Single Euploid Blastocyst Transfer Research Articles

YOUNG EGGS IN AN OLD BASKET – IS THERE A MATERNAL AGE LIMIT FOR EMBRYO TRANSFER?

The current paradigm for in vitro fertilization (IVF) with embryo transfer (ET) suggests that the age of the uterus does not impact pregnancy outcomes. However few studies to date have focused on patients of very advanced maternal age. The objective of this study was to compare ET outcomes of women of advanced maternal age utilizing donor oocytes to determine whether age plays a role in reproductive outcomes. This was a retrospective cohort study conducted at a large University-affiliated fertility center. All ET cycles from 2012 to 2020 with single euploid blastocyst transfer that utilized donated oocytes were included. Cycles were divided into four groups by maternal age: less than 40 years, 40-44 years, 45-49 years, and 50 years or older. Primary outcomes were rates of positive serum hCG, clinical pregnancy, ongoing pregnancy (defined as presence of fetal heartbeat at discharge), miscarriage, and live birth. Continuous variables were analyzed with one-way analysis of variance (ANOVA) or Kruskal-Wallis test, while categorical variables with Pearson’s Chi-squares or Fischer-exact test. A total of 1,080 cycles were included (Table 1). Average patient age was 42.4 ± 4.9 years with a mean BMI of 26.5 ± 6.1 kg/m2. Average donor age was 27.4 ± 3.8 years with a mean antimullerian hormone (AMH) of 4.7 ± 2.0 ng/ml. Patient BMI, donor age, and donor AMH were similar between groups. There was no difference in the rates of positive hCG, clinical pregnancy, ongoing pregnancy or live birth between groups. There was a higher rate of first trimester miscarriage in patients over 50 (p < 0.01).Table 1Pregnancy outcomes of patients undergoing single, euploid ET using donor oocytesPatient age in yearTotal (n = 1080)<40 (n = 289)40-44 (n = 441)45-49 (n = 309)50+ (n = 41)p valuePositive serum hCG, n (%)833 (77.1%)225 (77.9%)341 (77.3%)235 (76.1%)32 (78.1%)0.95ClinicalPregnancy, n (%)716 (66.3%)187 (64.7%)295 (66. 9%)206 (66.7%)28 (68.3%)0.70Ongoing pregnancy, n (%)636 (58.9%)167 (57.8%)270 (61.2%)179 (57.9%)20 (48.8%)0.39First trimester miscarriage, n (%)95 (8.8%)22 (7.6%)29 (6.6%)36 (11.7%)8 (19.5%)<0.01*Live Birth, n (%)608 (56.3%)156 (54.0%)264 (59.9%)169 (54.7%)19 (46.3%)0.18 Open table in a new tab Success rates of IVF remained excellent in older patients who utilized donated oocytes. However, these patients, particularly those aged 50 years and older, may have an increased risk of first trimester miscarriage.

RECURRENT IMPLANTATION FAILURE – ROLE OF PGT AND ERA TO OPTIMIZE REPRODUCTIVE OUTCOMES?

To assess the role of Pre-Implantation Genetic Testing for Aneuploidy (PGT-A) and Endometrial Receptivity Assay (ERA) in couples with Recurrent Implantation failure (RIF) to optimize reproductive outcomes. This is retrospective data of couples at our private fertility clinic during January 2014 to July 2019. Study population was divided into three groups; All women with history of RIF were only recruited or this study. Group A – Both PGT-A and ERA done (n=79) Group B – only PGT-A done, no ERA done – (n=54) Group C – No PGT-A / No ERA done (n=189) INCLUSION CRITERIA Women with at least two fresh/frozen embryos transfers with minimum 4 blastocysts transferred in total and never conceived were considered as RIF. Only women with one euploid embryo, who underwent frozen embryo transfers (FET) were recruited in this study in group A&B. Women of all age groups who had Blastocysts available for transfer were included in Group- C. Only self-gamete cycles were considered in this study. All the women with RIF underwent controlled ovarian stimulation and oocyte retrieval as per our clinic’s standard operating protocol (SOP). ICSI was the choice of insemination considering history of failed implantation, fertilized oocytes were cultured till blastocysts and freeze all policy was adopted. Blastocysts were biopsied and trophectoderm tissue was subjected to genetic testing through Next-Generation Sequencing (NGS). ERA was done as per our institutes SOP. The biopsy was done using a endometrial biopsy pipette supplied by the company at 120 hours (day 5) of the start of the progesterone supplementation. This procedure was repeated at 144 hours (day 6) also. This was done to cater for the reduction in the error that could occur due to the extrapolation of results in cases of pre-receptive samples. The samples that were collected were stored and transported in the prescribed method of the company. The same conditions were replicated in the (FET) transfer cycle. Group A women underwent elective Single Euploid Blastocyst transfer (eSET) in an artificial cycle as per the ERA timing. Group B underwent eSET of a Euploid Blastocyst in a FET cycle on 6th day post progesterone support, no ERA was done. Group C women underwent transfer with un-screened double Blastocyst Transfer (DET) in a FET cycle. Primary Outcome was Implantation Rate (IR). Mean of IR of all subjects were calculated: Group A - 53% Group B - 47% Group C - 42% Odds ratio and p value was calculated for IR between the groups and showed no statistical significance. Though Group A had the best implantation rates, there was no statistical significance with the other two groups. Role of PGT-A and ERA as an intervention to improve reproductive outcomes is still debatable and needs well designed RCTs to further infer. PGT-A and ERA for RIF couples do not seem to offer beneficial reproductive outcomes. Couples need to be counselled appropriately while offering treatment options for RIF.

