Embryo Morphology Evaluation Research Articles

Nomogram based on clinical and laboratory characteristics of euploid embryos using the data in PGT-A: a euploid-prediction model

BackgroundThe evaluation of embryo morphology may be inaccurate. A euploid prediction model is needed to provide the best and individualized counseling about embryo selection based on patients and embryo characteristics.MethodsOur objective was to develop a euploid-prediction model for evaluating blastocyst embryos, based on data from a large cohort of patients undergoing pre-implantation genetic testing for aneuploidy (PGT-A). This retrospective, single-center cohort study included data from 1610 blastocysts which were performed PGT-A with known genetic outcomes. The study population was divided into the training and validation cohorts in a 3:1 ratio. The performance of the euploid-prediction model was quantified using the area under the receiver operating characteristic (ROC) curve (AUC). In addition, a nomogram was drawn to provide quantitative and convenient tools in predicting euploid.ResultsWe developed a reliable euploid-prediction model and can directly assess the probability of euploid with the AUC (95%CI) of 0.859 (0.834,0.872) in the training cohort, and 0.852 (0.831,0.879) in the validation cohort, respectively. The euploid-prediction model showed sensitivities of 0.903 and specificities of 0.578.ConclusionsThe euploid-prediction model is a reliable prediction model and can directly assess the probability of euploid.

P–080 Higher sperm DNA fragmentation reduces the proportion of good quality embryos at day 5 on IVF and ICSI cycles from unselected males

Abstract Study question Does sperm DNA fragmentation (SDF) reduce the ratio of good-quality embryos in day 3 (D3) and day 5 (D5) of embryonic development? Summary answer High sperm DNA fragmentation (SDF >15%) is associated with poor embryo quality at blastocyst-stage per cycle in unselected patients undergoing IVF and ICSI. What is known already It has been shown that the proportion of spermatozoa with DNA fragmentation is higher in infertile men than in semen from fertile men. However, the controversy regarding the impact that sperm genome damage can have on IVF or ICSI treatments is evident in the published literature. The effects of SDF would become evident after activation of the embryonic genome at 8-cell stage, compromising not only the quality of the embryos obtained, but also the reproductive outcomes, as reduced implantation rates, higher miscarriages rates and thus, a decreased chance of pregnancy. Study design, size, duration This multicentric observational retrospective study included 1339 couples who underwent 2759 IVF-ICSI cycles using autologous oocytes from January 2000 to March 2019. All men have an SDF test in their ejaculated spermatozoa by TUNEL assay (Terminal deoxynucleotidyl transferase dUTP nick end labeling). The subjects were divided into two groups according to their sperm DNA integrity: low (≤15%) (n = 2287 cycles) or high (>15%) (n = 472) SDF. Participants/materials, setting, methods Embryo quality was assessed complying morphological standards at cleavage-stage on D3 and at blastocyst-stage on D5 (inner cell mass (ICM) and trophectoderm (TE) grade (A, B, C or D)) in according to ASEBIR’s embryo selection criteria, being embryos of good quality those categorized as A+B. The outcomes were calculated in relation to the total number of zygotes obtained. The results were compared by Student t test; p value <0.05 was considered significant. Main results and the role of chance The SDF average of the low group was 5.8% (95% CI 5.6–5.9) whereas in high group was 23.7% (95% CI 23.0–24.4). The female age was equal, 37.1 years (95%CI 37.0–37.2) and 37.1 years (95% CI 36.8–37.4) respectively. A total of 9796 embryos were evaluated. The optimal cleavage-stage embryo rate per cycle was 25.0% (95% CI 21.7–28.3) (8.0 average cells number, 1.5 embryo fragmentation average, symmetry 1, mononucleated cells) versus 26.7% (95%CI 19.1–34.2) (7.9 average cells number, 1.8 embryo fragmentation average, symmetry 1, mononucleated cells) when comparing between groups (p < 0.001). Blastocyst-stage arrival rate (number of embryos at D5) per cycle was 55.8% (95% CI 54.3–57.2) in ≤ 15% SDF group (embryo quality score was ICM A:12.1%, B:69.5%, C:8.8%, D:4.5%; TE A: 7.5%, B:42.2%, C:42.2%, D:8.1%) and 55.9% (95% CI 52.8–59.1) in the >15% SDF group (ICM A:12.0%, B:68.7%, C:10.6%, D: 5.2%; TE A:9.1%, B:44.8%, C:37.8%, D:8.3%) (p < 0.001). The good quality blastocyst rate per cycle was significantly higher in the group with SDF ≤15%, 27.7% (95%CI 26.5–29.0) versus SDF >15% (27.4% (95%CI 24.6–30.2)). Of the total number of blastocysts, the proportion of A+B blastocyst was 60.5% (95% CI 58.3–62.7) and 64.2% (95% CI 59.2–69.2) (p < 0.001), respectively. Limitations, reasons for caution The retrospective and multicenter nature of this study leads to uncontrolled biases derived from the clinical practice. Although the results were not adjusted for female’s age, it was not statistically different between groups. Embryo morphology evaluation was performed by senior embryologists, it still remains a subjective evaluation, though. Wider implications of the findings: In this study, a higher amount of data was compiled so that a large number of embryos were analyzed. The DNA integrity of the sperm may be an important consideration when poor quality embryos were obtained in the previous cycle when deciding on the next clinical strategy to apply. Trial registration number NA

