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Objectives: The aim of this study is to evaluate the association between changes in C reactive protein and pregnancy rate in IVF/ICSI. Patients and methods: A prospective cohort study that was conducted in Ain Shams University Maternity Hospital, Infertility Clinic during period time from September 2018 to September 2020. A total of 100 infertile women who were candidate for IVF/ICSI, over a period of 2 years, beginning in September 2018 were recruited to the study after they met the inclusion and exclusion criteria. The participants gave written informed consent and approval for the ethical aspects of the study. Results: This study showed no statistically significant differences according to clinical pregnancy regarding the sociodemographic data as BMI, duration of infertility and type of infertility and regarding Oocyte retrieval and embryo transfer, also no significant differences according to clinical pregnancy regarding CRP at baseline and at Oocyte pickup as well as CRP change at oocyte pickup. But age was significantly lower in cases with clinical pregnancy and CRP at embryo transfer and CRP change at embryo transfer were significantly higher in cases with positive pregnancy. Conclusion: Finally, we concluded that patients whose CRP level decreased on transfer day, had lower chance of pregnancy, whereas patients whose CRP level elevated on embryo transfer day had high chance of pregnancy

Introduction: The purpose of the present study to determine the presence of Von Willebrand factor (vWF) in the follicular fluid of women undergoing IVF/ICSI and to investigate the association of its concentration with oocyte and embryo quality. Methods: We conducted a prospective observational study based in women undergoing in vitro fertilization treatment. vWF concentrations in follicular fluid samples collected during transvaginal oocyte retrieval were measured using an enzyme-linked immunosorbent assay (Elisa). They were also analyzed by the VIDAS® vWF method for confirmation of the results. The latter is an automated quantitative test for the measurement of vWF in human plasma, using the ELFA technique (enzyme-linked fluorescent assay). vWF concentrations were correlated to oocyte and embryo quality. Results: Contrary to previous reports, vWF was determined in follicular fluid. A higher percentage of good quality oocytes were found in the presence of lower vWF follicular fluid levels. Similarly, better quality embryos were derived from oocytes retrieved from follicles with lower vWF follicular fluid levels. Conclusions: Contrary to existing evidence to date, vWF was found in the follicular fluid of women undergoing IVF/ICSI treatment. In addition, the quality of oocytes and embryos was adversely related to vWF follicular fluid levels. In fact, a better outcome of IVF/ICSI treatment was suggested when the levels of vWF in follicular fluid were low. In this context, vWF follicular fluid levels could be used as a prognostic factor for oocyte and embryo quality.

RESEARCH QUESTION: How do cumulative live birth rates (CLBR), cumulative multiple live birth rates (CMLBR) and dropout rates over six IVF and intracytoplasmic sperm injection (ICSI) cycles change over time? DESIGN: Prospective longitudinal cohort (n = 16,073 patients; 48,946 cycles) starting a first fresh assisted reproductive technology cycle between 1 January 2014 and 31 December 2016, with follow-up until 31 December 2017. Outcomes between the periods 2014-2017 and 2009-2012 were compared. RESULTS: Conservative estimates of CLBR after six complete cycles were significantly higher in women younger than 35 years after every cycle: one to three, adjusted P-value [p adj] < 0.0001; four, p = 0.01; five, p adj = 0.03; six, p adj = 0.04) and after the first cycle in women aged 35-37 years (p adj = 0.04) in 2014-2017 versus 2009-2012. For an optimal estimate, the CLBR was significantly higher after the first three cycles in women younger than 35 years (all p adj < 0.0001) and after the first cycle in women aged 35-37 years (p adj = 0.04). The CMLBR rate decreased from 5.1% ± 0.19 (SE) to 4.1% ± 0.16 for the conservative estimate and from 8.6% ±0.37 (SE) to 6.7% ± 0.30 for the optimal estimate after six complete cycles for the whole cohort. Dropout rates of complete cycles were 26.5% 29.4%, 33.4%, 38.9% and 47.3% after the first to fifth cycle, respectively. Compared with 2009-2012, the dropout rate in the current period was significantly higher for the first (P < 0.0001) and second (P = 0.0124) cycle. CONCLUSION: Over six complete IVF/ICSI cycles, CLBR and dropout rates increased and multiple live birth rates decreased when 2014-2017 was compared with 2009-2012. ispartof: REPRODUCTIVE BIOMEDICINE ONLINE vol:42 issue:4 pages:717-724 ispartof: location:Netherlands status: published

Introduction: Infertility is a global public health problem; cost-effective patient-friendly treatment modalities along with psychosexual intervention strategies are essential for infertility control/prevention/management among ethnically disparate populations. Objectives: This study aimed to assess differential in vitro fertilization (IVF) success trends among infertile women of South Indian ethnicity. Materials and Methods: Prospective, observational study designed in a hospital-based setting with active enrollment of infertile women undergoing IVF/intracytoplasmic sperm injection (ICSI) at Indira IVF Center, Chennai, Tamil Nadu, India (April-September 2019); inclusion criteria: age >35 years, South Indian ethnicity, married >1 year, absence of full-term clinical pregnancy, endometrial thickness Results: Mean age and endometrial thickness of study participants were 33.3 years (SD ±1.9) and 8.7 mm (SD ±0.5), respectively; average Body Mass Index (BMI) and anti-müllerian hormone (AMH) levels were 28.4 kg/m2 and 4.7 ng/mL, respectively. Embryos transferred/month: 7 in April/13 in May/24 in June/36 in July/24 in August/30 in September, and pregnancies: 4/8/17/26/20/22 for the months of April, May, June, July, August, and September 2019, respectively. Subgroup stratification demonstrated M-II vs total oocytes retrieved were 76%, 73%, 60%, 71%, 77%, and 77%. Overall IVF success rates were 57% in April/62% in May/71% in June/72% in July/83% in August/73% in September; frozen embryo transfer success was 0 in April-May/67% in June/73% in July/89% in August/100% in September 2019. English/Tamil-speaking infertile women self-reported treatment-related satisfaction rates of 80% to 100%. Conclusion: M-II oocytes’ yield, sociodemographics of infertile women, and increasing age/aberrant AMH/BMI profiles/endometrial receptivity/diminished ovarian reserve are promising predictors of IVF/ICSI success in genetically distinct patient population subset(s). Future multicentric gene epidemiology studies with larger sample size and precision-based psychiatric assessments/interventions are warranted for development of predictive biomarkers in infertility management.

Objective Ovarian response from a conventional ovarian stimulation protocol is a crucial step in IVF/ICSI treatments. This ovarian response encompasses a wide range of outcomes at the extremes, leading to either excessive responses with the risk of life-threatening conditions like ovarian hyperstimulation syndrome (OHSS), or poor ovarian response (POR) with poor outcomes. This study aims to integrate biochemical, ultrasonographic and demographic parameters into a mathematical formula able to predict ovarian response to stimulation in IVF/ICSI in gonadotropin-releasing hormone (GnRH) antagonist protocols. Methods This retrospective analysis included 147 patients submitted to an ovarian stimulation protocol combining recombinant FSH and gonadotropin-releasing hormone antagonist. All the parameters were correlated with the Spearman Rho and Pearson´s correlation coefficient. Once the data was normalized, we used the multiple linear regression models, checking the results with the progressive discriminating analysis. Results We classified the database according to the correlation with the number of oocytes retrieved; the progressive discriminating analysis resulted in the following equation: oocytes retrieved = 2.312-0.130 (FSH) + 0.562 (AFC). Conclusions The incorporation of 2 ovarian reserve parameters into a regression equation enables knowing the number of retrieved oocytes in each patient with 80.5% sensitivity and 55.4% specificity.

Background: Maternal subclinical hypothyroidism (SCH) is a risk factor for adverse pregnancy outcomes. However, it is still unclear whether SCH affects male fertility. The aim of this study was to ...

Our group carefully read the article published by Vieira and Colucci (2013) in your prestigious journal. We agree that any assisted reproductive technique causes stress to the couple and that this stress is higher in women in comparison with men (El Kissi et al., 2013). However, in the mentioned paper, we found several remarkable mistakes, which resulted in conclusions that were not supported by the data. First, the reported results have a serious methodological bias because the study was not randomized. Additionally, because the baseline patient characteristics were not presented, we do not know whether the groups (IVF-ICSI treatment and INVO treatment) were comparable. The psychological characteristics of the patients before ART were not shown. Second, the study did not provide any power analysis, and more importantly, no statistical comparisons were performed. This is the main problem, as the conclusions were based on different responses to stress in patients treated using either INVO or ICSI. However, the row data included in the article support statistical analysis using Fisher’s exact test; if the authors had performed such an analysis, no differences (P=0.5273) (http://www.graphpad.com/quickcalcs/contingency2/) would have been observed between the studied groups. Consequently, no statement arguing for differences between the two techniques can be made. Finally, the authors did not mention the clinical outcomes of the patients who underwent either ICSI or INVO treatment.

Kisspeptin, a crucial central regulator of reproduction, has been used as a trigger in in vitro fertilization (IVF) treatment. This study aimed to investigate the roles of kisspeptin in IVF treatment in infertile females ( n = 30); and in steroidogenesis in human granulosa-like tumor cell line (KGN). In the human study, blood was collected at three time points including (1) the beginning of gonadotropin stimulation (Phase I), (2) around eight days after gonadotropin stimulation (Phase II), and (3) on the day of ovum pick-up (Phase III). Follicular fluid (FF) was collected at Phase III. Serum human chorionic gonadotropin (hCG) was measured 15 days after embryo transfer and fetal heart beats were determined around 42 days of menstrual cycle to classify the subjects into successful and unsuccessful groups. FF kisspeptin levels were higher in successful compared with unsuccessful subjects ( P -2 mmol/L with FSH enhanced aromatase concentrations in the supernatant compared with FSH alone ( P

A grading system of human embryo is very important for embryo selection & predicting blastocyst formation from day - 1, day 2, & day – 3, were sequentially explained. We were designed a methods for grading of embryos from day – 1 (zygote as a pronuclear evaluation, PN), day – 2 (cleavage stage) and day - 3 (evaluation of developmental stage according to cell size and fragmentation) embryo according to cell size and degree of fragmentation and early compaction. In the first grading system pronuclear study and poly-spermy, Second grading system is based on the blastomere / cell number and the observation of fragmentation pattern and selection for embryo transfer, embryos vitrification and pregnancy outcome. Assessment of embryo quality in order to select the embryos that have higher chance to give pregnancy, it is critical goal in IVF cycle or assisted reproductive technologies. ET current trend in human infertility treatment with IVF / ICSI embryo transfer (IVF / ICSI ET) is to increase the chance of higher pregnancy and reduce the multiple pregnancies after multiple embryo transfer according to patient age and endometrium thickness as well as own ART Laboratory protocol. Morphological evaluation & grading of human embryo as a reliable and no-invasive method that provides valuable information & prediction of IVF/ICSI embryos which has developmental potential to reach till early compaction or blastocyst. This research paper describes the current status of morphological embryo evaluation from zygote to eight cell blastomeres or early compaction on late day 3. We found higher embryo development potential and early compaction during in-vitro embryo culture conditions and higher implantation rate in Grade A embryos in comparison transferred of Grade B embryos. Overall embryo development in-vitro and conceiving rate was seen 48.06 % after embryo transfer of both grades A and Grade B embryos in 233 patients in different age group with different endometrium thickness and multiple embryos transfer in one uterus depend on patient previous history. 935Embryos was selected from 2702 developing embryos for embryo transfer (ET) was performed in 233 patients. After this study we found implantation rate (IR) was 48.06% based on embryo quality, morphology and grade.