LIVE BIRTH RATES ARE SIGNIFICANTLY DECREASED FOLLOWING TRANSFER OF A EUPLOID BLASTOCYST WITH DIRECT UNEQUAL CLEAVAGE

Embryos can have direct unequal cleavage (DUC) during the first and/or second divisions. Although embryos with DUC have decreased blastulation rates, some develop into high quality euploid blastocysts. The objective of this study is to compare live birth (LB) rates following transfer of single frozen euploid blastocysts that either did or did not display DUC. This was a retrospective chart review of patients who underwent single frozen euploid blastocyst transfers from November 2018 to March 2020. Uninterrupted embryo culture was done in the Embryoscope up to 6 days post retrieval. Embryos were evaluated on day 2 for presence/absence of DUC and graded on day 5 or 6 according to the Gardner system. Blastocyst selection for frozen embryo transfer was based on euploidy status, developmental day, morphological score, and sex preference. Information regarding presence/absence of DUC was not accessed during blastocyst selection. Patient characteristics and LB rates were compared between the groups with and without DUC. Chi-square and Student’s t-tests were used for data analysis. A total of 114 single euploid blastocyst transfers were included, resulting in an overall LB rate per transfer of 63.2%. Twenty-two of the transferred blastocysts displayed DUC, whereas 92 did not. The overall LB rate in the DUC group was significantly lower than in the non-DUC group (31.8% vs 70.7%, p<0.01). Subgrouping blastocysts by developmental day and morphology showed significantly lower LB rates for day 6 and fair quality DUC blastocysts, compared with non-DUC blastocysts (18.2% vs 63.6%, p<0.05) (35.7% vs 70.6%, p<0.05). Live birth rates following single frozen euploid blastocyst transfer are significantly lower when blastocysts display DUC compared with blastocysts without DUC. When available, embryo division patterns should be considered when selecting a euploid blastocyst for frozen embryo transfer.

P–304 The endometrial preparation protocol does not affect the live-birth-rate after vitrified-warmed euploid single blastocyst transfers: an analysis of 1884 procedures

Abstract Study question Is the live-birth-rate (LBR) different when comparing artificial (AC) and modified-natural (M-NC) cycle for endometrial preparation to vitrified-warmed euploid blastocyst transfer? Summary answer The LBR after vitrified-warmed euploid blastocyst transfer seem independent of the endometrial preparation administered. What is known already Only the transfer of a competent embryo on a receptive endometrium might result in successful implantation. Three main protocols for endometrial preparation to vitrified-warmed embryo transfer exist: NC, M-NC, and AC. None among them, though, has been shown more appropriate than the others to date, especially since, only in a few studies, the analysis was restricted to single euploid blastocyst transfers to limit the impact of embryonic issues on implantation. In conclusion, no clear consensus exists and the choice is still largely based on menstrual/ovarian cycle characteristics and patient’s needs. Study design, size, duration All first vitrified-warmed single euploid blastocyst transfers performed between April–2013 and March–2020 were included in the analysis. Endometrial preparation was conducted with either an AC (N = 1211) or a M-NC (N = 673). The protocol was chosen based on patients’ logistical reasons. The primary outcome was the LBR per transfer. Sub-analyses based on blastocyst quality and day of development were conducted. Birthweight, gestational age, gestational and perinatal issues were secondary outcomes. Participants/materials, setting, methods AC: oral estradiol-valerate 3-times/day from day2–3 of the cycle until the endometrial thickness reached ≥7mm, then 600 mg/day of micronized progesterone. The transfer was conducted on day6 of progesterone administration. M-NC: an intramuscular dose of 10,000IU hCG was administrated when the leading follicle was &amp;gt;17 mm and the endometrium was thicker than 7mm and trilaminar, plus 400 mg/day of micronized-progesterone as luteal phase support starting 36–40hr post-hCG. The transfer was conducted on day7 after trigger. Main results and the role of chance The two groups were similar for maternal age at retrieval (38.0±3.3yr) and transfer (38.3±3.3yr), reproductive history, embryological outcomes of the IVF cycle, body-mass-index, basal hormonal levels, and blastocyst features (Gardner’s classification: AA = 73%, AB/BA=11%, BB/AC/CA=8%, CC/BC/CB=8%; day5=48%, day6=47%, day7=5%). The LBR was 46.7% (N = 565/1211) and 49.9% (N = 336/673) after AC and M-NC, respectively, resulting in an odds-ratio 1.14, 95%CI:0.94–1.37. The absence of significant differences was confirmed also when adjusted for blastocyst quality and day of full-development (1.16, 95%CI:0.96–1.41). Among the 565 and 336 deliveries, the birthweight was similar (3290.3±470.7 versus 3251.7±521.5 g, Mann-Whitney-U-test=0.5), the gestational age was similar (38.5±1.7 versus 38.4±1.9 weeks, Mann-Whitney-U-test=0.5). Also, the rates of newborns who were normal (81% versus 82%), large (8% versus 9%), and small (11% versus 9%) for gestational age were similar (Chi-squared-test=0.5). The rates of patients experiencing gestational (6% versus 7%) and/or perinatal issues (3% versus 3%) were also similar (Fisher’s-exact-tests=0.4). Limitations, reasons for caution This is a retrospective study conducted in poor prognosis patients indicated to preimplantation genetic testing for aneuploidies. Future randomized controlled trials and cost-effectiveness analysis are desirable, as well as studies in different patient populations. Lastly, each gestational/perinatal issue shall be analyzed per se (e.g. different placentation disorders). Wider implications of the findings: The absence of clinical and perinatal differences between the two protocols for endometrial preparation supports the adoption, whenever needed, of AC. This approach, in fact, allows a higher flexibility in patients’ and daily workload management. Trial registration number None

P–252 The correlation of first cleavage and blastulation timing to the euploid status of embryos after PGT-A