Comparison of PGS2.0 versus conventional embryo morphology evaluation for patients with recurrent pregnancy loss: a study protocol for a multicentre randomised trial

IntroductionPregnancy loss (PL) is an adverse life event, and there is no proven effective treatment for recurrent PL (RPL). Preimplantation genetic screening (PGS) can be performed to reduce the risks...

An inexpensive, automated artificial intelligence (AI) system for human embryo morphology evaluation and transfer selection

An inexpensive, automated artificial intelligence (AI) system for human embryo morphology evaluation and transfer selection

PGT-A and RCT proof in AMA and SMF couples

Aneuploidy is the most common genetic abnormality in human embryos. Preimplantation genetic diagnosis for aneuploidy (PGT-A) can increase pregnancy rates in infertile couples requiring assisted reproductive technology (ART). Large datasets from testing preimplantation embryos demonstrate that over half of embryos produced by in vitro fertilization (IVF) are aneuploid. Conventional embryo morphology evaluation as well as morphokinetics can not properly discriminate euploid from aneuploid embryos. Particularly, trisomic embryos that behave very similarly to euploid embryos. For these reasons, PGT-A started to be applied for several indications. In the present talk the results of two randomized trials (RCTs) in advanced maternal age (AMA) and severe male factor (SMF) will be discussed. In both studies, livebirths/ongoing pregnancy rates were compared among patients with blastocyst transfer as a control group and a study group in which day-3 biopsy was performed, and array CGH was applied for 24-chromosome analysis, with the transfer of euploid blastocysts. The final results of the AMA study conducted in women between 38 and 41 years of age included a total of 205 cycles. The PGT-A group exhibited significantly fewer embryo transfers (68.0%, vs. 90.5% for control; p=0.0001), and lower miscarriage rates (2.7%, vs. 39.0% for control; p=0.0007). Delivery rate after the first transfer attempt was significantly higher in the PGT-A group per transfer (52.9% vs. 24.2%; p=0.0002), and per patient (36.0% vs. 21.9%; p=0.0309). No significant differences were observed in the cumulative delivery rates per patient six months after closing the study. However, the mean number of embryo transfers needed per live birth was lower in the PGD-A group compared to control group (1.8 vs. 3.7), as was the time to pregnancy (7.7 vs. 14.9 weeks). The interim analysis of SMF study was performed when 101 cycles were completed and included women We can conclude that the benefits of testing embryos for common chromosomal abnormalities, include: increased ongoing implantation and pregnancy rates per transfer, decreased miscarriage rates per patient, and faster time to pregnancy when compared to conventional embryo scoring by morphology alone. Also, we concluded that PGT-A with the new approaches with throphectoderm biopsy and Next Generation Sequencing (NGS) the cost per treatment can decrease, being like a conventional cycle if we consider the endpoint of a healthy livebirth per cycle.

