients with HRT-FET cycles. This was a retrospective study and data was collected from prospectively maintained records of patients undergoing treatment at Miracles IVF and fertility center, Gurgaon, India between December, 2021 to November, 2023. Participants/materials, setting, methods All were HRT-FET cycles with vaginal progesterone supplementation of 800 mg/day. Serum progesterone levels were done on day of embryo transfer and if 5-10 ng/ml, rescue dose progesterone (100 mg intramuscular) was given. Single blastocyst embryo transfer was done. Inclusion criteria: 1.Age 25-35 years, 2.Endometrial thickness &amp;gt;7.5mm on day of start of progesterone. Exclusion criteria: 1.Transfer day progesterone &amp;lt;5ng/ml, 2. BMI &amp;gt; 30, 3. Previously failed embryo transfer The primary outcome was clinical pregnancy rate. Main results and the role of chance A total of 121 patients underwent 165 cycles during the study period with 93 patients included in the study after fulfilling the inclusion and exclusion criteria. 35 patients (37.6%) received rescue dose progesterone (RD group) and remaining 58 patients (62.4%) received no rescue dose (NRD group) as per their serum progesterone levels. All these patients were followed as per protocol and serum bhcg was done after 14 days of embryo transfer. Patients with positive values were monitored till positive clinical pregnancy. Biochemical pregnancy rates were 20 out of 35 (57.1%), 35 out of 58 ( 60.3%) and 55 out of 93 ( 59.1%) in RD group, NRD group and overall respectively. Clinical pregnancy rates were 18 out of 35 (51.4%), 34 out of 58 ( 58.6%) and 52 out of 93 ( 55.9%) in RD group, NRD group and overall respectively. RD group had significantly improved outcomes in terms of clinical pregnancy rates which were comparable to NRD group Limitations, reasons for caution Limitations, reasons for caution: Ours was a single center retrospective study with limited sample size. Though, our study revealed comparable outcome between the two groups but large volume multicenter studies are needed for making conclusive recommendations. Wider implications of the findings This study provides an insight into the role of serum progesterone levels on the day of embryo transfer. Rescue progesterone dosage for patients with sub-optimal serum progesterone levels may help in achieving similar clinical pregnancy rates and improving overall outcomes. Trial registration number not applicable

A multicenter protocol to assess the prognostic significance of the tumor microenvironment in patients with squamous cell carcinoma of the larynx.

The purpose of this multicenter study was to retrospectively investigate the prognostic significance of the tumor microenvironment, in relation to survival in a large cohort of patients with laryngeal squamous cell carcinoma (LSCC), using the method proposed by the International TILs Working Group in breast cancer. All consecutive patients with biopsy-proven LSCC who underwent total laryngectomy (TL) between January 2014 and January 2023 were retrospectively included in the study. A retrospective review of medical records including surgical, pathological and follow-up reports was performed. The density of TILs was determined according to the recommendations of the International TILs Working Group. The study group included 186 patients with LSCC. High TILs were statistically correlated with reduced size and extension of primary tumor (pT stage) with a statistically significant value (S: p = 0.01; P: p = 0.0003) and without needs of salvage therapy (S: p = 0.03; P: p = 0.004). Low TILs were indicative of worse prognosis. Our study confirmed the protective value of TILs and the prognostic role of the tumor microenvironment in LSCC; furthermore, our results showed that the score proposed by the International TILs Working Group for breast cancer can be applied to LSCC.

Cervicothoracic ventral-dorsal rhizotomy for treatment of brachial hypertonia in cerebral palsy.

Cervicothoracic ventral-dorsal rhizotomy (VDR) is a potential treatment of medically refractory hypertonia in patients who are not candidates for intrathecal baclofen, particularly in cases of severe upper limb hypertonia with limited to no function. A longitudinal cohort was identified to highlight our institutional safety and efficacy using cervicothoracic VDR for the treatment of hypertonia. Retrospective data analysis was performed for patients that underwent non-selective cervicothoracic VDR between 2022 and 2023. Non-modifiable risk factors, clinical variables, and operative characteristics were collected. Six patients (three female) were included. Four patients underwent a bilateral C6-T1 VDR, one patient underwent a left C7-T1 VDR, and another underwent a left C6-T1 VDR. Three patients had quadriplegic mixed hypertonia, one patient had quadriplegic spasticity, one patient had triplegic mixed hypertonia, and one patient had mixed hemiplegic hypertonia. The mean difference of proximal upper extremity modified Ashworth scale (mAS) was - 1.4 ± 0.55 (p = 0.002), and - 2.2 ± 0.45 (p < 0.001) for the distal upper extremity. Both patients with independence noted quality of life improvements as well as increased ease with dressing and orthotics fits. Caregivers for the remaining four patients noted improvements in caregiving provision, mainly in dressing, orthotics fit, and ease when transferring. Cervicothoracic VDR is safe and provides tone control and quality of life improvements in short-term follow-up. It can be considered for the treatment of refractory hypertonia. Larger multicenter studies with longer follow-up are necessary to further determine safety along with long-term functional benefits in these patients.