Background: Polycystic ovary syndrome (PCOS) is a common endocrine disorder of women, which occurs in 5% to 13% in women of reproductive age. It is one of the major reason of infertility in women. Therefore, the prediction of clinical pregnancy in women with PCOS is more challenging. Aim: Our aim was to investigate the IVF/ICSI outcomes in PCOD patients. Material and Methods: In the present retrospective study, total of 126 women in 133 fresh IVF cycles were involved, where 42 patient were PCOD and 84 patient were non-PCOD (other factor of infertility). All the patients’ medical data of age, duration of infertility, factor of infertility, AMH hormone level, number of total retrieved oocyte, matured (M2) oocytes, day 3 cleaved embryos and biochemical pregnancy rates were used for this study and data were statistically analyzed by Student T-Test. Result: After statistical analysis, significantly higher AMH level, higher number of total oocyte, mature oocyte and day3 cleaved embryos were observed. The biochemical pregnancy rate was also found to be higher in PCOD patient but it was not statistically significantly. Conclusion: In the present study, number of oocytes and cleaved embryos were found to significantly higher in PCOD group but biochemical pregnancy was not significant. Keywords: PCOD, AMH, Oocyte, Embryo, Biochemical pregnancy.

Background: Several studies have demonstrated that vitamin D (vitD) might play an important role in the reproductive system due to expression of vitD receptor and vitD-metabolizing enzymes in many reproductive tissues. VitD deficiency has been associated with increased risk of obstetric complications. However, the effect of vitD levels on in vitro fertilization (IVF)/ICSI outcomes is not fully understood. Evidence shows that women with adequate vitD levels might have higher pregnancy rates. This study evaluated the association between serum vitD levels and IVF/ICSI outcomes. Methods: This multicenter, retrospective cohort study was conducted at IVFMD, My Duc Hospital and IVFMDPN, My Duc Phu Nhuan Hospital, Ho Chi Minh City, Vietnam between November 2017 and July 2019. Vietnamese patients aged 18–40 years with serum vitD (25(OH)D) samples collected before starting controlled ovarian stimulation and undergoing embryo transfer were eligible. Patients were divided into four groups based on 25(OH)D levels: Results: Of 3779 patients recruited, 25(OH)D levels were Conclusions: In this analysis, serum vitD levels did not appear to be correlated with pregnancy outcomes in patients undergoing IVF/ICSI.

Objective: To compare the effectiveness of two methods in preventing ovarian hyperstimulation syndrome (OHSS) with cabergoline and coasting. Design: Systematic review and meta-analysis of randomized clinical trials (RCTs). Patients: Women were considered as have risk of OHSS undergoing fertility treatment. Interventions: Cabergoline, coasting. Result: There were included five RCT studies. The clinical pregnancy rate was no significantly difference between two groups (RR 1.22, 95% CI [0.86, 1.71]), implantation rate (RR 1.00, 95% CI [0.75, 1.32]), severe OHSS (RR 0.93, 95% CI [0.38, 2.31]), fertilization rate (SMD 0.70, 95% CI [-0.10, 1.50]), number of oocytes retrieved (SMD 0.80, 95% CI [0.30, 1.30]), number of embryo transfer (SMD-0.04, 95% CI [-0.24, 0.17]), E2 value on the day of HCG injection (SMD 0.21, 95% CI [-0.25, 0.68]), number of MII oocytes (SMD 0.71, 95% CI [0.32, 1.11]), abortion rate (RR 0.61, 95% CI [0.21, 1.83]), number of follicles > 17 mm on day of HCG (SMD -0.01, 95% CI [-0.26, 0.24]), number of follicles 15 - 17 mm on day of HCG (SMD -0.08, 95% CI [-0.33, 0.17]), number of follicles 10 - 14 mm on day of HCG (SMD -0.06, 95% CI [-0.31, 0.19]). Conclusion: Both cabergoline and coasting prevent the occurrence of OHSS, but no statistically significant difference between them. Compared with coasting group, a daily dose of 0.5 mg cabergoline significantly increased the number of oocytes retrieved, MII oocytes, and fertilization rate, but decreased the abortion rate.

Objective To compare pregnancy outcomes in patients undergoing artificial reproductive treatment (ART) and fresh embryo transfer (ET) who received twice-daily vaginal progesterone capsule (Cyclogest) alone verses twice daily vaginal progesterone capsule (Cyclogest) plus weekly intramuscular Hydroxyprogesterone Capronate (Proluton depot) for luteal phase support. Methods A retrospective cohort study that included 1162 patients who completed fresh ART/ET cycle from January 2015 to April 2018. Vaginal Cyclogest 400 mg twice daily was given to 985 patients following oocytes retrieval; whereas 177 patients received weekly intramuscular Proluton depot 250 mg in addition to twice-daily Cyclogest. The primary outcome was live birth rate. The secondary outcomes included biochemical pregnancy rate, clinical pregnancy, biochemical, and early and late pregnancy loss. Results There was no difference between the twice-daily vaginal progesterone and the addition of weekly intramuscular progesterone injections to the twice-daily vaginal progesterone regarding a positive pregnancy test (40.5% and 46.9%, respectively, p=0.112). There was no statistical difference in live birth rates between the groups (24% for group one, 26% for group two, p=0.582). Conclusions The administration of weekly intramuscular progesterone in addition to twice-daily vaginal progesterone capsule for luteal phase support post ART cycle does not result in higher live birth rate.

Background Thyroid autoimmunity (TAI) has been associated with adverse pregnancy outcomes. To settle with fertility problem, prescribing aspirin combined with prednisone (P+A) to women positive for antithyroid antibodies (ATA) is frequent in clinical practice, but the real effect remains controversial. Methods A multicenter, retrospective study was conducted in three reproductive centers from 2017 to 2020. We recruited 494 euthyroid infertile women positive for anti-thyroperoxidase and/or thyroglobulin antibodies (TPOAb and TgAb, respectively) with thyroid stimulating hormone (TSH) levels ranging 0.35-4.0mIU/L who were undergoing their first in vitro fertilization and embryo transfer (IVF-ET) cycle. Ultimately, 346 women were included of which 150 women were treated with prednisone (10mg/d) and aspirin (100mg/d), while the remaining 196 women were untreated (control group). Treatment started on the day of embryo transfer and continued until clinical pregnancy was determined. Results Clinical pregnancy rate (CPR) was 57.5% vs. 63.5% in the control and treated groups (P=0.414) for first fresh embryo transfer cycles and 57.8% vs. 61.8% for frozen-thawed embryo transfer cycles (P=0.606). Additionally, the live birth rate (LBR) at the fresh embryo transfer was 49.6% vs. 47.3% in the control and treated groups (P=0.762). Logistic regression revealed that P+A did not improve CPR or miscarriage rates (MR). Furthermore, we observed that low free triiodothyronine (FT3) was associated with high MR. Conclusions Utilizing an adjuvant treatment of P+A after the embryo transfer may be unnecessary in euthyroid women with TAI undergoing their first IVF-ET, regardless of embryo type (fresh or frozen).

BackgroundGonadotropin-releasing hormone (GnRH) analogues are commonly used in clinical practice to prevent premature luteinizing hormone (LH) surge during In-Vitro Fertilization/ Intra-Cytoplasmic Sperm Injection (IVF/ICSI) cycles. This review aimed to summarize the available evidence comparing the effects of conventional GnRH antagonist protocols, the most commonly used GnRH antagonist protocols, and GnRH agonist protocols on IVF/ICSI outcomes in women with polycystic ovary syndrome (PCOS). MethodsA comprehensive electronic search was carried out in Pubmed, Cochrane CENTRAL, Scopus, Web of Science, CINAHL, TRIP, ClinicalTrials.gov and ISRCTN registry from inception until 24 November 2020 without any language or date restrictions. In addition, reference lists of eligible studies and previous meta-analyses were hand-searched to identify relevant studies. Eligible randomized controlled trials were those designed to compare the effects of conventional GnRH antagonist protocols and GnRH agonist protocols on IVF/ICSI outcomes in PCOS subjects. The Cochrane ROB 2.0 tool was used to assess the risk of bias of each study, and the GRADE assessment was used to evaluate the overall quality of evidence. Data synthesis and analyses were done using Review Manager 5.3 with the assistance of Revman Web. A random-effects model was used for all meta-analysis. Dichotomous outcomes were reported as Relative Risk (RR) and continuous outcomes as Weighted Mean Difference (WMD), both with 95% CIs. 2 Resultswe included ten studies with 1214 randomized PCOS women. Using GnRH antagonist protocols led to a significantly shorter stimulation duration (WMD=-0.91; 95% CI: [-1.45 to-0.37] day, P=0.0009), lower gonadotropin consumption (WMD=-221.36; 95% CI: [-332.28 to-110.45] IU, P< 0.0001), lower E2 levels on hCG day (WMD=-259.21; 95% CI: [-485.81 to-32.60] pg/ml, P=0.02), thinner endometrial thickness on hCG day (WMD=-0.73; 95% CI: [-1.17 to-0.29] mm, P=0.001), lower number of retrieved oocytes (WMD=-1.82; 95% CI: [-3.48 to-0.15] oocytes, P=0.03), and lower OHSS rate (RR= 0.58; 95% CI: [0.44 to 0.77], P=0.0002). However, no significant differences in live birth rate, clinical pregnancy rate, ongoing pregnancy rate, multiple pregnancy rate, miscarriage rate and cycle cancellation rate were seen between the GnRH antagonist protocols and the long GnRH agonist one. Although more cycles were cancelled due to poor ovarian response in the GnRH antagonist protocol (RR= 4.63; 95% CI: [1.49 to 14.41], P=0.008), similar rates of cancellation due to risk of OHSS were noticed in both groups. DiscussionThe differences in IVF/ICSI outcomes may arise from the different patterns of gonadotropins suppression that the GnRH analogues exhibit during the early follicular phase of IVF/ICSI cycles and the divergent direct impacts of these analogues on ovaries and endometrial receptivity. The main evidence limitation was Imprecision. ConclusionsConventional GnRH antagonist protocols represent a safer and more cost-effective treatment choice for PCOS women undergoing IVF/ICSI cycles than the standard long GnRH agonist protocol without compromising the IVF/ICSI clinical outcomes.

In vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI).is associated with an increased risk of preterm (33rd - 37th gestational week), and early preterm birth (20th - 32nd gestational week). The underlying general and procedure related risk factors are not well understood so far. 4,328 infertile women undergoing IVF/ICSI were entered into this study. The study population was divided into three groups: a) early preterm birth group (n=66), b) preterm birth group (n=675 ) and c) full-term birth group (n=3653). Odds for preterm birth were calculated by stepwise multivariate logistic regression analysis. We identified seven independent risk factors for preterm birth and four independent risk factors for early preterm birth. Older (>39) or younger (

Background: Oral progestin has been used to prevent premature ovulation during follicle stimulation protocols performed in combination with a freeze-only strategy. However, no studies have determined how oral progestin clinically compares to gonadotropin-releasing hormone (GnRH) antagonists in women with normal ovulation. This study aimed to compare the efficacy and safety of controlled ovarian stimulation between progestin-primed ovarian stimulation (PPOS) protocol and GnRH antagonist (GnRH-ant) protocol.Methods: Young women with infertility and normal ovarian reserve who underwent in vitro fertilisation (IVF) treatments were screened and randomly allocated to the PPOS or GnRH-ant group. Women in the PPOS group underwent freeze-all and delayed embryo transfer, whilst fresh embryo transfer was preferred for those in the GnRH-ant group. The primary endpoint was the cumulative live birth rate (CLBR). Secondary endpoints included the incidence of premature luteinising hormone (LH) surge and the number of viable embryos.Results: CLBRs were similar in the PPOS and GnRH-ant group (55.75% vs. 52.87%, respectively, P > 0.05). No premature LH surge was observed during ovarian stimulation in the PPOS group, although six (3.45%) cases were observed in the GnRH-ant group. On the trigger day, LH level was lower in the PPOS group than in the GnRH-ant group (2.30 ± 1.78 mIU/ml vs. 3.66 ± 3.52 mIU/ml, P < 0.01). There were no differences in the number of retrieved oocytes, mature oocytes, or viable embryos between the two groups. Other clinical outcomes including implantation rates (37.27% vs. 36.77%), clinical pregnancy rates (55.75% vs. 55.89%), and miscarriage rates (12.28% vs. 13.76%) were comparable between the PPOS group and GnRH-ant group (P > 0.05). There was also no significant differences in newborn weights for singleton or twin births between the two groups (P > 0.05).Conclusion: Live birth outcomes are similar for PPOS and GnRH antagonist protocols in women with normal ovarian reserve. PPOS is likely to play a promising role in the freeze-only strategy given its simplicity and convenience for the patient. Trial registration: This trial was registered in the China Clinical Trial Registry on September 6, 2018 (number: ChiCTR1800018246).

Background: A number of publications have been studied the relation between blood group and female infertility including ovarian reserve, recurrent miscarriage and live birth, there is a lack of literature investigating joint wife/husband ABO blood type in large cohort. The aim of this study was to investigate the association between couple combinations for ABO blood groups and ART outcomes in patients undergoing IVF/ICSI.Methods: This retrospective cohort study included 30,717 couples underwent IVF cycles between 2010 and 2019. Clinical outcomes of IVF treatment was the primary outcome. History of spontaneous miscarriage, embryo quality, and birth sex, weights, defects rate were also studied. Multivariable logistic regression was used to examine the association. Odds ratios (OR) with 95% confidence intervals (CI) for each outcome were estimated.Results: There was no difference in the baseline demographics between the blood type groups. There was a statistically significantly positive association between the combination of female blood type AB and male blood type AB with biochemical pregnancy, clinical pregnancy, and live birth rate (OR 1.36; 95% CI, 1.05-1.78; P 0.02 and OR 1.31; 95% CI, 1.03-1.68; P 0.031 and OR 1.28; 95% CI, 1.01-1.63; P 0.041 respectively). No statistically significant difference was observed between joint wife/husband ABO blood types and high-quality embryo rate, early abortion rate, birth sex, birth weights and birth defects rate.Conclusions: Our findings suggest that husband/wife mating AB blood type may increase the success rate of IVF/ICSI cycles.

Objective Thyroid autoimmunity (TAI) per se has been demonstrated to be related to adverse pregnancy outcomes, including recurrent miscarriage and unexplained infertility. Although therapy of glucocorticoid to infertile remains controversial, aspirin combined with prednisone(P + A) are used frequently in antithyroid antibodies (ATA) positive infertile women in clinical practice. Methods A multicenter retrospective study was performed in the Second Affiliated Hospital of Zhejiang University School of Medicine, Ningbo Women and Children's Hospital and People's Hospital of Jinhua. We recruited 494 euthyroid infertile women positive for anti-thyroperoxidase antibody (TPOAb) and/or thyroglobulin antibody (TgAb) with thyroid stimulating hormone 0.35-4.0mIU/L, who undergoing first in vitro fertilization and embryo transfer (IVF-ET) cycle during the period of 2017 to 2020. 346 women were ultimately recruited, including 187 patients receiving fresh embryo transfer and 159 patients receiving frozen-thawed embryo transfer. Among them, infertile 150 women were treated with prednisone(10mg/d) and aspirin(100mg/d), while the other 196 infertile women were untreated as control group. The treatment commenced on the day of embryo transfer and continued until the clinical pregnancy was determined by ultrasonography around 30 days after embryo transfer. Results Clinical pregnancy rate (CPR) of the first fresh embryo transfer cycle was 57.5% vs. 63.5% in control vs. in treated group (P = 0.414) and CPR was 57.8% versus 61.8% in the two groups in the frozen-thawed embryo transfer cycle(P = 0.606). Additionally, the prevalence of delivering a live birth at the fresh embryo transfer was 49.6% in the non-treatment group and 47.3% in the P + A treated group(P = 0.762). Cumulative CPR was similar in the groups at the fresh embryo transfer (control vs treated group: 56.9% vs 61.0%, P = 0.534). By conducting logistic regression, combined treatment of P + A appeared to have no influence on CPR or miscarriage rates (MR) at the first embryo transfer. Furthermore, the low but within the reference level of free triiodothyronine (FT3) was seen as a poor predictor of spontaneous abortion at the fresh embryo transfer. Conclusions To conclude, with regard to euthyroid women with TAI undergoing the first IVF-ET cycle, adjuvant treatment of P + A after the embryo transfer may be unnecessary irrespective of the fresh or frozen-thawed embryo transfer.

Introduction: Embryo transfer into the uterus during IVF/ICSI treatment is followed by embryo implantation, which depends on the receptivity of endometrium and is a pre-requisite for the initiation of a successful pregnancy. The endometrium proliferates under the hormonal influence (estrogen and progesterone), which is reflected by its thickness which is measurable by transvaginal ultrasonic scanning (TVS). The thickness of the endometrium less than 8 mm has been linked with implantation failure.11 Endometrial thickness of 9-14 mm on the day of progesterone supplementation has been shown to have a higher implantation rate. Thus, the implantation potential of a good embryo remains low during IVF treatment despite ovarian stimulation methods of assisted fertilization and improved culture condition.Objectives: This study was planned to determine the impact of endometrial thickness on ET and pregnancy rate in Pakistani patients undergoing IVF/ ICSI.Methods: This was a prospective, cross-sectional study conducted at a private fertility clinic (Lahore institute of fertility and endocrinology, LIFE) in Lahore, Pakistan, from January 2015 to December 2015. In this study, all those cases were included that reached oocytes pickup and embryo transfer (ET). Two protocols, agonist and antagonist, were used. In all cases, human menopausal gonadotrophin (hMG) and follicle-stimulating hormone (FSH) were used for stimulation. Follicles development was monitored; at least 3 follicles were reached at the diameter of ≥16mm.Results: Thickness of endometrium on decision day was calculated at various ranges concerning both groups and find pregnancy rates. When endometrial thickness was ≤ 6mm, PR was 2.22%, 7mm pregnancy rate (PR) was 1.11%, 8 mm PR was 3.33%, 9mm PR was 17.78%, 10mm PR was 33.33%, 11mm PR was 32.22%, and more than 11 mm PR was 10%. So, PR was higher from 2.22% to 10%. The overall pregnancy rate was 45.45%.Conclusion: The study concluded that endometrial thickness has significant effects on embryo transfer and outcomes.

The aim of this study was to explore whether the presence of intra-cavitary fluid (ICF) influences the pregnancy outcomes of patients with caesarean section (CS) in embryo transfer cycles. A total of 8494 transferred cycles of 4924 women were enrolled in this retrospective study and separated into three subgroups by previous delivery method and the presence of intra-cavity fluid, a caesarean group with ICF (CS-ICF, n = 649), a caesarean group without ICF (CS-noICF, n = 3207), and the remaining 4638 cycles without ICF were included in the vaginal delivered group (VD, n = 4638). Baseline characteristics and clinical outcome were compared. Propensity score matching (PSM) was conducted to adjust confounding factors between groups. Patients in the CS-ICF group were of younger age (36.49 ± 4.19 vs 37.34 ± 4.25, 37.32 ± 4.86, P P P

Purpose To determine the utility of short gamete coincubation in in vitro fertilization (IVF‐S) combined with early rescue intracytoplasmic sperm injection (R‐ICSI) and split IVF‐ICSI in preventing low fertilization based on a retrospective cohort study. Methods Couples with a high risk of low IVF fertilization during the first ART cycle underwent IVF‐S with R‐ICSI or split IVF‐ICSI. Fertilization rate, embryo quality, and clinical outcomes were measured. Results After propensity score matching, we included 188 couples in the IVF‐S with R‐ICSI group as Group 1 and 720 in the split IVF‐ICSI group as Group 2. Normal fertilization rates were similar; however, Group 1 had a higher multiple pronuclei rate (10.42% vs. 4.50%, p

Purpose To compare assisted reproductive technology (ART) outcomes and preimplantation embryo development between underweight and normal weight women. Methods This retrospective cohort study included 26 underweight women (body mass index [BMI] 2) and 104 normal weight women (BMI > 20 and 2) who underwent a total of 204 in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles and 358 fresh/frozen embryo transfers (ET) in our institution between January 2016 and December 2018. Statistical analyses compared selected ART outcomes (ovarian stimulation, fertilization, and pregnancy) between both weight groups. Morphokinetic and morphological parameters were also compared between 346 and 1467 embryos of underweight and normal weight women, respectively. Results The mean ± standard deviation age of the underweight and normal weight women was similar (31.6 ± 4.17 vs 32.4 ± 3.59 years; p = 0.323). There were no differences in the peak estradiol levels, the number of retrieved oocytes, the number of metaphase II oocytes, and the oocyte maturity rates between the two groups. The IVF/ICSI fertilization rates and the number of embryos suitable for transfer or cryopreservation were similar for both groups. All morphokinetic parameters that were evaluated by means of time-lapse imaging as well as the morphological characteristics were comparable between low and normal BMI categories. There were no significant differences in pregnancy achievement, clinical pregnancy, live births, and miscarriage rates between the suboptimal and optimal weight women. Conclusion Underweight status has no adverse impacts on the outcomes of IVF/ICSI with either fresh or frozen ET or on preimplantation embryo development and quality.