How much have we learned from time-lapse in clinical IVF?

Can the time-lapse system (TLS) identify the best embryo for transfer? Although there are several studies that support this hypothesis, more research is required to improve the quality of the current evidence and also to assess live birth rate, miscarriage, stillbirth or clinical pregnancy in order to choose between a TLS or conventional incubation. In addition, although some authors report on effectiveness and safety in the use of TLS monitoring of embryo development in vitro, other authors that have not found relevant differences between the two systems for the culture and subsequence embryo selection. On the other hand, TLS has emerged as a novel technology and has been introduced into clinical practice in many laboratories to perform embryo morphology evaluation and study developmental kinetics in ART. However, most studies only assess blastocyst formation or implantation rate as the primary end-point and additional data are required, for example, about live birth, monozygotic twinning rates and health problems. Furthermore, the features of populations studies are varied; for example, female and male age, seminal characteristics and female factor. The embryo culture conditions and culture medium used also vary. For this review, a search of PubMed was conducted to retrieve relevant studies regarding use of TLS in embryo incubation and selection, and compare them with standard embryo culture and evaluation.

Reply: Clinical outcomes following selection of human preimplantation embryos with time-lapse monitoring: a systematic review

BACKGROUND Time-lapse monitoring (TLM) has emerged as a novel technology to perform semi-quantitative evaluation of embryo morphology and developmental kinetics in assisted reproduction. While this method has already been introduced into clinical practice in many laboratories, it is unclear whether it adds value to conventional morphology. Most studies only report blastocyst formation as the primary end-point. The aim of this systematic review is to provide a critical evaluation of the available studies that report clinical outcomes following embryo selection with TLM. METHODS A literature search in MEDLINE, Cochrane CENTRAL and ISI Web of Knowledge Science Citation Index was performed to identify studies that assess the clinical utility of kinetic markers for non-invasive selection of human embryos with high implantation potential. Only studies published before 31 December 2013 in the English language that report rates of implantation, clinical pregnancy or live birth were included. RESULTS Two hundred and fifty-one studies were identified by database search and reference list review; only 13 met eligibility criteria and were included in this analysis. The following morphokinetic parameters were assessed: pronuclear dynamics and morphology (n = 3), duration of first cytokinesis and reappearance of nuclei after cleavage (n = 3), time to various cleavage stages (n = 5), duration of various cleavage stages (n = 6), duration of cleavage cycles and mitotic synchronicity (n = 6), and time to morula, blastocyst and hatching (n = 4). Five studies used combined parameter grading to generate a cumulative score, and two studies retrospectively compared implantation rates following embryo selection by conventional morphology alone or with the addition of a hierarchal time-lapse classification. While several studies suggest higher implantation rates for fast-cleaving embryos and those with a timely duration (i.e. all time points within the defined ranges) of the 2-cell and 3-cell stages, no single morphokinetic parameter has been consistently shown to predict implantation potential. Furthermore, there was considerable disagreement regarding not only which parameters are useful, but also what constitutes normal and abnormal intervals for these measurements. CONCLUSIONS While TLM has the potential to revolutionize clinical embryology, there are currently no high-quality data to support the clinical use of this technology for selection of human preimplantation embryos. Our recommendations for the adoption of this technique are thus limited by the available literature and the lack of robust prospective studies reporting clinical outcomes. Sparse, often incomplete and largely heterogeneous data suggest that TLM may be able to distinguish between high and low-implantation potential embryos. Only one study demonstrated significantly improved clinical pregnancy rates when embryos were selected by TLM in addition to conventional morphology. Prospective studies are currently underway and hopefully will clarify the role of TLM. As more data become available, it is of the utmost importance that groups using TLM share a common nomenclature for measured time points; herein, we have proposed a standardized system for describing any milestone along the preimplantation developmental timeline. Furthermore, future studies must publish complete datasets in an effort to define patient-specific algorithms with the clinically meaningful end-point of implantation, prior to routine adoption in the assisted reproduction technology laboratory. Until such evidence accumulates, selection of embryos by TLM should remain an experimental strategy subject to institutional review and approval.