Abstract Study question To examine the correlation of first cleavage and blastulation timing on euploidy rates in IVF cycles after PGT-A. Summary answer The timing of blastulation is observed earlier in the euploid embryos. What is known already Embryo evaluation is one of the most critical processes that affect the clinical outcome in IVF cycles. Conventional morphologic assessment and morphokinetic assessment using time lapse technology are performed in order to select the embryo with the higher implantation potential to be transferred. It is stated that embryos with faster developmental potential, especially early forming blastocysts, show increased euploidy rate and higher implantation potential. Study design, size, duration This study includes ICSI/PGT-A treatments completed between May 2018 and December 2019. 117 blastocysts were biopsied and their euploidy status was analyzed by NGS. These embryos resulted from 32 different ICSI treatments. PGT-A was performed due to: a) repeated IVF failure, b) advanced maternal age, c) recurrent pregnancy loss.ICSI was implemented in all cases and blastocysts were vitrified awaiting the genetic results. Single euploid blastocyst transfer followed and clinical pregnancy rate was monitored. Participants/materials, setting, methods Based on the genetic results, the biopsied embryos were divided into two categories; group A representing the euploid embryos and group B the aneuploid embryos. The timing of 1st cleavage and the timing of blastulation, by means of forming a blastocoel, were investigated and compared between the two groups. The rate of early blastocysts in the two groups was also analysed. Early blastocysts are considered those formed at 96h ±2 of embryo culture post ICSI. Main results and the role of chance After the genetic analysis of the biopsied embryos, 37 blastocysts were included in group A-Euploid embryos and 80 blastocysts in group B-Aneuploid embryos. The mean time of the 1st cleavage division was similar between the two groups, with marginally no statistical significance (group A-euploid:25.9h, group B-aneuploid: 26.9h ,p&amp;gt;0.05). Regarding the blastulation time, it was achieved earlier in group A-Euploid, at a mean time of 102.6h, compared to the mean time of 106h in group B-Aneuploid (p &amp;lt; 0.05). Between the cohort of the Euploid embryos (group A), there was a higher rate of early blastulating embryos, compared to the cohort of aneuploid embryos (Group B) (24% VS 17.5%), although it was not statistically significant (p &amp;gt; 0.05). After transferring 1 euploid blastocyst, the ongoing pregnancy rate was monitored in 76.5%, independently of the 1stcleavage and blastulation time of the transferred embryo. Limitations, reasons for caution Further investigation in larger randomized studies is required, as only a limited number of cases were included in this study. Further analysis of the ongoing pregnancy rate between the euploid blastocysts, depending on other morphokinetic parameters would be of paramount significance, as well. Wider implications of the findings: High clinical pregnancy rates observed independently of the analyzed time points, indicate high success rates obtained after PGT-A/NGS. Additionally, success rates show that trophectoderm biopsy is not hazardous for the embryo viability, if performed properly. Concluding, genetic testing combined with time-lapse microscopy may provide further information to improve IVF outcomes. Trial registration number N/A

P–519 Investigation of embryo chromosomal constitution and live birth rate after vitrified-warmed euploid single blastocyst transfer across ranges of maternal body-mass-index

Abstract Study question Does maternal body-mass-index (BMI) associate with blastocysts’ chromosomal constitution and clinical outcomes in infertile patients undergoing preimplantation genetic testing for aneuploidies (PGT-A)? Summary answer A higher euploidy rate per biopsied blastocyst was reported among underweight women. Overweight women were instead subject to higher miscarriage (MR) and lower live-birth-rates (LBR). What is known already Different studies in the literature revealed an association between BMI and infertility, suggesting a J-shaped relationship: both underweight and overweight women can suffer from infertility issues. Even if IVF might increase the success rate in both these categories of patients, it seems insufficient per se to overcome the complex and multifactorial fertility impairment derived from unbalanced nutritional intakes. Miscarriage, in particular, is common in both underweight and overweight women. However, most of the literature is based on chromosomally-untested embryos. Study design, size, duration: Retrospective observational study. Only the first IVF cycle with ≥1 biopsied blastocyst from each woman was included. The primary outcome was the association between maternal BMI (underweight, BMI&amp;lt;18.5, n = 160; normal-weight, BMI=18–25, N = 1392; overweight, BMI&amp;gt;25, N = 259) and the mean euploidy rate per cohort of biopsied blastocysts (m-ER). The secondary outcomes were the association between maternal BMI with clinical (mainly MR and LBR), gestational and perinatal outcomes after first vitrified-warmed single euploid blastocyst transfers. Participants/materials, setting, methods We included 1811 women undergoing PGT-A at a private IVF center between April–2013 and March–2020. The secondary outcomes were investigated on 1125 first vitrified-warmed single euploid blastocyst transfers from all patients obtaining ≥1 transferable blastocyst. Only ICSI with ejaculated sperm and continuous culture in standard incubators were performed. Logistic regressions were conducted to identify putative confounders and adjust the results accordingly. Main results and the role of chance Except for a lower maternal age among underweight women (38.3±3.1 versus 38.9±3.4 yr, p &amp;lt; 0.01) and higher among overweight ones (39.3±3.6 yr, p = 0.04), no difference was reported with respect to normal-weight women in terms of duration of infertility, hormonal levels, main cause of infertility, sperm quality, and reproductive history. The mean number of biopsied blastocysts was ∼3 in all groups. The m-ER shows a decreasing trend as the maternal BMI increases between 17 and 22–23, to then plateau. In fact, a significant difference was reported between underweight (50.8%±36.4%) and normal-weight women (41.4%±37.5%, p &amp;lt; 0.01). A linear regression adjusted for maternal age confirmed this moderate association between increasing BMI and m-ER (unstandardized-coefficient-B –0.6%, 95%CI:–1.1% to –0.1%, p = 0.02). Morphological quality and day of full-blastulation among transferred euploid blastocysts was similar in the three groups. Overweight women showed higher MR per pregnancy (N = 20/75, 26.7%, 95%CI:17.4%–38.3% versus N = 67/461, 14.5%, 95%CI:11.5%–18.2%; OR 2.0, 95%CI:1.1–3.6, p = 0.01) and lower LBR per transfer (N = 55/154, 35.7%, 95%CI:28.3%–43.8% versus N = 388/859, 45.2%, 95%CI:41.8%–48.6%; OR adjusted for euploid blastocysts’ features 0.67, 95%CI:0.46–0.96, p = 0.03). Clinical outcomes were instead similar among underweight and normal-weight women. All gestational and perinatal outcomes were comparable in the tree groups. Limitations, reasons for caution Our study is limited by its retrospective nature, and the fact that maternal BMI was measured only before oocyte retrieval and not before embryo transfer. Moreover, the reduced sample size did not allow for further relevant sub-analyses among solely obese women. Wider implications of the findings: When possible nutritional/lifestyle modifications should be encouraged to adjust maternal BMI before IVF. Overweight patients should be especially informed of their higher risk for miscarriage. Yet, BMI is just a gross marker, future studies based on body fat localization and percentage (e.g. by bioelectrical impedance analyses) are desirable. Trial registration number None