Objective Recent studies revealed intriguing associations between cholecalciferol (D3) and reproductive functions. Seasonal changes of D3 concentrations are well known; however, they are not always considered in the context of reproductive functions. In this study, we analyzed D3 serum concentration in IVF/ICSI patients with respect to seasonal 3-month quartiles and anti-Muellerian hormone (AMH) referring to the impact on Assisted Reproductive Technologies (ART) outcome. Materials and research methods We studied 469 female patients, presenting between 2012 and 2018 for ART treatment in our fertility center. D3 as well as the AMH serum concentrations were measured at the beginning of the follicle stimulation (days 3–5 of menstrual cycles). Results were evaluated with respect to seasonal quartiles and outcome of the ART cycles. Results D3 concentrations showed significant fluctuations within annual quartiles with a pronounced peak in August–October and a minimum in February–April (26.0 vs. 20.5 mg/dl; p p = 0.010) and these were associated with significantly shorter stimulation periods during August–October (11.29 vs. 12.12 days; p = 0.042), higher number of fertilized oocytes between August and October (6.23 vs. 4.97; p = 0.05) along with a trend towards higher numbers of cumulus–oocyte complexes. However, no such differences were found for the numbers of MII oocytes or pregnancy rates. Conclusion Our data indicate seasonal 3-month quartile variations of AMH concentrations and characteristics of ART, such as days of ovarian stimulation and number of fertilized oocytes. Highest AMH concentrations were found between August and October and this quartile was associated with highest D3 concentrations.

Study question Do Gonadotropin-releasing hormone agonists (GnRHa) triggering improves oocyte maturation, clinical outcomes, and safety compared to human chorionic gonadotropin (hCG) triggering during controlled ovarian stimulation with an antagonist protocol? Summary answer The final triggering using GnRHa allows a higher number of retrieved and mature oocytes to be obtained with comparable clinical outcomes and lower OHSS risk. What is known already GnRHa represent an alternative to hCG for ovulation triggering after controlled ovarian stimulation with an antagonist protocol for IVF/ICSI. GnRHa triggering is thought to be more physiological due to the endogenous surges in LH and FSH levels. However, the benefit of GnRHa over hCG triggering on oocyte maturation remains controversial. Study design, size, duration A systematic review and meta-analysis of randomised controlled clinical trials. Searches were conducted from 01 January 1990 to 15 April 2020 on MEDLINE, EMBASE, the Cochrane Library, ClinicalTrials.gov and EudraCT, using the following search terms: ‘GnRH agonist’, ‘hCG’, ‘triggering’. Two independent reviewers carried out the study selection, the bias assessment using the RoB2 tool, and the data extraction according to Cochrane methods. Participants/materials, setting, methods The primary outcomes were the total number of retrieved oocytes and the number of mature oocytes. The main secondary outcomes were the number of embryos obtained, the clinical pregnancy rate, the early pregnancy loss rate, the live birth rate, and the incidence of ovarian hyperstimulation syndrome (OHSS). Random-effects meta-analysis was performed followed by prespecified sensitivity and subgroup analyses. Main results and the role of chance A total of 29 randomised controlled trials were included. The mean number of retrieved oocytes [difference in means (95% CI) 0.99 (0.21, 1.78); p = 0.01; n = 26] and of mature oocytes [0.68 (0.04, 1.33); p = 0.04; n = 12] were statistically significantly higher after GnRHa than after hCG triggering. A similar difference was observed for the number of embryos [0.94 (0.19, 1.68); p = 0.01; n = 10]. No differences in the clinical pregnancy rate [risk ratio 1.01 (0.90, 1.14); p = 0.83; n = 23], early pregnancy loss [1.27 (0.94, 1.71); p = 0.13; n = 16], and live birth rate [1.00 (0.77, 1.29); p = 0.97; n = 6] were noted. GnRHa was associated with a lower incidence of OHSS [odds ratio 0.25 (0.08, 0.74); p = 0.012; n = 20]. Limitations, reasons for caution The validity of meta-analysis results depends mainly on the quality and the number of the published studies available. Wider implications of the findings: In light of its safety and effectiveness, GnRHa should be the new standard for triggering in antagonist cycles, with dual triggering with hCG when the risk of OHSS is low and a fresh embryo transfer approach is used. Trial registration number NA

Study question Is there any association between stage of embryo at transfer based on extended in vitro culture and gestational trophoblastic disease risk during assisted reproduction? Summary answer No significant association between stages of embryo transfer from zygote stage to blastocyst stage was found after analysing 540376 cycles of IVF and ICSI. What is known already Gestational trophoblastic disease (GTD), commonly referred to as molar pregnancy, results from abnormal proliferation of the trophoblast with a reported incidence of ∼1 in 700 in the UK. Despite technological advances such as ICSI, PGT and selection of normally fertilised (2PN) embryos, there are reported cases of GTD following assisted reproduction. Blastocyst transfer is associated with higher pregnancy and live birth rates but evidence is lacking whether extended embryo culture to blastocyst stage influences implantation of an abnormal embryo or abnormal trophoblastic proliferation leading to GTD. Study design, size, duration A retrospective study was carried out using Human Fertilisation and Embryology Authority (HFEA) anonymised register data from 1999 to 2016. HFEA holds the longest running register for fertility treatment data in the world and is the national database for fertility treatment data in UK. A total of 540376 fresh IVF or ICSI assisted reproduction cycles were analysed. Participants/materials, setting, methods There were 1033588 treatment cycles during the study period but only 540376 cycles met the inclusion criteria of fresh IVF or ICSI. Cycles with incomplete data, frozen embryo transfers, donor treatment or surrogacy were excluded. A subgroup analysis of those with primary subfertility was performed after excluding subjects with secondary infertility in order to exclude an effect of a previous molar pregnancy. Multivariate regression analysis was used to adjust for possible confounders. Main results and the role of chance 78 molar pregnancies were reported in the original sample giving a prevalence of 4/10000 live births (78/228461), much lower than the prevalence given with natural pregnancies. Prevalence of molar pregnancy amongst the study population after meeting exclusion criteria was 4 /10000 livebirths (53/156683). Incidence of molar pregnancy was not statistically different between treatment type (0.0001 vs 0.00009). Significantly higher incidence of GTD was seen in the 40 to 42 age category compared to 18-34 category(OR 1.86(95% CI 8.7-3.75)),in par with known higher GTD risk in women above 40 in the general population. Of interesting note, although the incidence of molar pregnancy was significantly lower in women undergoing assisted reproduction increased risk with advancing age is not totally eliminated with treatment. There was no significant association between the occurrence of molar pregnancy with the type and cause for infertility and number of embryos transferred. Crude (1.06 (95% CI 0.852-1.31)) and adjusted odds ratios (1.07 (95% CI (0.857-1.32)) did not show any association between day of embryo transfer and molar pregnancy even after adjusting for age and excluding secondary infertility. Selection of blastocyst stage embryo after extended culture did not alter the likelihood of having a GTD compared to cleavage stage embryo. Limitations, reasons for caution The retrospective analysis of anonymised HFEA data limited adjustments for confounders such as smoking, previous history of GTD, ethnicity etc that predispose to GTD. Caution needs to be exercised for under-reporting of GTD to HFEA and lack of information on type of GTD identified. Wider implications of the findings Though GTD cannot be prevented by IVF/ICSI, the incidence is significantly low and extended culture is not associated with higher risk of abnormal trophoblastic proliferation or GTD occurrence with IVF/ ICSI treatment. These findings would aid informed implications counselling and reassurance of patients during assisted reproduction treatments. Trial registration number not applicable

Study question Is the incidence of miscarriage rates increased in frozen embryo transfer (ET) compared with fresh ET? Summary answer The frozen ET is not associated with an increased risk of miscarriage in comparison with fresh ET. What is known already Nowadays, the freeze-all policy has gained great popularity because it minimizes OHSS and it seems to improve the reproductive outcomes. This increase was possible thanks to progress in vitrification procedures. Nevertheless, some recent evidence suggests more caution on the wide use of elective frozen ET in a general IVF population for its possible effect on some pregnancy-related complications. Endometrial preparation schemes for frozen ET include natural cycles or, more frequently, artificial cycles in which the endometrium is prepared with exogenous steroids. In this last case, the corpus luteum is absent, but its demise could be correlated with some pregnancy-related complications. Study design, size, duration A meta-analysis, based on PubMed, Cochrane CENTRAL, EMBASE, was conducted to estimate and compare the miscarriage rate (MR) in frozen versus fresh ET. Following PICOS, inclusion criteria were: Population, patients undergoing IVF/ICSI; Intervention, frozen ET; Comparison, fresh ET; Outcome, MR (primary one); clinical pregnancy rate (CPR), ongoing pregnancy rate (OPR) and live birth rates (LBR) (secondary ones). Study design: RCT. Participants/materials, setting, methods Electronic and manual search, conducted from 1990 to 2020, yielded 269 studies. Two researchers reviewed the studies independently, excluding 247 and 14 studies after the first and second screening. The outcome data from the 8 included studies were combined using a Mantel–Haenszel model and applying the random effects models. The dichotomous data results of each study were expressed as risk ratio (RR) with 95% confidence intervals (CI). Heterogeneity was evaluated using the I2 statistic. Main results and the role of chance The eight studies comprise 6,934 participants, 3,450 undergoing frozen ET and 3,484 fresh ET (Ferraretti et al., 1999; Shapiro et al., 2011a; Shapiro et al.; 2011b; Chen et al., 2016; Shi et al., 2018; Le et al., 2018; Wei et al., 2019; Stormlund et al., 2020). Preliminary analyses excluded interaction between covariates. RR of miscarriage ranged from 0.29 (Chen et al., 2016) to 3.24 (Ferraretti et al., 1999). The level of heterogeneity (I2) was 48% and it was considered as moderate. MR resulted not significantly different in frozen [279/1,601 pregnancies] compared with fresh ET [308/1,458 pregnancies] (RR = 0.727, 95%CI=0.43–1.25, p = 0.25). By analysing the 5 trials in which the endometrium was prepared with steroids (Ferraretti et al., 1999; Shapiro et al., 2011a; Shapiro et al., 2011b; Chen et al., 2016; Le et al., 2018), MR resulted not significantly different in frozen [126/720] versus fresh ET [160/682] (RR = 0.605, 95%CI 0.22–1.66; p = 0.33; I2=43%). For the secondary outcomes, CPR (RR = 1.14; 95%CI=0.97–1.35; 0.56 vs 0.52 for frozen and fresh ET, respectively), OPR (RR = 1.12; 95%CI=0.95–1.33; 0.49 vs 0.46 for frozen and fresh ET, respectively) and LBR (RR = 0.93; 95%CI=0.60–1.44; 0.49 vs 0.47 for frozen and fresh ET, respectively) were not significantly different between the two groups. Limitations, reasons for caution The MR was not the primary outcome of the included RCTs. This could have determined a sample size not calibrated for the analysis of the primary outcome of the present meta-analysis. Wider implications of the findings: This meta-analysis indicates that MR was similar between frozen and fresh ET. The endometrial preparation, in artificial cycles without corpus luteum, does not seem to influence the good course of pregnancy. Based on these results, the choice of a fresh ET should not derive by the presumed reduction of MR. Trial registration number Not applicable