Real Implication of Fertility-Sparing Surgery for Ovarian Cancer: Reproductive Outcomes

Background: to prove the effectivity of fertility-sparing procedures in early-stage ovarian cancer by assessing pregnancy rates and obstetrical outcomes. Methods: we performed a retrospective multicenter study among 55 Spanish hospitals, collecting patients from 18 to 40 years old with diagnosis of early-stage ovarian cancer, epithelial (EOC) or non-epithelial (non-EOC), from January 2010 to December 2019. Data on the use of assisted reproductive techniques, pregnancy attempts and obstetrical outcomes were collected. Results: a total of 150 patients met inclusion criteria, 70 (46.6%) EOC and 80 (53.4%) non-EOC. Pregnancy attempts were reported in 51 (34%) patients, with 42 (28%) patients carrying the pregnancy to term. Among them, 30 (71.4%) underwent surgery alone and 12 (28.6%) had additional postoperative chemotherapy. A total of 32 (76.1% patients) had spontaneous pregnancies and 10 (23.9%) required in vitro fertilization. There was only one (2.4%) complication reported. Vaginal delivery was reported in twenty-nine (69%) patients and cesarean section in five (11.9%) patients. Conclusions: fertility-sparing management for ovarian cancer seems to be an option with proven good pregnancy rates and low complications. The selection of patients must consider strict criteria in order to maintain a good prognosis.

Biomarkers In Prediction of Acute Mesenteric Ischaemia: a prospective multicentre study (BIPAMI study): a study protocol

BackgroundAcute mesenteric ischaemia (AMI) is a life-threatening disease where early diagnosis is critical to avoid morbidity and mortality from extensive irreversible bowel necrosis. Appropriate prediction of presence of bowel necrosis is currently not available but would help to choose the optimal method of treatment.The study aims to identify combinations of biomarkers that can reliably identify AMI and distinguish between potentially reversible and irreversible bowel ischaemia.MethodsThis is a prospective multicentre study. Adult patients with clinical suspicion of AMI (n = 250) will be included. Blood will be sampled on admission, at and after interventions, or during the first 48 h of suspicion of AMI if no intervention undertaken. Samples will be collected and the following serum or plasma biomarkers measured at Tartu University Hospital laboratory: intestinal fatty acid-binding protein (I-FABP), alpha-glutathione S-transferase (Alpha- GST), interleukin 6 (IL-6), procalcitonin (PCT), ischaemia-modified albumin (IMA), D-lactate, D-dimer, signal peptide-CUB-EGF domain-containing protein 1 (SCUBE-1) and lipopolysaccharide-binding protein (LBP). Additionally, more common laboratory markers will be measured in routine clinical practice at study sites.Diagnosis of AMI will be confirmed by computed tomography angiography, surgery, endoscopy or autopsy.Student’s t or Wilcoxon rank tests will be used for comparisons between transmural vs. suspected (but not confirmed) AMI (comparison A), confirmed AMI of any stage vs suspected AMI (comparison B) and non-transmural AMI vs transmural AMI (comparison C). Optimal cut-off values for each comparison will be identified based on the AUROC analysis and likelihood ratios calculated. Positive likelihood ratio > 10 (> 5) and negative likelihood ratio < 0.1 (< 0.2) indicate high (moderate) diagnostic accuracy, respectively. All biomarkers with at least moderate accuracy will be entered as binary covariates (using the best cutoffs) into the multivariable stepwise regression analysis to identify the best combination of biomarkers for all comparisons separately. The best models for each comparison will be used to construct a practical score to distinguish between no AMI, non-transmural AMI and transmural AMI.DiscussionAs a result of this study, we aim to propose a score including set of biomarkers that can be used for diagnosis and decision-making in patients with suspected AMI.Trial registrationNCT06212921 (Registration Date 19–01-2024).