P–515 Clinical outcome of mosaic-blastocyst transfer versus euploid-blastocyst transfer in single frozen blastocyst transfer cycles

Abstract Study question What is the clinical outcome of transferring a mosaic blastocyst versus a euploid blastocyst in single frozen blastocyst transfer (sFBT) cycles? Summary answer Single mosaic blastocyst transfer has similar clinical outcome to single euploid blastocyst transfer. What is known already Embryonic mosaicism occurs when there are two or more distinct cell lines found in preimplantation embryos derived from IVF. Data from recent studies show that mosaic blastocysts have the potential to implant and can result in healthy live births. As a result, patients now have the option of transferring mosaic blastocyst when they do not have any euploid blastocyst available for transfer. However, the clinical outcome of transferring mosaic blastocyst has not been definitively reported. Thus, a retrospective study was conducted to compare the clinical outcome of mosaic sFBT and euploid sFBT. Study design, size, duration A total of 602 patients underwent frozen blastocyst transfer in Alpha IVF from January to October 2019 and had their blastocysts screened for aneuploidy. These patients were divided into 2 groups: 26 patients with mosaic blastocysts transferred (Group A, age ranged 19–44), and 576 patients with euploid blastocysts transferred (Group B, age ranged 21–44). The mean age of patients from Group A and B were 34.0 and 32.8 respectively (p &amp;gt; 0.05). Participants/materials, setting, methods All samples had their DNA libraries constructed for sequencing using Next Generation Sequencing according to manufacturer’s specification (IonTorrent, USA). All blastocysts were frozen for subsequent sFBT cycle (Cryotech, Japan). All thawed blastocysts for sFBT survived with morphologically intact inner cell mass and trophectoderm cells. The importance of antenatal confirmation of the fetal chromosome status was emphasized in patients from Group A. The clinical outcomes of both groups were analysed and compared. Main results and the role of chance No significant differences were seen in the clinical pregnancy and implantation rate of Group A and B (65.4% vs 63.0%; p &amp;gt; 0.05). The miscarriage rate of Group A and B were 23.5% and 14.0% respectively. Albeit the higher miscarriage rate in Group A, there was no statistical significance between these two groups (p &amp;gt; 0.05). Group A was further divided into two subgroups, Subgroup A1: low risk mosaic blastocyst transfer; Subgroup A2: high risk mosaic blastocyst transfer. In the comparison of Group A subgroups, the clinical pregnancy and implantation of Group A1 is higher than Group A2 (76.9% vs 44.4%). In addition, the miscarriage rate of Group A1 and A2 were 23.1% and 0.0% respectively. Interestingly, there was no statistical significance in clinical pregnancy rate, implantation rate and miscarriage rate between these two subgroups. Limitations, reasons for caution This is a retrospective study and the sample size was comparatively smaller in the mosaic blastocyst transfer group than the euploid blastocyst transfer group. Further studies with a larger sample size should be carried out to ascertain the clinical outcome. Wider implications of the findings: Single mosaic blastocyst transfer has similar clinical outcome to single euploid blastocyst transfer. Thus, mosaic blastocyst can be considered for transfer when no euploid blastocyst are available. Nevertheless, stringent antenatal surveillance for chromosomal abnormalities to confirm the chromosomal status of the fetus must be followed. Trial registration number Not applicable

P–415 Clinical and In Vitro Fertilization laboratory parameters that contribute to clinical miscarriage after single euploid embryo transfer