Study question The purpose of this systematic review is to calculate dropout-rates of IVF/ICSI treatment by analysing the published cumulative live birth rates of IVF/ICSI treatment. Summary answer One out of three patients stop their treatment after their first IVF/ICSI cycle and dropout-rates tend to increase per consecutive cycle. What is known already Cumulative live birth rates (CLBRs) have created the possibility to present realistic probabilities of having a live birth after IVF/ICSI treatment. However, it is noted that a significant percentage of the patients stop their treatment before having a child (“dropout”). Possible reasons and predicting factors for dropout of treatment are already extensively investigated. However, only a few studies try to report about the incidence of dropout. Publications on CLBRs of large numbers of patients allow the extraction of dropout-rates. These rates will provide insight in the extent of the problem and could be used as a reference for interventional studies. Study design, size, duration Four databases (PubMed, The Cochrane Library, EMBASE, DoKS) were systematically searched from 1992 to December 2020. Search terms referred to “cumulative live birth” AND “ART/IVF/ICSI”. No restrictions were made on the type or language of publication. Studies were included if they reported absolute numbers of patients and live births per consecutive complete IVF/ICSI cycle or per consecutive embryo transfer cycle, starting from the first IVF/ICSI cycle for each patient. Participants/materials, setting, methods Dropout-rates per cycle were calculated in two manners: “intrinsic dropout-rate” with all patients that started the particular IVF/ICSI cycle in the denominator, and “potential dropout-rate” with all patients who did not achieve a live birth after IVF/ICSI (and potentially could have started a consecutive cycle) in the denominator. Dropout-rates were analysed for consecutive complete cycles and consecutive embryo transfer cycles, because these two manners are used in reporting CLBRs, often related to the reimbursement policy. Main results and the role of chance This review included 29 studies and almost 800,000 patients from different countries and registries. Regarding the patients who started their first IVF/ICSI cycle, trying to conceive their first child by IVF/ICSI, intrinsic dropout-rate was 33% (weighted average) after the first complete cycle, meaning they did not return for their second oocyte retrieval cycle. After the first embryo transfer cycle, intrinsic dropout-rate was 27% (weighted average), meaning those patients did not return for their next frozen-thawed embryo transfer cycle or for the next oocyte retrieval cycle. Regarding the patients who did not achieve a live birth after the first complete cycle, potential dropout-rate was 48% (weighted average), and 37% (weighted average) after the first embryo transfer cycle. Both potential and intrinsic dropout-rates for both consecutive complete and embryo transfer cycles tended to increase with cycle number. One study on second IVF/ICSI conceived children showed a potential dropout-rate after the first complete cycle of 29%. From studies on women >40 years of age, the potential dropout-rate after the first complete cycle was 45% (weighted average) and from studies with the uses of testicular sperm extraction, the potential dropout-rate after the first complete cycle was 34% (weighted average). Limitations, reasons for caution Our analysis was hampered by the different ways of reporting on CLBRs (complete cycles versus embryo transfer cycles), informative censoring, patients changing clinics and spontaneous pregnancies. Dropout-rates were potentially overestimated given that spontaneous pregnancies were not taken into account. Wider implications of the findings The extent of dropout in IVF/ICSI treatment is substantial and has an important impact on its effectiveness. Therefore, it is a challenge for fertility centers to try to keep patients longer on board, by taking into account the patients’ preferences and managing their expectations. Trial registration number PROSPERO Registration number: CRD42020223512

Study question Can frozen embryo transfer (FET) be offered immediately after a stimulated IVF/ICSI cycle without compromising live birth rate (LBR)? Summary answer FET in the menstrual cycle immediately following the stimulated IVF/ICSI cycle was associated with a slightly higher LBR compared to standard postponed FET. What is known already It is standard clinical practice to postpone FET for at least one menstrual cycle following a failed fresh transfer or a freeze-all cycle. This practice is thought to minimize any possible residual negative effect of ovarian stimulation, with excessive steroid levels and multiple corpora lutea, on the resumption of a normal ovulatory cycle and receptivity of the endometrium. Even so, elective deferral of FET is an empirical strategy based on suggestions rather than solid scientific evidence and may unnecessarily delay time to pregnancy, causing frustration and decreased quality of life to couples. Study design, size, duration Systematic review and meta-analysis according to PRISMA guidelines. Original studies on subfertile women aged 18-46 with any indication for treatment with IVF/ICSI investigating the timing of FET after IVF/ICSI were included. Intervention was defined as FET in the menstrual cycle immediately following the stimulated IVF/ICSI cycle. Comparator was defined as FET in the second or subsequent menstrual cycle following IVF/ICSI. Risk of bias was assessed using Robins-I and quality of evidence using GRADE. Participants/materials, setting, methods PubMed (MEDLINE) and EMBASE databases were searched for MeSH and Emtree terms, as well as text words related to timing of FET, up to March 2020. There were no limitations regarding year of publication or duration of follow-up but to English language. The primary outcome was LBR. Secondary outcomes were implantation rate, pregnancy rate, clinical pregnancy rate (CPR), time-to-pregnancy, miscarriage rate (MR), cycle cancellation rate and patient wellbeing. Main results and the role of chance Out of 4124 search results, 15 studies were included in the review. Studies reporting adjusted odds ratios (aOR) for LBR, CPR and MR were included in meta-analyses. All studies (n = 15) were retrospective cohort studies involving a total of 6,304 immediate FET cycles and 13,851 postponed FET cycles including 8,019 matched controls. Twelve studies of very low to moderate quality reported no difference in LBR with immediate versus postponed FET. Two studies of moderate quality reported a statistically significant increase in LBR with immediate FET and one small study of very low quality reported better LBR with postponed FET. Trends in rates of secondary outcomes followed trends in LBR regarding timing of FET. The meta-analyses showed a significant advantage of immediate FET (n = 2,076) compared to postponed FET (n = 3,833), with a pooled aOR of 1.20 (95% CI 1.01-1.44) for LBR and a pooled aOR of 1.22 (95% CI 1.07-1.39) for CPR. Limitations, reasons for caution Limitations include the retrospective design and heterogeneity of studies included, limiting comparison and pooling of data. With little transparency regarding cancellation rates, the risk of selection bias is apparent. Further, confounding by embryo quality is a limitation. Small sample sizes are a limitation to subgroup meta-analyses. Wider implications of the findings The standard clinical practice of postponing FET for at least one menstrual cycle following a failed fresh transfer or a freeze-all cycle may not be best clinical practice. Randomized controlled trials including data on cancellation rates are highly needed to provide high grade evidence regarding clinical practice and patient counseling. Trial registration number not applicable

Study question Does time between ejaculation and processing, and time between processing and insemination/injection affect fertilization rate (FR) and ongoing pregnancy rate (OPR) in IVF/ICSI treatments? Summary answer Increasing time between processing and insemination significantly decreased the OPR after IVF. FRs after IVF/ICSI and OPR after ICSI were not affected by different time-intervals. What is known already The choice for IVF or ICSI depends on semen quality, however, this doesn’t affect the outcome of IVF/ICSI treatments (Mariappen et al 2018). After ejaculation, the percentage of motile spermatozoa decreases progressively at a rate of about 10%/hour (Makler 1979). According to the ESHRE-guideline, semen should be processed within 1 hour after ejaculation. In our laboratory, a validation was performed that confirmed a decrease in sperm motility after ejaculation. During incubation at 37 °C after processing, the sample remained stable in incubation medium (unpublished data). Therefore, we analyzed the effect of handling time and incubation time with regard to IVF/ICSI outcomes. Study design, size, duration This retrospective data analysis examines the effect of time between ejaculation and processing using density-gradient centrifugation (handling time) and time between processing and insemination (IVF)/injection (ICSI) (incubation time) on the FR and OPR, irrespective of the initial semen quality. A total of 1488 oocyte pickups (844 IVF, 644 ICSI) were included from 1060 patients undergoing fertility treatment between 2017 and 2019. Oocyte pickups without oocytes, with oocyte vitrification, or with donor oocytes were excluded. Participants/materials, setting, methods Anonymized data were obtained from the laboratory database ProMISe. Handling time and incubation time of the semen incubated at 37 °C and 5% CO2 were analyzed in relation to the occurrence of TFF (Total Fertilization Failure), FR and OPR. Linear and logistic regression was performed in SPSS version 25. In case of significant association, the data were adjusted for potential confounders, such as woman’s age, semen quality before and after preparation, and number of oocytes. Main results and the role of chance This study shows that increasing the incubation time of the semen significantly reduced the OPR per ET in IVF treatments (from 30,8% within 3,5 hours to 24,1% after 6 hours) even after adjusting for the potential confouders. However, the OPR in ICSI treatments was not significantly affected by the incubation time (rather, there was an opposite trend). Also, the handling time of the semen did not significant effect the FR per OPU and the OPR per ET in IVF/ICSI treatments. The overall percentage of TFF was 3,5% and did not differ significantly between the IVF and ICSI treatments. Both handling time and incubation time did not have a significant effect on the occurrence of TFF. An explanation for the decrease in OPR in IVF treatments may be that increasing the incubation time at 37 °C reduces the sperm quality as the capacitation reaction takes place too early, energy levels are reduced, DNA damage increases, or vacuoles arise in the sperm heads (Thijssen et al 2014, Jackson et al 2010, Peer et al 2007). Incubation at room temperature and reduction of the insemination time may improve OPR. Limitations, reasons for caution Retrospective study limitations (bias), no data on DNA fragmentation, incubation of semen only at 37 °C after preparation. Wider implications of the findings: Although it is recommended to produce semen at the IVF-department, our results show that an exception can be made, when production of a semen sample in a clinical setting is stressful, with no negative effect on the outcome. Furthermore, incubation-time at room temperature may have a positive effect on OPR. Trial registration number Not applicable

Study question What is the influence of time on the vaginal microbiome-based prediction of IVF/ICSI success? Summary answer Time influences the vaginal microbiome-based prediction of IVF/ICSI success. What is known already The association between the microbiome of the lower female reproductive tract and subfertility is discussed extensively suggesting its importance for fertility and fertility treatment. Using a modified next generation sequencing technique, an assay of the vaginal microbiome that predicts the pregnancy chances before starting the IVF/ICSI procedure has been developed and validated (1) displaying profiles associated with a low, medium and high chance of implantation. The vaginal microbiome is already known to change over time (2). However, it remains unclear to what extent spontaneous improvement from a low score can occur and over what time period. Study design, size, duration To investigate the spontaneous reversal capacity and associated time period of a low score microbiome profile in IVF-ICSI patients, an observational prospective cohort study of 77 women was performed using the ReceptIVFity assay. Women with medium or high profiles were encouraged to proceed with their ART treatment, whereas women with a low profile were suggested to delay the treatment for 1 month until a subsequent swab was taken with a maximum of 4 repeats. Participants/materials, setting, methods The study was carried out in a University based single center setting. Ethical approval was obtained (6259R MPG§23b). Patients between 24 and 41 years of age were included when eligible for their first, second or third IVF or IVF-ICSI attempt. Exclusion criteria were: antibiotic treatment in the 3 months prior to the test, women who have started with hormone treatment in the last 2 months in the context of ovarian stimulation, or downregulation of endometriosis. Main results and the role of chance Of the 77 patients included, 53 had a high or medium profile and proceeded with their treatment. 24 had a low profile and were supposed to delay the treatment in favor of a subsequent test. The low profile patients were followed up as indicated in the study description. Unfortunately, 11 of the 24 low score patients dropped out of the study. This relatively high number can only in parts be explained by unswayable medical reasons as no fertilization or embryo arrest but a comparable number of patients dropped out most likely due to Corona restrictions or Corona-related anxiety reasons. In the low score group, 1 month after the initial test, 12 patients repeated the swab; 4 remained low (33,33%), whereas 8 shifted to the medium or high (66,67%) groups. After 2 months, 4 patients had another test; 1 remained low (25%), 3 shifted to medium and high (75%). Therewith, in two months’ time 91,7% shifted from low to a better (medium/high) profile. So far, only 1 patient of the initial lows remained low for 5 months. The 12 shifters had a clinical pregnancy rate of 40% after the first embryo transfer after changing the microbiome profile from low to medium/high. Limitations, reasons for caution The results described were generated from a smaller group than intended initially due to a relative high dropout rate for no medical reasons. Wider implications of the findings: Patients suffering from infertility have a clinical benefit from performing a ReceptIVFity test before ART treatment and to delay treatment, when the result is low, since the spontaneous conversion time to a better profile, and therewith a higher pregnancy chance, occurred within 2 month in almost all patients. Trial registration number 2018124928