P-714 Prevalence of chromosomal abnormalities in oocyte donor candidates: a French survey of over 8200 karyotypes

Abstract Study question Is karyotyping worth considering for fertile and childless oocyte donor candidates? Summary answer Karyotyping reduces the genetic risk of utilizing oocytes with balanced structural chromosomal rearrangements for recipient couples. What is known already While some countries including France consistently advocate for karyotype to be routinely performed, others suggest it as optional, specifically advising it for donors with a medical history indicating chromosomal rearrangements. Ravel et al. have demonstrated a higher incidence of balanced structural chromosomal rearrangements among 1196 french fertile oocyte donor candidates (FODC) compared to unselected newborns, raising the question whether oocyte donor recruitment could be biased towards chromosomally at-risk women. In 2016, France authorized childless oocyte donor candidates (CODC) to enhance recruitment, yet as of now, there is no reported study on their genetic status. Study design, size, duration A retrospective observational multicentric study conducted from January 2005 to October 2021. Participants/materials, setting, methods ODC were recruited within the French CECOS (Centre d’Etude et de Conservation des Œufs et du Sperme humain) network. Both FODC and CODC were compared to a control group composed of female newborns from literature. Quantitative and qualitative data were expressed as the median [25%–75%] or n (‰), respectively. Non-parametric Mann-Whitney tests were employed for quantitative parameters, while Chi-square tests or Fisher’s exact tests were used, as appropriate, to compare frequencies between groups. Main results and the role of chance A total of 8229 ODC from 22 CECOS centers were included in this study. FODC (n = 6339) and CODC (n = 1890) were compared to 8102 control female newborns. In total, 65 chromosomal abnormalities (7.9‰) were identified, leading to the exclusion of the ODC. This outcome was comparable to the 53 abnormalities observed in the control group (6.54‰, p = 0.3059). The occurrence of balanced structural chromosomal rearrangements was increased in ODC population (4.8‰) compared to controls (2.34‰, p = 0.0086). The occurance of reciprocal translocations was increased 5-fold in CODC (3.70‰) compared to the control group (0.74‰, p = 0.013). The number of gonosomal mosaics was notably increased in FOCD, with 17 cases (2.68‰) compared to the control group with six cases (0.74‰, p = 0.0052). Among chromosomal aberration carriers only two childless women (one reciprocal translocation and one gonosomal mosaïc) had fertility issues in their medical history with a diagnosis of premature ovarian failure. Limitations, reasons for caution This retrospective study was designed to examine ODC within the framework of French bioethics laws. ODC should adhered to specific criteria: being volunteers, anonymous, aged 18 to 38, and without any financial compensation. Since 2016, donors were permitted to retain around 50% of their gametes for personal oocyte cryopreservation. Wider implications of the findings Karyotyping is a straightforward and cost-effective approach to identify structurally balanced chromosomal abnormalities. Thus, chromosomal analysis limits genetic risks whithout resorting to the ethically debated expanded genetic screening in the ODC population. Nevertheless, it is crucial to communicate to gamete recipients the inherent genetic risks associated with any conception. Trial registration number not applicable

Does Antrum Size Matter in Sleeve Gastrectomy? Volume II—A Retrospective Multicentric Study with Long-Term Follow-Up

Background: Laparoscopic sleeve gastrectomy (LSG) is the most widespread bariatric procedure due to its safety and efficacy. Despite continuous refinement, achieving a globally standardized procedure remains challenging. Moreover, due to its wide adoption, numerous studies have focused on complications associated with the technique, such as gastroesophageal reflux disease (GERD). This study evaluates the impact of antrum size (wide antrectomy versus small antrectomy) in LSG on long-term anthropometric outcomes and complications in patients with morbid obesity. Methods: Body mass index (BMI), percentage of excess weight loss (%EWL) at a 5-year follow-up, GERD Health-Related Quality-of-Life (GERD-HRQL) scores, and obesity-related diseases of patients undergoing LSG with gastric resections starting 2 cm and 6 cm from the pylorus were retrospectively evaluated. Results: Between January 2015 and November 2019, 597 patients who met the criteria for LSG were included in the study. Group A (241 patients) underwent wide antrectomy, while Group B (356 patients) underwent small antrectomy. Weight, BMI, %EWL, and %TWL significantly improved at 6 and 12 months in the wide-antrectomy group. However, these differences diminished by 24 months, with no significant long-term differences in weight loss outcomes between the two groups at 5 years. Conversely, GERD-HRQL scores were significantly better in the small-antrectomy group until 24 months; thereafter, results were comparable between groups over the long term. Conclusions: Therefore, while wide antrectomy may offer superior short-term anthropometric outcomes, both techniques yield similar long-term results regarding weight management and GERD incidence. Larger prospective studies are needed to further address this issue.