The importance of the cleavage stage morphology evaluation for blastocyst transfer in patients with good prognosis.

To evaluate: (i) the influence of morphology at cleavage stage on blastocyst formation and implantation, and (ii) whether the transfer of low-quality embryos on day-three would be a better approach than the transfer at blastocyst stage. This study included 8,444 embryos obtained from 1,125 patients undergoing ICSI cycles between January/2011 and September/2013. The influence of the quality of the embryo on days-two and -three on blastocyst formation and implantation success was evaluated. Moreover, the implantation potential of low-quality embryos, at cleavage stage, transferred on day-three was compared with the implantation potential of low-quality embryos, at cleavage stage, transferred on day-five. Low-quality embryos on day-two had an approximate 20% decreased chance of achieving the blastocyst stage, and blastocysts derived from low-quality embryos on day-two had a nearly 40% decrease in the implantation chance. Low-quality embryos on day-three had a 30% decreased chance of achieving the blastocyst stage, and blastocysts derived from low-quality embryos on day-three had an almost 40% decreased implantation chance. The implantation rate didn't differ when low-quality embryos on the cleavage stage were transferred on day-three or left in culture and transferred on day-five. The transfer of low-quality embryos on day-three is a better approach than transfer at the blastocyst stage. In addition, the embryo morphology evaluation at the cleavage stage is still needed for the selection of the embryo with the best implantation potential in extended embryo culture programmes.

Clinical outcomes following selection of human preimplantation embryos with time-lapse monitoring: a systematic review.

Time-lapse monitoring (TLM) has emerged as a novel technology to perform semi-quantitative evaluation of embryo morphology and developmental kinetics in assisted reproduction. While this method has already been introduced into clinical practice in many laboratories, it is unclear whether it adds value to conventional morphology. Most studies only report blastocyst formation as the primary end-point. The aim of this systematic review is to provide a critical evaluation of the available studies that report clinical outcomes following embryo selection with TLM. A literature search in MEDLINE, Cochrane CENTRAL and ISI Web of Knowledge Science Citation Index was performed to identify studies that assess the clinical utility of kinetic markers for non-invasive selection of human embryos with high implantation potential. Only studies published before 31 December 2013 in the English language that report rates of implantation, clinical pregnancy or live birth were included. Two hundred and fifty-one studies were identified by database search and reference list review; only 13 met eligibility criteria and were included in this analysis. The following morphokinetic parameters were assessed: pronuclear dynamics and morphology (n = 3), duration of first cytokinesis and reappearance of nuclei after cleavage (n = 3), time to various cleavage stages (n = 5), duration of various cleavage stages (n = 6), duration of cleavage cycles and mitotic synchronicity (n = 6), and time to morula, blastocyst and hatching (n = 4). Five studies used combined parameter grading to generate a cumulative score, and two studies retrospectively compared implantation rates following embryo selection by conventional morphology alone or with the addition of a hierarchal time-lapse classification. While several studies suggest higher implantation rates for fast-cleaving embryos and those with a timely duration (i.e. all time points within the defined ranges) of the 2-cell and 3-cell stages, no single morphokinetic parameter has been consistently shown to predict implantation potential. Furthermore, there was considerable disagreement regarding not only which parameters are useful, but also what constitutes normal and abnormal intervals for these measurements. While TLM has the potential to revolutionize clinical embryology, there are currently no high-quality data to support the clinical use of this technology for selection of human preimplantation embryos. Our recommendations for the adoption of this technique are thus limited by the available literature and the lack of robust prospective studies reporting clinical outcomes. Sparse, often incomplete and largely heterogeneous data suggest that TLM may be able to distinguish between high and low-implantation potential embryos. Only one study demonstrated significantly improved clinical pregnancy rates when embryos were selected by TLM in addition to conventional morphology. Prospective studies are currently underway and hopefully will clarify the role of TLM. As more data become available, it is of the utmost importance that groups using TLM share a common nomenclature for measured time points; herein, we have proposed a standardized system for describing any milestone along the preimplantation developmental timeline. Furthermore, future studies must publish complete datasets in an effort to define patient-specific algorithms with the clinically meaningful end-point of implantation, prior to routine adoption in the assisted reproduction technology laboratory. Until such evidence accumulates, selection of embryos by TLM should remain an experimental strategy subject to institutional review and approval.