Study question Does an elevated SDF (>15%) increase the odds of adverse maternal and neonatal outcomes in autologous oocyte IVF-ICSI cycles from unselected couples? Summary answer No adverse effects of high SDF on obstetric and neonatal outcomes have been found in couples with sperm fragmentation undergoing IVF-ICSI cycles with own eggs. What is known already Sperm chromatin integrity assessment has been implemented as an additional tool in the clinical evaluation of sperm quality in infertile patients undergoing an assisted reproduction treatment. All of the published reports to date appraise its effect on clinical outcomes, and how it impacts embryo quality and the pregnancy chances after IVF and ICSI cycles. Sperm DNA integrity has also been hypothesized to affect offspring health but not many studies have reported in humans if an elevated SDF raises the risks of obstetric, delivery and neonatal outcomes. Study design, size, duration Multicentric retrospective cohort study of all IVF-ICSI cycles using autologous oocytes between January 2000-March 2019 at Spain IVIRMA clinics of couples with a SDF test on their ejaculated semen. The sperm fragmentation index was measured in all men with TUNEL assay. The database included 228 couples which had a delivery with at least a newborn. Subjects were divided into two study groups according to their level of SDF: ≤15% (low SDF) or > 15% (high SDF). Participants/materials, setting, methods Patients with missed information on maternal and neonatal outcomes were not counted for the analysis. The obstetric outcomes were gestational age, gestational diabetes, preeclampsia (hypertension with proteinuria after 20 weeks of gestation) and type of delivery. Neonatal outcomes were sex, birth weight, length, head circumference, Apgar score at 1, 5, 10 minutes, and neonatal intensive care unit (NICU) admission. Student’s t-test and Fisher’s test were used for statistical analysis. A p-value Main results and the role of chance Maternal age mean was 37.4 years (95%CI 36.9-38.0) in ≤ 15%SDF group and 37.2 years (95%CI 36.1-38.4) in > 15%SDF group (p = 0.8). Similar gestational age was found, 41.8 weeks (95%CI 41.3-42.2) in ≤ 15%SDF and 41.3 weeks (95%CI 40.4-42.3) in > 15%SDF. Gestational diabetes incidence was higher in > 15%SDF compared to ≤ 15%SDF group (3.5% versus 1.7% (OR = 2.0 (95%CI 0.03-39.8), p = 0.5). Equally, the incidence of preeclampsia was 3.6% in patients with high SDF versus 1.7% in couples with low SDF, OR = 2.1 (95%CI 0.03-41.3), p = 0.5. Type of delivery frequency was in the ≤15%SDF group 61.9% vaginal and 38.1% cesarean, while in the >15%SDF group 62.1% vaginal and 37.9% cesarean (OR = 1.0 (95%CI 0.4-2.6), p = 1.0). The overall proportion of singleton pregnancies was 87.2% (95%CI 82.4-91.2) and twins 12.8% (95%CI 8.8-17.6). There were no statistically differences between groups in the rate of delivery of twins and in the sex ratio of the newborns. When comparing the newborns of ≤ 15%SDF with >15%SDF group, the average of weight was 3011.7g (95%CI 2912.2-3111.2) versus 2986.4g (95%CI 2753.1-3219.7), of length was 49.2cm (95%CI 48.3-50.0) versus 49.5cm (95%CI 49.2-49.9), of head circumference was 34.9cm (95%CI 34.6-35.2) versus 34.3cm (95%CI 33.4-35.2). No statistically differences were observed for Apgar punctuation and for NICU admission. Limitations, reasons for caution Due to the retrospective nature of the study we have missing data from the lack of follow-up of many patients after the confirmation of the ongoing pregnancy. Although pregnancies of couples with elevated SDF have a higher incidence of gestational diabetes and preeclampsia, the sample size evaluated is a limitation. Wider implications of the findings This is one of the first reports to evaluate the relationship between paternal DNA damage and obstetric risks and neonatal health in couples with high SDF who underwent IVF-ICSI in our centers. Despite SDF did not jeopardize the maternal and neonatal outcomes, more studies are needed to confirm this conclusion. Trial registration number NA

Study question Is reproductive success measured as CLBR per inseminated oocyte, per embryo transfer and per embryo transferred affected by paternal age in autologous IVF-ICSI cycles? Summary answer The number of embryo transfers and embryos transferred until live birth, but not the number of inseminated oocytes, were significantly different among the age groups. What is known already In recent years, there has been an increase of the average paternal age at which the first child is conceived. Therefore, there is a growing interest on the study of the impact of male age on the reproductive outcomes in assisted reproduction cycles (ART). Several studies have shown negative effects of advanced paternal age on semen parameters, embryo aneuploidy, miscarriage, male infertility,.... However, other studies have found no association between them. Hence, the impact of paternal age on reproductive outcomes still remains uncertain, leading to a need of more research on this topic, which this study tries to address. Study design, size, duration This retrospective observational multicentric cohort study has included autologous IVF-ICSI treatments (n = 6295) performed to couples with etiology of male infertility (non-normozoospermic) in Spain IVIRMA clinics between January 2008 and March 2020 using patients’ own sperm sample. Paternal age ranged from 20 to 75 years. The study population was categorized in 5 groups following the criterion of homogenizing the number of observations between groups: 20–34 (A), 34–37 (B), 37–39 (C), 39–42 (D) and 42–75 (E) years. Participants/materials, setting, methods Considering that male age could be a factor affecting reproductive outcomes, we evaluated men with different age that performed an autologous IVF-ICSI treatment with their own semen, etiology of male infertility and known age. Data was exported in order to obtain the clinical database and Kaplan-Meier was used for data analysis. P Main results and the role of chance This study considered approximately 2976 patients and 4385 embryo transfers. The CLBR per inseminated oocyte showed no significant difference between the study groups: A (6.43%, 48.24%, 81.38%), B (5.74%, 52.14%, 82.87%), C (6.14%, 49.83%, 83.69%), D (5.89%, 53.60%, 81.16%) and E (6.61%, 47, 52%, 77.85%) for 4, 13 and 21/22 inseminated oocytes, respectively. In terms of CLBR per embryo transfer, the results obtained for each of the age groups were: A (31.81%, 71.89%, 87.63%), B (28.89%, 67.87%, 82.63%), C (27.10%, 68.87%, 88.17%), D (23.45%, 64.63%, 100.00%) and E (22.88%, 55.48%, 63.31%) for 1, 4 and 7 embryo transfers, respectively. There were statistically significant differences in the CLBR per embryo transfer between the studied age groups (p Limitations, reasons for caution The retrospective nature of this study leads to biases derived from the clinical practice and to the presence of missing data (limiting sample size). Moreover, this study considered autologous cycles, therefore the results were not adjusted for female factors. Wider implications of the findings: Our study showed that reproductive outcomes measured by CLBR per embryo transfer and embryo transferred until live birth were significantly different between the paternal age groups in autologous IVF-ICSI cycles of infertile males. Hence, paternal age could be affecting reproductive outcomes and it should be considered for improving infertility counselling. Trial registration number NA

Study question Does GCSF by intrauterine route leads to better result in the treatment of thin endometrium as compared to GCSF by the subcutaneous route, in IVF-ICSI Cycles? Summary answer Yes, GCSF by intrauterine route leads to better result in the treatment of thin endometrium as compared to subcutaneous-GCSF, in ART Cycles? What is known already GCSF, is a member of the colony stimulating factor family of cytokines and growth factors. GCSF receptors are expressed in high concentration on dominant follicle, particularly at preovulatory stage.The endometrium also shows an increased expression of these receptors. GCSF concentration rises in the follicular fluid at the same time. Serum levels of GCSF are found to be in direct correlation with levels of GCSF in follicular fluid. Serum levels increase progressively from the day the embryo-transfer to the day of implantation. GCSF has been found to be beneficial in patients with thin endometrium and recurrent implantation failure. Study design, size, duration This was a RCT conducted between 2018–2019. 30 patients with thin endometrium were enrolled in each group. In either group, GCSF was given if endometrium was less than 7mm on day 14, maximum of two doses were administered. Patients undergoing frozen embryo transfer were recruited in the study, after meeting the inclusion and exclusion criteria. Primary outcome measured was increase in endometrium thickness and the secondary outcome was the clinical pregnancy rate and abortion-rate. Participants/materials, setting, methods 60 patients with thin endometrium were randomly divided into two groups. Group A: Inj. GCSF (300 mcg/1 ml) subcutaneously on Day 14 onwards alternate days for two doses. Group B: Inj. GCSF (300 mcg/1 ml) instilled slowly into the uterine cavity using an intrauterine insemination (IUI) catheter under USG guidance. Endometrial thickness was assessed after 48 h. If endometrial thickness was found to be Main results and the role of chance In the subcutaneous group (group-A) the mean endometrial thickness before GCSF injection was 5.8 ± 0.6 mm and, after injection it increased to 6.9 ± 0.4 mm. Similarly, in the intrauterine group (group-B) the mean endometrial thickness before GCSF was 5.9 ± 0.7 which increased to a mean of 7.9 ± 0.5 after GCSF instillation. The difference between endometrial thickness before and after intrauterine infusion of GCSF was more than that in the subcutaneous group. In group-A, 08 patients conceived out of 30 patients ( clinical pregnancy rate 26.6%) and in group B 11 conceived out of 30 patients in whom GCSF was instilled intrauterine (pregnancy rate 36.6%). Thus, there was a difference in the clinical pregnancy rate in the two groups, the intrauterine group yielding a higher clinical pregnancy rate, but it was not statistically significant. Because of the thin endometrium, we found an abortion rate of 25% (2/8) in the subcutaneous-GCSF group, and an abortion rate of 18% (2/11) in the intrauterine GCSF group. Limitations, reasons for caution There are few potential limitations because of the small sample size. Confounders such as obesity, smoking and alcohol intake, presence of adenomyosis and endometriosis, were not taken into consideration. Though prevalence of obesity is usually low in Indian women. Habits of smoking and alcohol are exceedingly uncommon in Indian women. Wider implications of the findings: Use of GCSF plays an important role in management of patients of thin endometrium undergoing embryo transfer. It is an easily available and economical preparation in developing countries and the intrauterine instillation of GCSF can be easily practiced in an ART unit with good results in resistant thin endometrium patients. Trial registration number Not applicable