O-006 Oocyte factors as predictors of abnormal, mononucleated and trinucleated, zygotes and assessment of their morphokinetic differences

Abstract Study question Are there oocyte factors associated with the occurrence of mononucleated and trinucleated zygotes after ICSI? Does morphokinetics differ in embryos from normal and abnormal zygotes? Summary answer Formation of 1PN zygotes relates to vitrificated and donor oocytes. 3PN links to age and own oocytes. 1PN embryo morphokinetics are slower than 2PN, 3PN. What is known already In assisted reproduction, after monitoring fertilization, it is common to observe zygotes with one or more than 2 pronuclei. These abnormal zygotes are often discarded due to potential chromosomal abnormalities. The formation of these anomalous zygotes can be attributed to various causes, such as non-activation of the male pronucleus, non-extrusion of the second polar body, etc. Simultaneously, limited data exist on the morphokinetics of abnormal zygotes, explored in a few studies. The embryonic kinetics of 1PN zygotes seem slower than those of 2PN and 3PN zygotes, but further research is needed to confirm this aspect. Study design, size, duration Retrospective multicentre study involved time-lapse monitoring of 28,218 ICSI zygotes (26,252 2PN, 1,190 1PN, and 776 &amp;gt;3PN) generated between January 2019 and December 2023. The study includes donor and patient oocytes with various infertility diagnoses, whether fresh or vitrified. The comparison of embryonic morphokinetics from abnormal/normal zygotes was conducted on 254 embryos from a single centre, with 45 originating from 1PN zygotes, 198 from 2PN zygotes, and 11 from zygotes with 3 or more pronuclei. Participants/materials, setting, methods The study involved 1,053 couples treated in four different centres. Uniform culture media were used, and oocytes were cultivated in time-lapse technology-equipped incubators across all centres. A logistic regression analysis aimed to identify oocyte factors linked to an abnormal pronuclei count. Embryonic morphokinetics were analysed using the CHLOE artificial intelligence system (Fairtility, Israel). Evaluated parameters included: tPNa, tPNf, t2, t3, t4, t5, t6, t7, t8, t9+, tM, tSB, tB, and tEB. Main results and the role of chance Donor oocytes exhibited a 1.32 times higher risk of producing 1PN zygotes than 2PN or 3PN zygotes (p &amp;lt; 0.001). Similarly, vitrified oocytes had a 1.63-fold higher risk of generating 1PN zygotes than 2PN or 3PN zygotes (p &amp;lt; 0.001). Additionally, we found that both the use of own oocytes and advanced age were significantly associated with 3PN zygotes (p &amp;lt; 0.001). Regarding age, the risk increases by 10% for each additional year. The average durations for all morphokinetic parameters of 2PN and 3PN embryos were similar. In contrast, all events, except t3 (p = 0.2), occurred more slowly in embryos derived from 1PN zygotes compared to embryos from 2PN zygotes. Thus, the disappearance of pronuclei had a p-value &amp;lt;0.005, and from t4 to tEB, the p-value was &amp;lt;0.001. When comparing embryos from 1PN and 3PN zygotes, only tPNa, t4, t8, t9+, and tM were significantly different. If only zygotes that reached the blastocyst stage were used for analysis, the results were less conclusive due to a significantly reduced number of cases. Limitations, reasons for caution Embryos were cultured in an EmbryoScope incubator at three centres, while Geri was used in the fourth. Wider implications of the findings Using donor or vitrified oocytes may lead to an increased occurrence of zygotes with 1PN, as will occur with 3PN when patient or aged oocytes are used. The kinetics of embryos from 3PN oocytes resemble those from 2PN, while those from 1PN zygotes exhibit slower kinetics. Trial registration number not applicable

P-134 Fatty acid supplementation during warming improves pregnancy outcomes after frozen blastocyst transfers: a propensity score-matched study