Pronuclear morphology evaluation in in vitro fertilization (IVF) / intracytoplasmic sperm injection (ICSI) cycles: a retrospective clinical review

BackgroundThe assessment of the embryo quality is crucial to maintain an high pregnancy rate and to reduce the risk of multiple pregnancy. The evaluation of the pronuclear and nucleolar characteristics of human zygote have been proposed as an indicator of embryo development and chromosomal complement. The aim of the current study was to assess the role of pronuclear morphology evaluation in vitro fertilization (IVF) / intracytoplasmic sperm injection (ICSI) cycles.MethodsRetrospective clinical analysis on 755 non-elective transfers of only one embryo (ET). Embryo assessment was performed in days 1 and 2. Clinical and biological data were recorded and analyzed according to embryo and/or pronuclear morphology.ResultsBoth pronuclear and embryo morphology were significantly related to clinical pregnancy and live-birth rates. No significant difference in clinical pregnancy and live-birth rates was detected when the pronuclear and embryo morphology assessments were combined. Embryo morphology and maternal age were the only independent predictors of favorable outcome by logistic regression analysis.ConclusionsPronuclear evaluation is effective to select the best zygotes if ET is performed at day 1, whereas it did not improve the clinical outcomes when combined with embryo morphology evaluation in day 2.

Advanced cleavage embryos submitted to PGS-FISH have a high frequency of aneuploidy

OBJECTIVE: Embryo morphology evaluation is the standard procedure to select the best embryo to be transferred to the uterus. Theoretically, the ideal cleavage rate yields embryos with 4 cells on day 2 and 8 cells on day 3. Sometimes in vitro culture conditions produces embryos with a faster cleavage rate showing ≥ 5cells on day 2 and ≥ 10 cells on day 3, possibly demonstrating a better performance. We sought to investigate aneuploidy rates (AR) by preimplantation genetic screening (PGS-FISH) of advanced cleavage embryos (ACE) in comparison to AR of historic controls. DESIGN: Retrospective study. MATERIALS AND METHODS: Embryos produced in vitro during 2008 and submitted to PGS-FISH had their 3rd quartile of number of cells on days 2 and 3 of culture computed. Embryos above the third quartile of the number of blastomeres on day 2 and day 3 during in vitro culture were considered ACE. Aneuploidy rates of ACE were compared to AR of historic controls for the same age, dividing women by age in 35-39 years old and above 40 years old by Z test for two proportions. RESULTS: Seventy-five embryos from 43 patients (40±4 years old) were above the third quartile of development (#cells D2=4; #cells D3=8) and were considered ACE (#cells D2=5±0.8; #cells D3=10±0.7). Abnormal cleaved embryos were submitted to PGS-FISH for 5(n=65; 86.6%) or 9(n=11;13.4%) chromosomes and our data showed an overall aneuploidy rate of 69.4%. The proportion of abnormal embryos was higher in ACE if compared to historic controls including women aging 35-39 years old (49.7%; 95% confidence interval-CI=-0.3; -0.08; p=0.001) or women above 40 years old (55.6%; 95%CI=0.01; 0.,2;p=0.02). CONCLUSIONS: Our data suggests that embryos with accelerated cleavage rate, more than 5 cells on day 2 and 10 cells on day 3, although seem to produce a better performance, is associated to an increased aneuploidy rate after PGS-FISH evaluation and should not be considered to be transferred.

Identification of novel non-invasive biomarkers of oocyte aneuploidy

OBJECTIVE: Although evaluation of embryo morphology provides useful information concerning the implantation potential of individual embryos, it is acknowledged that most morphological criteria are only weakly linked to IVF outcome. Thus, the ability to identify viable embryos for transfer based upon morphological assessment is limited. Genetic analysis of the embryo (i.e. PGD) can provide important information concerning the presence of lethal chromosome anomalies, supplementing morphological evaluation.