Abstract Study question To investigate which factors, excluding embryo aneuploidies, are associated with miscarriage in patients who have undergone a single euploid blastocyst transfer. Summary answer Miscarriage was related to the body mass index (BMI), the type of cycle and the thickness of the endometrium. What is known already Preimplantation genetic testing for aneuploidies (PGT-A) is widely used in-vitro fertilization (IVF) to select euploid embryos. Several studies have shown that embryo aneuploidy is the main contributing factor for IVF failure, reinforcing the relevance of PGT-A as a method to select chromosomally normal embryos. A recent meta-analysis confirmed that patients undergoing PGT-A have a lower miscarriage rate than women that conceived naturally (9% vs 28%, respectively). Even though most of the studies show that PGT-A significantly reduces miscarriage rate, still some women do lose their pregnancies. We investigated which other reasons may be related to this early pregnancy loss. Study design, size, duration Retrospective, observational, and multicenter study of 6910 patients undergoing single euploid blastocyst transfer after PGT-A from January 2017 to December 2019 in our institution. Several laboratory and clinic variables were analyzed to study the effect of these variables on the miscarriage rate Participants/materials, setting, methods Indications for PGT-A were advanced maternal age, implantation failure, recurrent pregnancy loss and male factor. Embryos were cultured 5% O2 concentration and 6.5% CO2 concentration. Trophectoderm biopsy was performed on day 5/6 of development and analyzed through Next Generation Sequencing (NGS); embryos were vitrified until transfer was performed. Single euploid embryo transfer was performed in all cases. We performed a multivariate regression analysis to compare the different variables and search for there are significant differences. Main results and the role of chance We studied a total of 6910 patients undergoing PGT-A to describe which factors, excluding embryo aneuploidies, were correlated with miscarriage in patients who underwent single thawed euploid embryo transfer. When considering embryo morphology (embryo grading, quality of inner cell and quality of trophectoderm), we did not find differences in miscarriage rate among groups (high quality= 15.9%; normal quality= 14.3%; low quality= 15.0%; poor quality= 14.8%) p = 0.833. BMI was significantly associated with miscarriage rate (odds ratio [OD] 1.04; 95% confidence interval [CI], 1.012–1–076 p = 0.006) and miscarriage rate. We observed a weak association between endometrial thickness and miscarriage rate ([OD] 0.65; 95%, 0.528–0.778 p = 0.04) and also between type of endometrial preparation (natural cycle or hormone replacement cycle) ([OD] 0.77; 95%, 0.528–0.778) p = 0.04. Body mass index, according to our findings, was the main variable correlated with miscarriage rate. We did not find any association with the other variables studied (biopsy day, maternal age, male age, duration infertility, cycle length, previous miscarriage, previous live birth, previous cycles IVF, endometrial pattern and diagnosis). Limitations, reasons for caution The retrospective study design limits the generalization of our results but offers a good insight to be validated in prospective trials. Wider implications of the findings: According to our findings, BMI, endometrial thickness the day of the embryo transfer, and the type of endometrial preparation should be considered when transferring an euploid blastocyst. Trial registration number NO APLICA

Which factors affect the likelihood of miscarriage after single euploid blastocyst transfer?

Research questionWhich parameters affect the likelihood of miscarriage after single euploid frozen–thawed blastocyst transfer (FBT)? DesignIn this retrospective study, clinical and laboratory data from 1051 single euploid FBTs were evaluated. Exclusion criteria were endocrine or systemic pathologies, uterine anomalies or pathologies, unilateral or bilateral hydrosalpinx, karyotypic abnormalities (either maternal or paternal) or thrombophilia. Patients were divided into two groups according to pregnancy outcome: live birth and miscarriage. ResultsBody mass index (BMI) (25.98 ± 0.5 versus 24.36 ± 0.21, P = 0.019), duration of infertility (6.62 ± 0.54 versus 4.92 ± 0.18, P = 0.006) and number of previous miscarriages (1.36 ± 0.13 versus 0.79 ± 0.05, P < 0.001) were significantly higher in the miscarriage group (n = 100) than in the live birth group (n = 589). Although the trophectoderm and inner cell mass (ICM) percentage scores were not statistically different among the miscarriage and live birth groups, the percentage of day-6 biopsied embryos was significantly higher in the miscarriage group. Binary logistic regression analysis revealed that BMI (OR 1.083, 95% CI 1.013 to 1.158, P = 0.02) and number of previous miscarriages (OR 1.279, 95% CI 1.013 to 1.158, P = 0.038) were independent factors for miscarriage. Patients with elevated BMI and a higher number of miscarriages were at increased risk of miscarriage. ConclusionAfter a single euploid FBT, BMI and number of previous miscarriages are predictors of miscarriage. Lifestyle interventions before FBT may decrease miscarriage rates.

Deep learning neural network analysis of human blastocyst expansion from time-lapse image files

Research questionCan artificial intelligence (AI) discriminate a blastocyst's cellular area from unedited time-lapse image files using semantic segmentation and a deep learning optimized U-Net architecture for use in selecting single blastocysts for transfer? DesignThis platform was retrospectively applied to time-lapse files from 101 sequentially transferred single blastocysts that were prospectively selected for transfer by their highest expansion ranking within cohorts using a 10 h expansion assay rather than standard grading. ResultsThe AI platform provides expansion curves and raw data files to classify and compare blastocyst phenotypes within both cohorts and populations. Of 35 sequential unbiopsied single blastocyst transfers, 23 (65.7%) resulted in a live birth. Of 66 sequential single euploid blastocyst transfers, also selected for their most robust expansion, 49 (74.2%) resulted in live birth. The AI platform revealed that the averaged expansion rate was significantly (P = 0.007) greater in euploid blastocysts that resulted in live births compared with those resulting in failure to give a live birth. The platform further provides a framework to analyse fragmentation phenotypes that can test new hypotheses for developmental regulation during the preimplantation period. ConclusionsAI can be used to quantitatively describe blastocyst expansion from unedited time-lapse image files and can be used to quantitatively rank-order blastocysts for transfer. Early clinical results from such single blastocyst selection suggests that live birth rates without biopsy may be comparable to those found using single euploid blastocysts in younger, good responder patients.

Oocyte competence is independent of the ovulation trigger adopted: a large observational study in a setting that entails vitrified-warmed single euploid blastocyst transfer.