Abstract Study question Does warming with fatty acids (FAs) affect the viability of human blastocysts and pregnancy outcomes after a single vitrified-warmed blastocyst transfer (SVBT)? Summary answer FA-supplemented warming solutions promoted the blastocyst outgrowth by decreasing the degeneration of trophoblast cells and improved pregnancy outcomes after SVBTs. What is known already Vitrification is considered a well-established technique and has been used worldwide. However, previous studies demonstrated the detrimental effects of vitrification on pre-implantation development and term pregnancy in oocytes and embryos. Vitrification reduces the intracellular lipid content and decreases the competence for pre-implantation development. The FA supplementation into the warming solution restores the lipid content in the ooplasm and improves developmental competence by promoting the β-oxidation pathway in mice. Although, in humans, the FA-supplemented warming solutions reportedly improved pregnancy outcomes after vitrified-warmed cleavage-stage embryo transfers, the effectiveness of these solutions on blastocyst warming remain unclear. Study design, size, duration The experimental study included 401 discarded blastocysts. The blastocysts were warmed using the FA-supplemented (FA group) or non-supplemented (control group) solutions. The blastocyst adhesion, outgrowth area, and incidence of outgrowth degeneration were assessed. The retrospective cohort study included 4,426 treatment cycles from 4,426 women who underwent SVBTs between January 2023 and August 2023 (January–May 2023, control group; June–August 2023, FA group). Pregnancy outcomes were compared statistically between the groups. Participants/materials, setting, methods The implantation competence was experimentally examined using an outgrowth model. The blastocysts were placed on fibronectin-coated dishes and cultured for 96 hours. The embryos were designated as degenerated when the outgrowth area decreased by &amp;gt; 10%, compared with the maximum area. For the clinical study, multiple covariates that affected the outcomes were used for propensity score matching. The patients in the FA group were matched with controls using a propensity score in a 1:1 ratio. Main results and the role of chance The adhesion rate after the outgrowth culture was comparable between the control and FA groups. However, the outgrowth was larger in the FA group than in the control group. The multivariate linear regression analysis also demonstrated similar results. Furthermore, the incidence of outgrowth degeneration was significantly decreased in the FA group. Furthermore, the subgroup analyses revealed that FA-supplemented warming solutions were effective in blastocysts donated from young patients, day 5 blastocysts, and morphologically good blastocysts. For the retrospective cohort study, the implantation, clinical pregnancy, and ongoing pregnancy rates were significantly higher in the FA group than in the control group. The multivariate logistic regression analysis also demonstrated that warming with FA-supplemented solutions significantly increased the probability of ongoing pregnancies. However, the rates of early pregnancy loss and miscarriage in the first trimester were comparable between the groups. The subgroup analyses demonstrated a similar trend to in vitro experiments; the FA-supplemented solutions were more effective in good prognosis cases (young patients, day 4/5 blastocysts, and morphologically good blastocysts) than in poor prognosis cases (advanced age patients, day 6/7 blastocysts, and morphologically poor blastocysts). Limitations, reasons for caution This was a single-centre, retrospective study, necessitating further multicentre studies to ascertain the generalizability of these findings to other clinics with different protocols and/or patient demographics. Despite using propensity score matching and multivariate models to adjust for the confounders, we cannot adjust for all possible confounders. Wider implications of the findings FA-supplemented warming solutions improved pregnancy outcomes after SVBTs, particularly in patients with a good prognosis. These warming solutions could contribute to maximizing clinical outcomes, shortening treatment periods, and reducing patient burden. Trial registration number not applicable

O-278 The vaginal and endometrial microbiome: do they tell the same story?