Zygote score and status 1 or 2 days after cleavage and assisted reproduction outcome

Objective:To determine whether pronuclear morphology is a useful additional criterion when selecting embryos for transfer. Method:A modified Tesarik and Greco scoring system was used in this retrospective study to assess the pronuclear morphology of 883 zygotes from 214 in vitro fertilization or intracytoplasmic sperm injection cycles. Pronucleus size and presence of cytoplasmic halo were also noted. Embryo selection was performed on Day 2 or 3 following oocyte retrieval. The χ2, analysis of variance, Kruskal–Wallis, and Fisher exact tests were used where appropriate. Results:A stepwise logistic regression revealed that a better embryo quality was associated with both pattern 0 of the scoring system and pronuclei of equal size on Days 2 or 3. Higher pregnancy and multiple gestation rates were obtained when at least 1 embryo classified as pattern 0 was included in the set of embryos transferred. Conclusion:Combined with embryo morphology evaluation on Days 2 and 3, a scoring system based on pronuclear morphology seems to provide a good criterion when selecting embryos for transfer.

Comparative Studies on the mRNA Expression of Development-Related Genes in an Individual Mouse Blastocyst with Different Developmental Potential

The evaluation of embryo morphology, widely used for selecting mammalian embryos before transfer, is not an adequate standard for selecting nuclear-transferred (NT) embryos. To search for markers useful for predicting the potential of NT embryos to develop into young, we examined the relation between the morphology of embryos with different developmental potential and gene expression of Oct 4, Nanog, Stat3, FGF4, Stella, and Sox2. In the present study, we examined pronuclear-exchanged blastocysts and morula blastomere, embryonic stem (ES) cell, and cumulus cell NT blastocysts, and in vivo-developed and in vitro-developed blastocysts. Based on the small variations in the gene expression levels among the in vivo-developed blastocysts, and the significant differences in gene expression between in vivo-developed (high developmental potential), and ES cell and cumulus cell NT blastocysts (low developmental potential), down-regulation of Sox2 and Oct4 genes is considered to be a candidate marker for the low potential of NT embryos to develop into young.

P-314: Do interventions necessary to overcome technical difficulties encountered in trial embryo transfer and employed during real embryo transfer really improve outcome in IVF cycles?

To evaluate the impact of intervention techniques employed in real embryo transfer to overcome technical difficulties presented during trial transfer on the outcome of IVF cycles. Prospective observational study of 418 consecutive IVF/ET cycles. Trial embryo transfer (TET) to map the cervical canal was performed with a soft polyethylene catheter and a firm outer Teflon introducer (Wallace, SIMS Portex, UK) in all cycles immediately before the real transfer. All trial and real embryo transfers (RET) were performed under abdominal ultrasound guidance 72 hours after oocyte retrieval and only RETs considered uneventful (n=364) were compared. Interventions suggested by TET that were employed in RET to overcome difficulties included the use of: Group A (n=184)- soft catheter, no interventions; Group B (n=97)- soft catheter, insertion of the outer introducer through the endocervical canal and internal os, with or without the help of a tenaculum and; Group C (n=83)- firm catheter (Tefcat, Cook, USA), with or without the help of a tenaculum. All catheters were loaded with a maximum of four embryos in 20-30μl of culture medium after embryo morphology evaluation and grading. After RET, the following variables were recorded: presence of blood and/or mucus in or on the transfer catheter; cramping or patient discomfort; retention of embryos in the transfer catheter. There was no difference between groups with respect to patients’ main demographic and baseline characteristics, embryo development rate, mean number and quality at transfer. The overall clinical pregnancy rate/transfer for the different interventions was 48.4%, 41.2% and 25.3% in groups A, B and C, respectively, showing a significant difference (p=0.02) between groups A and C. Likewise, the implantation rate was significantly higher (p=0.005) in group A (26.2%) than in group C (15.6%) and revealed a trend (p=0.08) toward better results when compared to group B (19.7%). Clinical pregnancy/implantation rates showed no difference when groups B and C were compared. Contamination of the catheter with blood and/or cervical mucus was more frequent in group B (68.0%) when compared to groups A (37.5%) and C (41.0%) (p=0.0001, p=0.0004; respectively). The occurrence of uterine cramping was 3.6%, 10.2% and 18.6% in groups A, B and C, respectively, showing a significant difference (p=0.001) between groups A and C and no difference when groups A and B or B and C were compared. There was no difference between groups in regard to the incidence of retained embryos. The use of more invasive and potentially traumatic interventions by clinicians to efficiently overcome a difficult TET during RET may, by themselves, significantly impair the outcome of IVF cycles. This is reflected in the lower pregnancy and implantation rates observed in the study, regardless of degree of ease in execution of the procedure. Every maneuver should be tried with a softer catheter before changing it to a firmer one, since it is clearly shown by this study that the latter has a negative impact on the overall outcome. A balance between the choice of the intervention necessary to overcome the difficulties encountered during TET, and the expected decrease in pregnancy/implantation rates should always be taken into account by the clinician in charge of the embryo transfer, since different intervention techniques may have negative influence on the outcome of IVF cycles to varying degrees.