Study question After one failed IVF/ICSI cycle, does the endometrial microbiota composition differ between women who will or will not reach a live birth within 12 months? Summary answer The endometrial microbiota composition did not significantly differ in women with one failed IVF/ICSI cycle with or without live birth, but statistical power was low. What is known already Evidence for the presence of an indigenous endometrial microbiome is mounting, and its composition may be associated with implantation success. However, a ‘core’ endometrial microbiome has not yet been defined, and its role in embryo implantation is still poorly understood. Further investigation of this topic may allow improvement and personalisation of clinical care for infertile couples. Endometrial microbiome analysis in infertile women has not yet been performed using transcervically obtained endometrial tissue. Using endometrial tissue instead of swabs or fluid may increase the bacterial DNA yield and therefore the precision of microbiome analyses. Study design, size, duration Endometrial tissue was obtained from a cohort of 141 infertile women undergoing endometrial scratching within a randomised controlled trial (RCT) (SCRaTCH trial, NL5193/NTR5342). Briefly, women aged 18-44 years with failed implantation after one full IVF/ICSI cycle and planning a subsequent IVF/ICSI cycle, were eligible. Participants were followed-up until 12 months after randomisation, with the primary outcome being live birth, defined as the delivery of at least one live foetus after 24 weeks of gestation. Participants/materials, setting, methods Endometrial tissue was obtained with an endometrial biopsy catheter in the midluteal phase of a natural cycle preceding subsequent IVF/ICSI, snap-frozen and stored at -80 °C until use. Total DNA was isolated from these biopsies, followed by 16S rRNA sequencing (V3-V4 region) to determine the endometrial microbiota composition. Positive (mock communities) and negative controls (DNA extraction and PCRs) were included. QIIME2 and DADA2 were used for the data analysis, followed by statistical analysis in R studio. Main results and the role of chance During the 12-month follow-up, 61/141 women (43.3%) reached a live birth. While endometrial microbiota profiles of all 141 women were analysed, only samples with ≥100 reads were included in the analysis, resulting in a total of 46 samples (32.6%) that were included in the analysis, which consisted of samples from 25 women who did not have and 21 women who did have a live birth within 12 months. The median number of reads per sample was not significantly different between the two groups (respectively 2,317 (IQR 651-19,031) and 1,335 (IQR 296-3,180), p = 0.29 by Mann-Whitney test). The endometrial microbiota detected, were bacterial genera frequently reported within the vaginal microbiota (e.g. Lactobacillus, Atopobium and Gardnerella). A clear dominance of Lactobacillus (relative abundance 55-100%, n = 22) or an unclassified bacterium genus (relative abundance 52-76%, n = 18) was observed in the majority of the samples; however, this dominance was not associated with the outcome of live birth. In addition, the samples dominated by Lactobacillus genera were mostly dominated by one species of Lactobacillus each (L. crispatus, L. iners, L. gasseri or L. jensenii). Limitations, reasons for caution The low biomass and the low ratio of bacterial versus human DNA in endometrial tissue were limiting factors in endometrial microbiota analysis. Furthermore, tissue was obtained transcervically, and contamination with vaginal/cervical microbiota could therefore have occurred. In the SCRaTCH trial no vaginal swabs were taken to serve as internal controls. Wider implications of the findings Future endometrial microbiota studies should consider the use of samples with a lower proportion of human DNA to maximize bacterial DNA yield. Furthermore, for endometrial microbiota research, sampling devices avoiding cervicovaginal contamination are desirable and may be developed in the future. Trial registration number SCRaTCH trial, NL5193/NTR5342

ObjectiveTo study the influence of endometriosis activity on the pregnancy outcomes of patients with recurrent implantation failure (RIF) in in-vitro fertilization/intra-cytoplasmic sperm injection (IVF/ICSI) cycles. The pregnancy outcomes were compared between RIF patients with endometriosis who received treatment at different occasions to explore the appropriate treatment plan for these patients and to optimize the pregnancy-support strategies.DesignAmbispective cohort study.MethodsA total of 330 patients with endometriosis were enrolled from 2008 to 2018 and included 1043 IVF/ICSI cycles. All patients were diagnosed with RIF after IVF/ICSI. Patients were assigned to three subtypes according to different control states of endometriosis, including the untreated, early-treatment, and late-treatment groups. The clinical pregnancy rate, live birth rate, and cumulative live birth rate of endometriosis patients with RIF were the main outcomes; additionally, the fertilization rate, available embryonic rate, and high-quality embryonic rate were also compared.ResultsThe early-treatment and late-treatment groups showed higher cumulative live birth rate than the untreated group (early-treated 43.6% vs. late-treated 46.3% vs. untreated 27.7%, Pvs. 85.20%, 76.50% vs. 64.47%). Kaplan–Meier curve showed that patients in the untreated group needed a mean of 23.126 months to achieve one live birth; whereas those in the treated group needed a comparatively shorter duration (early-treated: 18.479 ± 0.882 months and late-treated: 14.183 ± 1.102 months, respectively).ConclusionEndometriosis has a negative influence on IVF/ICSI outcome. The control of endometriosis activity can result in a higher cumulative live birth rate in patients. It is necessary for endometriosis patients to receive medical treatment to achieve a better prognosis especially for those with RIF.

Study question Does the endometrial synthesis and inactivation of 17β- estradiol differ between receptive and non-receptive endometrium in women undergoing IVF/ICSI? Summary answer The synthesis and inactivation of 17β-estradiol is similar in the endometrium of women who did and did not achieve a clinical pregnancy through IVF/ICSI. What is known already Implantation failure of high-quality embryos is a main concern in IVF/ICSI treatment. Blood sex-steroid concentrations do not reflect their corresponding concentrations in endometrial tissue. This is in line with the concept that blood steroids (and precursors) are locally converted to bioactive metabolites and vice versa, by expressing steroid-metabolising enzymes, such as 17β-hydroxy steroid dehydrogenase (17β-HSD). Studies indicate that alterations in intracrinology might modulate endometrial receptivity. We hypothesize that the local 17β-HSD activity during the window of implantation (WOI) differs between pregnant and non-pregnant IVF/ICSI patients. Study design, size, duration Case-control study of 40 patients that were recruited in the SCRaTCH study (NL5193/NTR5342), a randomised trial exploring whether ‘endometrial scratching’ in patients with a previous IVF/ICSI cycle failure affects pregnancy outcome in a subsequent IVF/ICSI cycle. For the present investigation, 20 endometrial biopsies from women who achieved clinical pregnancy after fresh embryo transfer (ET) were compared with 20 endometrial biopsies of women that did not conceive after fresh ET. Participants/materials, setting, methods Endometrial biopsies and serum were obtained at LH + 5-8 days (urinary test) in a natural cycle, prior to the fresh ET cycle. Cases (negative pregnancy test, n = 20) and controls (clinically pregnant, n = 20) were matched for primary vs. secondary infertility, embryo quality and age. Reduction of estrone to 17β-estradiol (synthesizing 17β-HSDs) and oxidation of 17β-estradiol to estrone (inactivating 17β-HSDs) were determined by high-performance liquid chromatography (HPLC). Results were compared with the Wilcoxon-Mann-Whitney Rank Sum Test. Main results and the role of chance Activity of 17β-HSDs responsible for the reduction of estrone to 17β-estradiol (mainly 17β-HSD type 1) was detected in all samples and ranged from 55 to 1864 pmol 17β-estradiol formed/mg protein/24 h. The values obtained from pregnant women (median: 1054) were not significantly different to those obtained from non-pregnant women (median: 997), p = 0.97. The activity of enzymes responsible for the oxidation of 17β-­ estradiol (mainly 17β-HSD type 2) into the less active estrone ranged from 32 to 1731 estrone formed/mg protein/24 h. The values obtained from pregnant women (median: 737) were not significantly different to those obtained from non-pregnant women (median: 624), p = 0.90. The ratio of 17β-HSD type 1:17β-HSD type 2 had a median of 1.63 in the pregnant woman compared to 1.95 in the group of non-pregnant woman (p = 0.57). Limitations, reasons for caution The study is pilot in nature and study population is small. Primary and secondary infertility patients were analysed together. A chance phenomenon could have occurred as included women were included after their first IVF/ICSI cycle, hence not every included study person met the criteria for repeated implantation failure (RIF). Wider implications of the findings 17β-estradiol metabolism takes place during the WOI, controlling the final 17β-estradiol level. Although the present investigation did not show differences between pregnant and non-pregnant women, it remains important to explore if estrogen balance deviations, e.g. in other cycle phases plays a role in clinical conditions such as primary infertility/RIF. Trial registration number NL5193/NTR5342

Study question Is the incidence of ectopic pregnancy (EP) increased in fresh compared with frozen embryo transfer (ET)? Summary answer The fresh ET is not associated with an increase of the incidence of EP in comparison with frozen ET. What is known already EP represents the first cause of mortality in the first trimester of pregnancy. Assisted reproductive technologies (ART) are associated with an increased EP risk. The reasons of this effect are inadequately explained and may be associated with variables patients-related as tubal diseases and ART-related as the number of embryos transferred, the depth of insertion of the catheter tip during ET and the supraphysiological estradiol levels during ovarian stimulation affecting endometrial receptivity and tubal function. The role of this last factor seems to be corroborated by higher incidence of EP in fresh versus frozen ET, as highlighted by some retrospective studies. Study design, size, duration A meta-analysis, based on PubMed, Cochrane CENTRAL, EMBASE, was conducted to estimate and compare the EP rate in fresh versus frozen-thawed ET. Following PICOS, inclusion criteria were: Population, patients undergoing IVF/ICSI; Intervention, fresh ET; Comparison, frozen/thawed ET; Outcome, EP (primary one), clinical/ongoing pregnancy and live birth rates (secondary ones). Study design: RCT. Participants/materials, setting, methods Electronic and manual search, conducted from 1990 to 2020, yielded 269 studies. Two researchers reviewed the studies independently, excluding 247 and 16 studies after the first and second screening. The outcome data from the 6 included studies were combined using a Mantel–Haenszel model and applying the random effects models. The dichotomous data results of each study were expressed as risk ratio (RR) with 95% confidence intervals (CI). Heterogeneity was evaluated using the I2 statistic. Main results and the role of chance The six studies included in the present review comprise in total 6,675 participants, 3,320 undergoing frozen ET and 3,355 fresh ET (Ferraretti et al., 1999; Chen et al., 2016; Le et al., 2018; Shi et al., 2018; Wei et al., 2019; Stormlund et al., 2020). Preliminary analyses excluded interaction between covariates, defining intervention/control groups and outcome. In particular, the incidence of tubal infertility was comparable between the two groups. Risk ratio of EP ranged from 0.03 (Ferraretti et al., 1999) to 2.77 (Shi et al., 2018). The level of heterogeneity (I2) between studies was 42% and it was considered as moderate. EP incidence resulted not significantly different in fresh ET [56/1,703 pregnancies] compared with frozen ET [44/1,799 pregnancies] (RR = 0.450, 95% CI 0.13–1.81, p = 0.29; I2=42%). Analysis of the secondary outcomes was conducted on 5 studies; results showed that clinical pregnancy rate was not significantly different between fresh (0.52) and frozen ET (0.56) (RR = 1.11; 95% CI = 0.93–1.34); at the same manner, the ongoing pregnancy rate (RR = 1.11; 95%CI=0.91–1.37; 0.46 vs 0.49 for fresh and frozen, respectively) and the live birth rate (RR = 0.93; 95%CI=0.60–1.44; 0.47 vs 0.49 for fresh and frozen, respectively) resulted not significantly different between the two groups. Limitations, reasons for caution The EP incidence was not the primary outcome of the included RCTs. This could have determined a sample size not calibrated for the analysis of the primary outcome of the present meta-analysis. Wider implications of the findings: This meta-analysis indicates that EP incidence is similar between fresh and frozen ET. The possible role on EP risk of the supraphysiological estradiol levels during ovarian stimulation should be reconsidered. Based on these results, the choice of a frozen ET should not derive by the presumed reduction of EP risk. Trial registration number Not applicable