Abstract Study question Is the composition of the endometrial microbiome comparable to that of the vagina and can vaginal sampling provide sufficient information to characterize the uterine microbiome? Summary answer The compositions of the endometrial and vaginal microbiome are largely comparable and therefore, in most cases, vaginal sampling is sufficient to characterize the uterine microbiome. What is known already A dysbiotic endometrial microbiome (i.e., a shift from a predominance of beneficial bacteria to less favourable species) has been reported in association with adverse pregnancy outcomes in assisted reproductive treatment (ART). If dysbiosis is detected, antibiotics or probiotics can be administered to restore an optimal microbiome. However, characterization of the endometrial microbiome is challenging, involving invasive sampling, and technical difficulties due to the low microbial mass in the endometrium. In contrast, vaginal sampling is considered minimally invasive and microbial analysis is technically less complex. It is important to clarify whether the microbiomes of the endometrium and the vagina are comparable. Study design, size, duration A prospective multi-centre study including 60 ART patients was conducted from June 2022 to August 2023. During the luteal phase (on average day-19), paired vaginal swabs and endometrial biopsies were collected sequentially and stored in a preservation medium for further analysis. Patients and physicians were also asked to complete a questionnaire to identify any lifestyle or medical factors that may have differentially affected the microbial composition of the vagina and endometrium. Participants/materials, setting, methods A carefully validated qPCR methodology was used to accurately assess the presence and relative abundance of the four major lactobacillus species associated with eubiosis (crispatus, gasseri, iners, and jensenii) and the 15 most common bacterial species implicated in dysbiosis. Additionally, the relative abundance of microbial and human DNA was examined. Of note, endometrial analysis required additional tissue lysis. Furthermore, due to the low microbial DNA content in endometrial samples, preamplification was necessary prior to qPCR. Main results and the role of chance The proportion of microbial versus human DNA was approximately 90 times higher in vaginal samples compared to endometrial samples (p &amp;lt; 0.0001). However, the microbial compositions of both the endometrium and the vagina showed remarkable similarities. Lactobacillus species were predominant at both sites, accounting for an average of 86% and 89% of the bacterial population in the vagina and endometrium, respectively (p = 0.6). Of the dysbiotic species, Gardnerella vaginalis was the most common, accounting for 8% (vagina) and 7% (endometrium) of the total bacterial population (p = 1). There was no difference in the average number of distinct microbial species between the two niches (1.8 vs. 2.1; p = 0.3). Direct comparison of paired samples highlighted the high similarity of microbial compositions between both sites and showed strong correlations of species abundances in individual patients (L.crispatus: r = 0.9, L.iners: r = 0.9, L.gasseri: r = 1, L.jensenii: r = 0.9, G.vaginalis: r = 0.7; p &amp;lt; 0.0001/all species). However, the microbial composition differed in four patients (6.6%), with three having a dysbiotic vaginal profile (≥50% dysbiotic species) but a eubiotic endometrial profile (≥50% lactobacillus species) and one showing the reverse pattern. The medical conditions of two patients, including chronic endometritis and use of antibiotics and probiotics in the preceding cycle, may account for the observed differences. Limitations, reasons for caution The species analysed were carefully selected to ensure a comprehensive analysis. Nevertheless, the potential presence of unevaluated species could impact the results. While the included patients displayed diversity in their reproductive histories, it is important to acknowledge that the observed results may still be influenced by underlying causes of infertility. Wider implications of the findings The close similarity between the microbiomes at the two sites investigated suggests that minimally invasive vaginal sampling may be sufficient to characterise the endometrial microbiome. The ease of vaginal sampling and processing is expected to improve patient recruitment in microbiome research and facilitate easier application of clinical microbiome tests. Trial registration number not applicable

P-514 Unequal pronucleus size is indicative of higher rates of diploidy in 3PN blastocysts

Abstract Study question What proportion of blastocysts developed from 3PN embryos are diploid, and is this associated with the relative size of the pronuclei? Summary answer Most 3PN blastocysts with even-sized pronuclei are triploid, while most 3PN blastocysts with uneven-sized pronuclei are diploid. What is known already Fertilised oocytes with three pronuclei (3PN, or tripronuclear) are generally considered unsuitable for use due to the high likelihood of triploidy, which is not compatible with an ongoing healthy pregnancy. Recent data on 3PNs with uneven-sized pronuclei (micro-3PN, or 2.1PN) indicates that many may actually be diploid if they form a good quality blastocyst. Study design, size, duration Prospective multi-centre study from October 2020 to December 2023. 81 3PN blastocysts from 69 patients were biopsied. Participants/materials, setting, methods ICSI-inseminated embryos were assessed on Day 1 and pronuclear size at 18 h post insemination was measured using EmbryoViewer software. Biopsy was performed at the blastocyst stage and genetic testing was performed using the Human Karyomap-12 SNP array to ascertain ploidy status. Pronuclei were classified as “Equal”, “Unequal” (smallest pronucleus is 61%-75% of the largest), or “Micro” (smallest pronucleus is &amp;lt; 60% of the largest). Main results and the role of chance 94% of the 3PN blastocysts with equal size pronuclei were triploid, and the remaining 6% were complex aneuploid or had genome wide uniparental disomy (n = 36). Of the 3PN blastocysts with unequal pronuclei (n = 19), 42% were triploid, whereas none of the embryos with a micro pronucleus were triploid (n = 26). Limitations, reasons for caution Only 3PN embryos that formed a good quality expanded blastocyst were biopsied. Only ICSI-inseminated embryos were included, not IVF. No 3PN embryos were transferred in the study, so any additional pregnancy and birth risks from diploid 3PN embryos are unknown. Wider implications of the findings Blastocysts that develop from 3PN oocytes with uneven pronuclei have a greater than 80% chance of diploidy and could be considered for clinical use with PGT. More data is needed to inform clinical guidelines. Trial registration number N/A

370 Do Prolonged Surgery Waiting List Times Affect the Incidence of Post-Tonsillectomy Haemorrhage, in the Context of the COVID-19 Pandemic?