To assess whether the GnRH-agonist or urinary-hCG ovulation triggers affect oocyte competence in a setting entailing vitrified-warmed euploid blastocyst transfer. Observational study (April 2013-July 2018) including 2104 patients (1015 and 1089 in the GnRH-a and u-hCG group, respectively) collecting ≥1 cumulus-oocyte-complex (COC) and undergoing ICSI with ejaculated sperm, blastocyst culture, trophectoderm biopsy, comprehensive-chromosome-testing, and vitrified-warmed transfers at a private clinic. The primary outcome measure was the euploid-blastocyst-rate per inseminated oocytes. The secondary outcome measure was the maturation-rate per COCs. Also, the live-birth-rate (LBR) per transfer and the cumulative-live-birth-delivery-rate (CLBdR) among completed cycles were investigated. All data were adjusted for confounders. The generalized-linear-model adjusted for maternal age highlighted no difference in the mean euploid-blastocyst-rate per inseminated oocytes in either group. The LBR per transfer was similar: 44% (n=403/915) and 46% (n=280/608) in GnRH-a and hCG, respectively. On the other hand, a difference was reported regarding the CLBdR per oocyte retrieval among completed cycles, with 42% (n=374/898) and 25% (n=258/1034) in the GnRh-a and u-hCG groups, respectively. Nevertheless, this variance was due to a lower maternal age and higher number of inseminated oocytes in the GnRH-a group, and not imputable to the ovulation trigger itself (multivariate-OR=1.3, 95%CI: 0.9-1.6, adjusted p-value=0.1). GnRH-a trigger is a valid alternative to u-hCG in freeze-all cycles, not only for patients at high risk for OHSS. Such strategy might increase the safety and flexibility of controlled-ovarian-stimulation with no impact on oocyte competence and IVF efficacy.

Comparison of single euploid blastocyst transfer cycle outcome derived from embryos with normal or abnormal cleavage patterns

Research questionTo assess incidence of abnormal cleavage among biopsied blastocysts; to compare euploidy rates of the blastocysts with abnormal and normal cleavage; and to compare single euploid blastocyst transfer (SEBT) outcome derived from embryos with normal or abnormal cleavage. DesignRetrospective analysis of prospectively collected data in a private IVF clinic. Consecutive 554 patients (749 cycles) undergoing preimplantation genetic testing for aneuploidy (n = 497; 671 cycles) or monogenic diseases (n = 57; 78 cycles) were included. All assessments for abnormal cleavage were carried out retrospectively; presence of abnormal cleavage was not a factor in deciding which euploid embryo to transfer. A total of 1015 blastocysts were biopsied and 295 SEBT procedures were carried out. Main outcome measure was live birth rate (LBR). ResultsIncidence of reverse cleavage, direct cleavage, and reverse plus direct cleavage, were 7.7%, 6.4% and 2.3%, respectively. Of the 1015 biopsied blastocysts, 35.0% were euploid. Blastocysts with abnormal cleavage, in total, had a significantly higher euploidy rate compared with blastocysts with normal cleavage (44.6% [74/166] versus 33.1% [281/849]; P = 0.017). The LBR after SEBT with normal, reverse and direct cleavage, and direct cleavage plus reverse cleavage, was 133/238 (55.9%), 6/26 (23.1%), 8/24 (33.3%) and 0/3 (0.0%) (P < 0.001). Generalized estimating equation analysis showed that the presence of abnormal cleavage pattern was the only independent predictor of LBR (OR 0.316; 95% CI 0.115 to 0.867; P = 0.013). ConclusionsBlastocysts with direct or reverse cleavage should be biopsied in preimplantation genetic testing cycles if they are morphologically eligible. Euploid blastocysts with abnormal cleavage, however, have approximately half the LBR of those euploid blastocyst with normal cleavage, hence, blastocysts with abnormal cleavage should have lower priority for transfer.

Endometriosis shows no impact on the euploid blastocyst rate per cohort of inseminated metaphase-II oocytes: A case-control study

ObjectiveTo evaluate the true impact of endometriosis on oocytes’ competence defined as blastulation, euploidy and implantation rates. DesignRetrospective multicenter case-control study involving infertile couples undergoing ICSI with qPCR and trophectoderm biopsy-based PGT-A. Patients affected from endometriosis (n = 210) were diagnosed through transvaginal sonography or surgical history with histological confirmation. Each case was matched to two controls (n = 420) according to IVF clinic, maternal age at retrieval (38.6 ± 2.7 yr), number of previous failed IVF treatments (0.5 ± 0.8) and number of metaphase-II oocytes retrieved (6.1 ± 3.7 per patient). The primary outcome was the mean euploid blastocyst rate per cohort of inseminated metaphase-II oocytes. Other embryological, clinical, obstetric and neonatal outcomes were also evaluated. ResultsThe mean euploid blastocyst rate per cohort of inseminated metaphase-II oocytes was identical in the two groups (18 %±22 %) independently of maternal age. No difference was shown for all embryological outcomes investigated. The live birth rates per vitrified-warmed single euploid blastocyst transfer were also similar (67/158, 42 % in patients affected from endometriosis versus 132/327, 40 % in matched-controls). No difference was reported in the gestational and neonatal outcomes. The cumulative live birth delivery rates among completed cycles were also identical (61/201, 30 % versus 117/391, 30 % in endometriosis and matched-control groups, respectively) independently of maternal age. ConclusionsEndometriosis might not impair oocyte developmental and reproductive competence, although its potential impact on the number of metaphase-II oocytes retrieved cannot be ignored. This information is critical for clinicians during counseling to outline an effective strategy to treat infertile patients affected from this condition. Future prospective studies are needed to evaluate the impact of endometriosis stage on euploidy rates.

The euploid blastocysts obtained after luteal phase stimulation show the same clinical, obstetric and perinatal outcomes as follicular phase stimulation-derived ones: a multicenter study.

The euploid blastocysts obtained after luteal phase stimulation show the same clinical, obstetric and perinatal outcomes as follicular phase stimulation-derived ones: a multicenter study.