Study question Is the outcome of IVF/ICSI cycles done with continuous (single step ) embryo culture system different from that with sequential embryo culture system ? Summary answer Yes the outcome of IVF / ICSI cycles done with continuous (single step ) embryo-culture system is better than that with sequential embryo-culture system . What is known already Embryo culture media are important factors in IVF, which can significantly influence the clinical outcome of IVF/ICSI cycles. However it is not clear which formulation is most optimal and whether sequential or continuous media (single step) should be favored. Sequential media complies with embryo demands based on developmental stage , taking into account metabolic changes embryos undergo in-vivo, while moving from the oviduct to the uterus. The embryos in the early cleavage stage prefer to use pyruvate to produce energy, whereas once development nears the blastocyst stage , the embryos start using glucose in the process of glycolysis . Study design, size, duration A prospective RCT was carried out at our centre between 2018–2019 and IVF-ICSI patients meeting inclusion criteria (at least six normal MII - Oocytes) were included in this study. The aim of study was to compare blastocyst formation rates after embryo-culture in two different culture media systems. 436 metaphase II Oocytes from 62 women were randomly and equally divided to be fertilized and cultured to the blastocyst stage in either sequential media or single-step media. Participants/materials, setting, methods In this prospective trial with sibling oocytes, 436 metaphase II oocytes from 62 women were randomly and equally divided to be fertilized and cultured to the blastocyst stage in either sequential media ( n = 218 MII oocytes) or a single medium ( n = 218 MII oocytes). In both groups, embryos were cultured in an interrupted fashion with media changes on day 3. Embryo transfer was performed on day 5. Main results and the role of chance Blastocyst formation rates on day 5 were significantly higher following culture in single step media 60.55% (132 / 218 ) as compared to sequential media 34.86% ( 76 / 218) . The percentage of good quality blastocysts was also significantly higher in single step media. In conclusion, culture in single step media was associated with higher blastocyst formation rates compared to sequential media , suggesting that the single medium may provide better support to the developing embryo. The proportion of poor quality embryos was significantly higher in the sequential media group. Results indicate that embryo culture in continuous media could be as efficient as embryo culture in sequential media. A significant difference observed was the proportion of poor quality embryos on day 5 , which was significantly higher when the embryos were cultured in sequential media. Our results suggest that the type of embryo culture media can influence the quality of embryos both at the cleavage stage and blastocyst stage. The use of continuous embryo culture media does not seem to cause an adverse effect; in fact, their use can lower the workload in busy IVF labs and lower the stress that embryos are exposed to during handling. Limitations, reasons for caution Although single-step-medium for extended culture has practical advantages and blastocyst formation rates appear to be higher, there is insufficient evidence to recommend either sequential or single-step media as being superior for the embryo-culture to days 5/6. Further studies comparing these two media systems in well-designed trials should be performed. Wider implications of the findings: When employing sequential media for embryo culture , it is necessary to transfer the embryos from one medium to another ( cleavage stage medium to blastocyst stage medium) which increases stress related embryo damage . Therefore, single-step media is beneficial as the embryos can develop undisturbed till blastocyst stage. Trial registration number Not applicable

Study question Are the ovarian reserve parameters and the outcome of ovarian stimulation for IVF / ICSI influenced by ethnicity? Summary answer Ethnicity influences ovarian reserve parameters and the outcome of ovarian stimulation for IVF / ICSI What is known already Infertility affects couples worldwide and due to a lack of a standardized reporting system, the real number, especially in developing countries, might be underestimated. The etiology of infertility may differ around the world and is often subjected not only to social, cultural and religious peculiarities, but also to different genetic influences. Published data suggest that ethnicity influences the ovarian reserve as well as the outcome of Assisted-Reproductive-Techniques (ART)-treatments. Key players of a successful ART outcome are the ovarian reserve and consequently the number of oocytes retrieved. Until today, the impact of ethnical differences is not sufficiently addressed in research. Study design, size, duration Prospective observational study, performed in 10 infertility centers worldwide (Europe (4 centers), Middle East North Africa (MENA) region (2 centers), Iran (2 centers), South America (1 center), India (1 center)) between May 2019 and September 2020, evaluating ovarian reserve and outcome parameters of ovarian stimulation treatments for IVF/ICSI. The study was approved by the Ethical Committee of each participating center. Participants/materials, setting, methods Couples with primary / secondary infertility and an indication for an IVF/ICSI treatment were included into this study. Besides anamnestic data regarding the history of the infertility and self-reported ethnicity (Arab, Caucasian, Hispanic, Ohters, Persian and South Asian), data obtained during the basic fertility assessment on the ovarian reserve parameters (Antral follicle count (AFC) and Anti-Muellerian-Hormone (AMH)) as well as stimulation parameters from the ovarian stimulation treatment were collected and analyzed. Main results and the role of chance This study comprised 1032 couples with the following distribution of the ethnicities: Arab 21.5%, Caucasian 15.9%, Hispanic 5%, Others 1.2%, Persian 33.4%, and South Asian 23%. The unadjusted means, SD and 95%CI (Confidence Interval) of AMH (ng/ml) for the groups were 3.33±0.19 [2.95–3.71]; 2.03±0.25 [1.55–2.52]; 2.43±0.74 [0.97–3.88]; 2.76±0.96 [0.88–4.64]; 3.10±0.16 [2.79–3.41]; 3.62±0.19 [3.25–3.98], for AFC 15.52±0.53 [14.49–16.55]; 12.00±0.67 [10.69–13.31]; 12.69±1.08 [10.57–14.81]; 15.11±2.60 [10.01–20.21]; 13.58±0.42 [12.75–14.41]; 13.49±0.51 [12.49–14.48] and for the number of retrieved oocytes (rCOC) 14.08±0.61 [12.88–15.27]; 9.84±0.71 [8.44–11.24]; 7.94±1.26 [5.48–10.41]; 9.92±2.62 [4.78–15.05]; 10.83±0.49 [9.87–11.79]; 17.06±0.59 [15.90–18.21], respectively. Univariate analysis of AMH, AFC and rCOC with the ethnicities revealed highly statistically significant differences for AMH and rCOC (p As age is a major confounder for the ovarian reserve, multivariate analyses were performed. After adjusting for age, AMH was significantly different between Arab-Persian, Arab-South Asian and Arab-Caucasian (p Limitations, reasons for caution Limitations of the study are an unequal number of included patients per ethnicity and that the data for the ovarian reserve parameters and the stimulation outcome were not available for all of the included patients. Wider implications of the findings: Counselling of couples with infertility have to take, besides all other factors, also the ethnicity of the couple into account as ethnicity influences the ovarian reserve parameters as well as the number of retrieved oocytes in ovarian stimulation cycles for IVF/ICSI. Trial registration number ClinicalTrials.gov Identifier: NCT03927417

Study question To investigate differences in In Vitro Fertilization (IVF)/Intracytoplasmic Sperm Injection (ICSI) outcomes between endometriosis women who do or don’t have a concomitant autoimmune disease. Summary answer Despite a higher oocyte yield, a trend for reduction in clinical pregnancy rates was observed in the autoimmunity group compared to women without concomitant autoimmunity. What is known already Endometriosis is an inflammatory chronic gynaecological disorder with a known detrimental impact on fertility. Endometriosis pathogenesis is still unclear. It has been postulated a role of both innate and adaptive immune system. The coexistence of endometriosis and autoimmunity is a well-documented occurrence Some recent findings have revealed an increased risk to have concomitant autoimmune disease in women with endometriosis, but no study has so far investigated whether this association could affect IVF/ICSI outcomes. Indeed, autoimmune phenomena, including proinflammatory cytokines and auto-antibody production, may result in diminished quality of oocytes/embryos with lower pregnancy rates among these patients. Study design, size, duration This was a retrospective observational study carried out at the Fertility Unit of IRCSS San Raffaele Hospital (Milan). We reviewed medical patients’ notes of women with a confirmed diagnosis of endometriosis who referred to our Fertility Unit from October 2018 to January 2021. Participants/materials, setting, methods Out of 1441 patients undergoing IVF/ICSI, 98 women had surgical/histopathological diagnosis of endometriosis. 25 of them had a clinical and/or serological diagnosis of autoimmunity. Autoimmunity was assessed by clinical data (blood tests for auto-antibodies or rheumatological records) obtained from the electronic patient files stored in the database of our Fertility Centre. Clinical pregnancy was defined as the presence of at least one intrauterine gestational sac with a viable embryo at week 6 after transfer. Main results and the role of chance 25/98 (25.5%) endometriosis women with a concomitant autoimmune disease (cases) were compared with 73/98 (74.5%) endometriosis patients without autoimmunity (controls). The mean age was 37.36±3.63 and 36.93±3.79 (p=.623) in cases and controls respectively. The mean number of oocytes retrieved was higher in cases (5.78±4.07) than in controls (3.82±2.69;p=.041); similarly, cases showed an higher number of embryos (2.13±1.93 vs. 1.19±1.37;p=.041) and blastocysts (1.89±2.02 vs. 0.85±1.61;p=.041) obtained. A total of 47 fresh embryo transfer (ET) were performed. Considering all the endometriosis patients, the clinical pregnancy rate (CPR) per cycle was 34.0% (16/47); when stratifying for the presence of autoimmunity the CPR was 23.1% (3/13) in cases, and 38.2% (13/34) in controls (p=.494). Limitations, reasons for caution This is a retrospective study based on data extraction from electronic records of our Fertility Centre. The sample size is limited and some information about past medical history could be missed. Results should be interpreted with caution until validated by future research providing more standardized data collection. Wider implications of the findings: Despite significantly higher numbers of oocytes retrieved and embryos/blastocysts formed, the presence of concomitant autoimmune disease in patients with endometriosis may impair pregnancy rates. Whether this finding is confirmed and whether it could be due to a defect in embryo/blastocysts quality or in endometrial receptivity deserves further studies. Trial registration number Not applicable