Pronuclear morphology evaluation with subsequent evaluation of embryo morphology significantly increases implantation rates

Objective To elucidate the relative predictive value of implantation markers at different stages of preimplantation development. Design Correlation of pronuclear morphology with embryo morphology and implantation rates in retrospective and prospective analysis of in vitro fertilization/intracytoplasmic sperm injection (IVF-ICSI) treatment cycles. Setting Private infertility center. Patient(s) A total of 441 couples undergoing infertility treatment. Intervention(s) None. Main outcome measure(s) Size of pronuclei and distance between them, the number and polarization of nucleolus precursor bodies (NPB) at the one-cell stage, embryo cleavage and fragmentation rates on days 2 and 3, and pregnancy and implantation rates. Result(s) Polarization of the NPB in both pronuclei had a statistically significant correlation with normal membrane breakage during ICSI (40%, compared with 33% easy, and 31% difficult membrane breakage) and also with faster cleavage and lower fragmentation rates of embryos. Sixty-one percent of implanting embryos had polarization of the NPB in both pronuclei compared with 37% for all embryos. Larger distance between pronuclei and their unequal size had a statistically significant correlation with slower cleavage and inferior embryo quality. Embryo selection based on only pronuclear morphology or on only day-3 embryo morphology yielded implantation rates of 15.1% and 12.1%, respectively. Embryo selection based on sequential evaluation of both pronuclear morphology and embryo morphology on day 3 resulted in a 21.1% implantation rate. Conclusion(s) Polarization of NPB in both pronuclei is as reliable marker of implantation as embryo morphology on day 3. However, pronuclear morphology assessment improves embryo selection only when it is combined with embryo morphology evaluation on day 3.

Embryo quality and andrological study of two subfertile bulls versus five control bulls with normal fertility

Andrological studies and embryo morphology evaluation of superovulated cows were performed on 2 randomly selected subfertile dairy bulls whose semen was used for artificial insemination and on 5 control bulls with normal fertility. Neither sperm motility studies, nor sperm morphology or testicular measurements differed between the subfertile and the control bulls. Altogether 315 ova were recovered from 41 superovulated cows inseminated with semen collected from either the subfertile or the normal control bulls. The spermatozoa of one of the 2 subfertile bulls was shown to have a decreased ability to fertilize superovulated ova, while the other subfertile animal, the bull with the lowest noreturn rate, was found by chromosome analysis to have a reciprocal translocation (60, XY, rcp 20:24), causing embryonic death. We suggest that subfertile bulls should not be used in commercial embryo transfer programs nor in artificial insemination and that andrological studies on subfertile bulls with good sperm motility should include evaluation of 6- to 7-day-old ova from superovulated cows to determine if the fertilization rate is normal or impaired. A chromosome analysis should also be performed when a subjertile bull has a normal fertilization rate of ova.