Parameters impacting the live birth rate per transfer after frozen single euploid blastocyst transfer.

BackgroundTo assess the predictive value of patient characteristics, controlled ovarian stimulation and embryological parameters on the live birth outcome of single euploid frozen-warmed blastocyst transfer (FBT).MethodsThis was a retrospective cohort study including 707 single FBTs after preimplantation genetic testing for aneuploidy (PGT-A) that were performed from October 1, 2015, to January 1, 2018. The effects of patient-, cycle- and embryology-related parameters on the live birth outcome after FBT were assessed.ResultsIn the subgroup analysis based on live birth, patients who achieved a live birth had a significantly lower body mass index (BMI) than patients who did not achieve a live birth (22.7 (21.5–24.6) kg/m2 vs 27 (24–29.2) kg/m2, p<0.001). The percentage of blastocysts with inner cell mass (ICM) A or B was significantly higher among patients achieving a live birth, at 91.6% vs. 82.6% (p<0.001). Day-5 biopsies were also more prevalent among patients achieving a live birth, at 82.9% vs 68.1% (p<0.001). On the other hand, the mitochondrial DNA (mtDNA) levels were significantly lower among cases with a successful live birth, at 18.7 (15.45–23.68) vs 20.55 (16.43–25.22) (p = 0.001). The logistic regression analysis showed that BMI (p<0.001, OR: 0.789, 95% CI [0.734–0.848]), day of trophectoderm (TE) biopsy (p<0.001, OR: 0.336, 95% CI [0.189–0.598]) and number of previous miscarriages (p = 0.004, OR: 0.733, 95% CI [0.594–0.906]) were significantly correlated with live birth. Patients with elevated BMIs, cycles in which embryos were biopsied on day-6 and a higher number of miscarriages were at increased risks of reduced live birth rates.ConclusionA high BMI, an embryo biopsy on day-6 and a high number of miscarriages negatively affect the live birth rate after single euploid FBT.

Pregnancy outcomes after frozen-thawed single euploid blastocyst transfer following IVF cycles using GNRH agonist or HCG trigger for final oocyte maturation.

To assess whether GnRH agonist trigger impacts the implantation potential of euploid embryos. Retrospective cohort study done at an academic IVF center evaluating frozen-thawed embryo transfer (FET) cycles in which single-euploid blastocysts were transferred between 2014 and 2019. All embryos were generated in an IVF cycle which used GnRHa or hCG trigger and then were transferred in a programmed or natural FET cycle. Only the first FET cycle was included for each patient. Primary outcome was ongoing pregnancy rate or live birth rate (OPR/LBR). Secondary outcomes were implantation rate (IR), clinical pregnancy rate (CPR), clinical loss rate (CLR), and multiple pregnancy rate (MPR). Logistic regression was performed to control for confounding variables. A p value of < 0.05 was considered statistically significant. Two hundred sixty-three FET cycles were included for analysis (GnRHa = 145; hCG = 118). The GnRHa group was significantly younger (35.2 vs. 37.5 years) and had higher AMH values (4.50 ng/ml vs. 2.03 ng/ml) than the hCG group, respectively (p < 0.05). There was no significant difference in OPR/LBR (64.1% (93/145) vs. 65.3% (77/118); p = 0.90) between the GnRHa and hCG groups, respectively. There was also no significant difference in IR, CPR, CLR, or MPR between groups. After controlling for confounding variables, the adjusted odds ratio for OPR/LBR was 0.941 (95% CI, 0.534-1.658); p = 0.83) comparing GnRHa to hCG. Pregnancy outcomes did not significantly differ when groups were stratified by age (< 35 vs. > 35 years old). Our findings confirm that euploid embryos created after hCG or GnRHa trigger have the same potential for pregnancy.

In vitro Fertilization with Single-nucleotide Polymorphism Microarray-based Preimplantation Genetic Testing for Aneuploidy Significantly Improves Clinical Outcomes in Infertile Women with Recurrent Pregnancy Loss: A Randomized Controlled Trial

Objective: To evaluate the effect of preimplantation genetic testing for aneuploidy (PGT-A) in infertile patients with recurrent pregnancy loss (RPL). Methods: A prospective randomized clinical trial was performed in a university-affiliated fertility center in Shanghai, China. Patients in the PGT-A group underwent blastocyst biopsy followed by single-nucleotide polymorphism microarray-based PGT-A and single euploid blastocyst transfer, whereas patients in the control group underwent routinein vitro fertilization/ICSI procedures and frozen embryo transfer of 1–2 embryos selected according to morphological standards. Results: Two hundred and seven infertile patients with RPL were included in this study and randomly assigned to either the control or the PGT-A group. Baseline variables and cycle characteristics were comparable between the two groups. The results showed that PGT-A significantly improved the ongoing pregnancy rate (55.34% vs. 29.81%) as well as the live birth rate (48.54% vs. 27.88%) and significantly reduced the miscarriage rate (0.00% vs. 14.42%) on a per-patient analysis. A significant increase in cumulative ongoing pregnancy rates over time was observed in the PGT-A group. Subgroup analysis showed that the significant benefit diminished for patients who attempted ≥2 PGT-A cycles. Conclusions: PGT-A significantly improved the ongoing pregnancy and live birth rate, while reduced miscarriage rate in infertile RPL patients. However, the significance diminished in patients attempting ≥2 cycles; thus, further studies are warranted to explore the most cost-effective number of attempts in these patients to avoid overuse.