Abstract Aim The United Kingdom was affected by the COVID-19 pandemic with the reduction of many services within the NHS, including suspension of elective surgical procedures from March 2020. We hypothesised that delayed surgery caused by the pandemic may increase the risk of post-tonsillectomy haemorrhage due to increased episodes of tonsillitis. The aims of this review were to investigate the relationship between prolonged waiting times and post-tonsillectomy haemorrhage. Method We performed a single-site, retrospective review from March 2019 to August 2023, with data collected from electronic hospital records. Both adults and children were included in the analysis and only procedures performed for recurrent tonsillitis were included. Results We identified 234 cases: 184 adeno/tonsillectomies in the pre-COVID (March 2019-20) and 50 in the post-COVID (August 2020-21) cohort. In the pre-COVID cohort, 7.6% (n=14) experienced a post-tonsillectomy haemorrhage, with 1.1% (n=2) experiencing a primary haemorrhage and 6.5% (n=12) experiencing secondary haemorrhage. The return to theatre rate was 2.2% (n=4). Median waiting time for surgery was 78 days. In the post-COVID cohort, secondary haemorrhage occurred in 6.0% (n=3). No patients experienced a primary haemorrhage and return to theatre rate was 0%. Median waiting time was 345 days. Simple logistic regression was performed which showed no significant association between waiting times and post-tonsillectomy haemorrhage across the whole (p = 0.90, 95% CI [-0.0002, 0.0002]) cohort. Conclusions Our cohort demonstrated no significant relationship between prolonged waiting times and post-tonsillectomy haemorrhage. Small sample size was a significant limitation; we recommend a larger scale, multi-centre study to investigate this further.

P-354 Advanced Paternal Age (APA) amplifies cryopreservation-induced stress in human spermatozoa

Abstract Study question Does paternal age intensify the negative effects of cryopreservation on sperm quality? Summary answer Paternal age exacerbates the adverse impact of cryopreservation on sperm quality. What is known already Despite mounting evidence indicating age-related declines in sperm quality, the impact of APA on male fertility remains unclear. APA raises questions about DNA fragmentation and mitochondrial activity, a robust indicator of sperm functionality. While sperm cryopreservation is extensively employed in fertility preservation and assisted reproductive technology (ART), its potentially detrimental effects on cells compartments, including mitochondria, persist. The mechanisms of APA on sperm susceptibility to cryopreservation-induced cellular damage still need to be addressed. Study design, size, duration This prospective multicentric study, conducted between January and October 2023, comprised 200 men. Only normozoospermic samples were included in the research (n = 81). Then, based on their age, the participants were categorised into two groups: ages ≤35 (non-APA, n = 40) and ages ≥42 (APA, n = 41). The cohorts were stratified into distinct subgroups, encompassing both fresh and frozen/thawed specimens of APA and non-APA sperm, and subjected to evaluations encompassing mitochondrial functionality, acrosomal reaction, DNA fragmentation and Tyrosine phosphorylation. Participants/materials, setting, methods Semen samples were collected after 2-7 days abstinence and analysed according to WHO 2021 guidelines. Acrosomal status was determined using FITC-conjugate PSA. DNA fragmentation was examined with the Halosperm kit. Mitochondrial functionality was gauged using MitoTracker Red CMXRos through the percentage of stained cells and fluorescence intensity and PCR for mitochondrial DNA copy number (mtDNAcn). Tyrosine phosphorylation was assessed through immunofluorescence and Western Blotting. Main results and the role of chance Non-APA group exhibited significantly superior fresh semen characteristics. DNA fragmentation was notably elevated in fresh semen from APA group, reaching 21.7%, compared to 15.4% non-APA individuals. After cryopreservation, the DNA fragmentation index further increased in men over 42 years to 26.7%. Mitochondrial functionality analysis indicated a significant decrease in staining intensity in cryopreserved sperm, notably in the APA group, with mean fluorescence intensity decreasing from 3.35±1.5 to 1.8±0.5. In contrast, the non-APA group experienced a drop from 3.33±1.6 to 2.53±1.6. Additionally, in the APA group, a significant increase in mtDNAcn was observed in both fresh and frozen/thawed sperm compared to the younger counterparts, indicative of poor mitochondrial quality. Cryopreservation negatively impacted acrosome integrity in both age groups, with a significant decrease observed in unreacted acrosomes from 71.5% to 57.7% in the non-APA group and from 75% to 63% in the APA group. Furthermore, exploration of capacitation-like changes, like tyrosine phosphorylation, demonstrated a significant decrease with age and a negative impact of cryopreservation on tyrosine phosphorylation in sperm from young males. Limitations, reasons for caution Main limitations are the small number of examined samples and the inclusion of only normozoospermic patients. Therefore, future investigations are needed to provide a more comprehensive understanding of the impact of paternal age on cryopreservation-induced stress in human spermatozoa. Wider implications of the findings Our research sheds light on the influence of paternal age on sperm cryopreservation, revealing higher vulnerability of mitochondria and elevated DNA fragmentation in APA. These findings should be considered when advising older men on choosing cryopreservation techniques in assisted reproductive treatments. Trial registration number Not applicable