Timing of blastocyst development is the sole predictor of positive pregnancy outcome in advanced maternal age patients following single euploid blastocyst transfer

Improving embryo quality and selection is imperative to achieve a shorter time to pregnancy and successful live birth outcome, specifically for women of advanced maternal age (AMA). Preimplantation genetic testing for aneuploidy (PGT-A) allows for selection of euploid embryos, beneficial for this older population that are at increased risk for oocyte aneuploidy. The purpose of this study was to identify effective predictors of reproductive outcomes for AMA patients in conjunction with a euploid single embryo transfer (SET). Retrospective cohort study. AMA patients (≥ 38 years; n=847) underwent routine oocyte retrieval, ICSI, blastocyst biopsy, PGT-A and subsequent euploid SET at a single large infertility clinic. Patients presented with various infertility diagnoses and without significantly compromised ovarian reserve. Reproductive outcomes were divided into three groups; successful live birth (n=504), positive implantation followed by pregnancy loss (ultrasound ± FHT; n=81) and negative/biochemical (negative, or rise and subsequent direct fall of βhCG; n=262). Statistical analyses included Chi Square test for independence and ANOVA where appropriate, with significance at P<0.05. Maternal age at oocyte retrieval was significantly associated with the likelihood of an embryo transfer, as the percent of aneuploid-only cycles increased significantly with age; 38-40 years (20%), 41-42 years (46%), 43+ years (76%; P<0.0001). Reproductive outcomes following euploid SET were not associated with any of the following: maternal age at oocyte retrieval (mean 40.0 years), total number of oocytes retrieved, total number of MII oocytes, number of blastocysts available for biopsy, % euploid blastocysts, or the inner cell mass (ICM) and trophectoderm grade. Pregnancy outcomes following euploid SET were only significantly associated with the timing of blastocyst development and the appearance of the ICM (P<0.0001). Euploid blastocysts that resulted in negative/biochemical outcomes were significantly over-represented by slower blastocyst development (52.3%) compared to those with live birth (37.5%) and pregnancy loss (38.3%) (P<0.0001). Remarkably, no differences were observed between successful live birth and positive implantation followed by pregnancy loss for any parameters analyzed. Interestingly, a second euploid SET for AMA patients with prior pregnancy loss resulted in 58.3% successful live births (n=36), comparable to the overall first SET live birth rate (59.5%; ns). The timing of blastocyst development and the appearance of the ICM was the sole predictor of positive pregnancy in AMA patients following a euploid SET, and thus should be highly considered for euploid blastocyst selection to achieve the fastest time to pregnancy. Further investigations are essential to identify potential predictors of euploid pregnancy loss, and the stochastic occurrence of this adverse outcome.

Correlation between blastocyst stage of expansion and clinical outcome: a retrospective analysis of 810 single euploid blastocyst transfer cycles at a single IVF center

Correlation between blastocyst stage of expansion and clinical outcome: a retrospective analysis of 810 single euploid blastocyst transfer cycles at a single IVF center

Does euploidy on day 5 or day 6 matter for clinical outcome in frozen single blastocyst transfers?

Introduction Comprehensive chromosomal screening at the blastocyst stage by aCGH/NGS analysis has become increasingly common approach in order to obtain high implantation rates in ART. Although utilization of chromosomally screened blastocyst-stage embryos is known to provide superior clinical outcome, not all blastocysts display the same pace of embryo development and considerable number of embryos show delay in blastulation towards day 6 or day 7 instead of day 5. Attitudes of clinics towards delayed blastocysts largely differ in their PGT-A programs. Also, current literature on the comparative analysis of aneuploidy and clinical outcome of single euploid blastocyst transfer (SEBT) cycles involving a day 5 or a day 6 euploid blastocyst is scarce and contradictory. This study was conducted to analyze and compare the laboratory and clinical outcome of SEBT cases involving day 5 or day 6 euploid blastocysts screened by NGS. Material & Methods This retrospective study was conducted in Bahceci Fulya IVF Centre, between January 2016 and December 2017. It included a total of 1938 PGT-A cycles with indications of advanced maternal age, recurrent implantation failure or pregnancy loss. For all suitable embryos, a trophectoderm(TE) biopsy was performed on either on day 5 or day 6 and next generation sequencing (NGS) was used for comprehensive chromosomal screening (CCS) analysis. Of them, 658 frozen embryo transfers were performed as a single euploid blastocyst transfer (SEBT) cycle, with embryos biopsied on either day 5 (Group I) or day 6 (Group II). Major laboratory as well as clinical parameters were compared between groups. Results Main patient & cycle parameters were found to be similar in both SEBT groups (p>0.05). A total of 3501, 3111, 3010 (96.8%) and 1496 (49.7%) embryos in Group I and a total of 1540, 1280, 1242 (97.0%) and 511 (41.1%) embryos in Group II were biopsied, analyzed, diagnosed and detected as euploid respectively. Aneuploidy rate was found to be significantly high in embryos biopsied on day 6 (p=0.0024). Regarding laboratory parameters, although no difference was observed in terms of aneuploidy rates among embryos with different ICM scores, percent of embryos with trophectoderm score C was significantly high in Group II (p=0.018). Clinical outcome is shown in Table 1. Table 1: Clinical outcome between SEBT groups utilizing a day 5 or a day 6 blastocyst. Group I Group II p Num. of cycles (n) 498 160 Pregnancy rate (b-hCG+, %) 79.1 55.6 p Clinical pregnancy rate (Sac+, %) 72.7 52.5 p Live birth/Ongoing (%) 64.1 42.5 p Missed abortus (%) 11.3 17.9 NS Conclusions Our data and results have shown that, although they generate acceptable clinical pregnancies, euploid embryos that are obtained from embryo biopsies performed on day 6 result in significantly lower clinical outcome when compared to euploid embryos that are biopsied on day 5. This result further suggests that when there are more than one euploid embryo for SEBT, priority should be given to day 5-biopsied embryos in order to obtain better clinical outcome.