P-138 Effect of biofeedback from the blastocysts on the cumulative pregnancy rate

Abstract Study question The study is to compare cumulative pregnancy rates of embryos transferred with and without spent culture media to confront biofeedback’s role in clinical outcomes Summary answer Transferring the blastocysts with spent culture media improved the pregnancy rates. Biofeedback was observed from the blastocyst stage. The blastocyst freezing and transfer approach improved. What is known already Biofeedback is often encountered in regulating menstrual cycles and other events in continuing gestations with estrogen, progesterone, beta hCG..etc. A similar biofeedback is possible with growing blastocyst for implantation. Since the in-vitro culture of embryos is devoid of its occurrence, it may lead to asynchrony of subsequent events and implantation. There is no scientific evidence from metabolomics on the characterization of these substances or secretions from spent culture medium that includes cell-free DNA giving feedback and its role in improving clinical outcomes. Study design, size, duration This retrospective multi-centric study included the 3591 frozen embryo transfers from 1st Apr 2022 to 31st Nov 2023 out of 4623 women who underwent oocyte retrieval from 1st Jan 2022. Patients were divided into study (group A) and control (group B). In group A, 1035 women received the transfers from spent culture media; in group B, 2556 women received transfers after shifting the blastocysts to fresh culture media before loading into the ET catheter. Participants/materials, setting, methods All the transfers considered in this study were day-5 blastocysts, irrespective of their frozen state. Day-5 frozen-thawed were incubated short-time (∼1 hour) before the transfer. Whereas, the frozen embryos and morulas were cultured till day 5. In the study group, all cultures were conducted in 100ul of culture media under oil per embryo to maintain consistency amongst the group. The rest of the laboratory and clinical procedures were consistent between the two groups. Main results and the role of chance The cumulative pregnancy rate was significantly higher in group A than in group B (60.9% Vs 55.6%; OR = 1.23; 95% CI, 1.07 -1.44, Z-score 2.901, P &amp;lt; 0.003). The effect of spent culture media was further observed by regrouping the day-3 or 4 frozen (group A1) and day-5 frozen patients (group A2). Day-5 thawed blastocysts transferred after short incubation to allow the secretions to directly deposit on the endometrium. Group A1 might have a high concentration of secretions due to its accumulation from the embryo or morula stages. Pregnancy rates in group A1 were higher than A2 but not statistically significant (61.7% Vs 59.7%; OR = 1.08, 95% CI, 0.84-1.4, Z-score 0.654, P = 0.51). The less impact indicates that the feedback came from blastocysts rather than embryos. Limitations, reasons for caution It is not evident how these secretions contribute to biofeedback. Quantification, identifying their time of release and recovery from the qualitative tests for their therapeutic use are the limitations. Nevertheless, spent media can be employed if found beneficial. Wider implications of the findings We have taken a large possible data set for control against the test group and took cumulative pregnancy rate as the primary outcome indicator to prove biofeedback from blastocysts can improve the clinical outcomes irrespective of the age and other factors of infertility. Trial registration number Not applicable