Comprehensive Chromosome Screening Research Articles

High normal CGG repeat number on the fragile x mental retardation 1 (FMR1) gene is not correlated with diminished ovarian reserve

OBJECTIVE: Inversions are structural chromosome abnormalities that are associated with infertility due to the increased risk of producing unbalanced gametes from recombination events within the inverted chromosome segment. However, there is also evidence indicating the presence of an inter-chromosomal effect during chromosome segregation influencing the generation of other chromosome aneuploidies. The aim of this study was to evaluate the nature of chromosome errors in IVF blastocysts from carriers of balanced chromosome inversions. DESIGN: Research study. MATERIALS AND METHODS: Infertility patients with balanced chromosome inversions (n1⁄430; female carriers1⁄420, male carriers1⁄410) and maternal age-matched infertile controls with normal karyotypes that concurrently cycled (n1⁄430), consented with IRB approval to an IVF cycle with comprehensive chromosome screening (CCS). All embryos were cultured to the blastocyst stage with a trophectoderm biopsy performed for CCS using SNP microarray (RMA-NJ). Only euploid or balanced blastocysts were transferred in a subsequent frozen embryo transfer. Statistical analysis of aneuploidy and outcome data was performed using a two sided Fisher’s exact test with p value of 0.05 for significance. RESULTS: Maternal ovarian reserve, paternal age, oocytes retrieved, fertilization rate and the number of blastocysts biopsied were comparable between the two groups. The chromosomes that contained the balanced inversions varied but were mostly the larger chromosomes including 1, 2, 3, 5, 7, 9 and 17. The incidence of aneuploidy, excluding the chromosome with the inversion, was significantly higher for the inversion patients compared to the maternal age matched controls (Inversion1⁄459% vs. Control1⁄449%; P<0.001). However, following transfer of a euploid blastocyst there was an equivalent live birth rate (Inversion1⁄473.7% vs. Control1⁄478.3%; ns). CONCLUSION: Carriers of balanced chromosome inversions exhibited higher aneuploidy rates for chromosomes that were not involved in the inversion compared to maternal age-matched controls, signifying the occurrence of an inter-chromosomal effect. However, despite a higher aneuploidy rate, carriers of balanced inversions experienced excellent ART outcomes following the transfer of euploid blastocysts. These results provide valuable information for clinical management of infertility patients carrying balanced inversions prior to infertility treatment.

Serum anti-mullerian (AMH) levels do not predict embryo ploidy after comprehensive chromosomal screening (CCS) of trophectoderm cells during preimplantation genetic screening (PGS)

Serum AMH concentrations decline with increasing age, constituting a sensitive marker for ovarian aging. On an embryo by embryo basis, it is not clear whether abnormal AMH levels predicts aneuploidy. This study seeks to determine if serum levels of AMH can be associated with the percentage of embryos deemed aneuploid using CCS of trophectoderm cells during PGS.

Prolonged time to first cytokinesis and the interval between the five cell stage and early cavitation are associated with embryonic mosaicism

The use of time lapse imaging through day 3 of development provides insight into the probability a cleavage stage embryo will blastulate. This raises the question of whether these parameters are correlated with other biologic phenomena occurring during the same time interval. One such parameter is mosaicism. Mosaicism is a particularly problematic abnormality as it may result in misdiagnoses from comprehensive chromosome screening (CCS) and is a potential reason why some embryos designated as euploid fail to deliver.

Blastocysts available for biopsy on day 5 are more likely to result in a viable pregnancy

Comprehensive chromosomal screening has become a routine test in many IVF practices around the globe. Reliable information about euploidy status of the embryos created a background for significant improvement in IVF treatment outcomes (Munné et. al., 2010). Many patients have euploid embryos with similar morphological characteristics biopsied on day 5 and day 6. The objective of this study was to evaluate pregnancy rates in frozen embryo transfer cycles in which blastocyst were biopsied on day 5 versus day 6.

Endometrial thickness has no impact on implantation rates or clinical outcomes in euploid embryo transfers

Past attempts to correlate endometrial thickness (EnT) with implantation rates (IR) and pregnancy rates (PR) have been biased due to variability in embryo quality and the unidentified genetic composition at embryo transfer (ET). With the utility of trophectoderm biopsy (TB) and comprehensive chromosome screening (CCS); a more precise analysis can now be performed. To evaluate our hypothesis that a thicker endometrium is more associated with a desired clinical outcome, we assessed whether EnT impacts implantation rate.

High cellularity of trophectoderm biopsy adversely affects pregnancy outcomes

Comprehensive chromosome screening (CCS) is becoming more frequently utilized to aid in embryo selection and optimize live birth rates. Level I data supports the safety and accuracy of trophectoderm (TE) biopsy, but not all TE biopsies are equal. Whether the size of the biopsy impacts reproductive potential is unknown. Here, the association between TE biopsy cellularity and pregnancy outcomes is assessed.

An embryo cohort which contains all aneuploid embryos is not indicative of future embryo cohort aneuploidy

Use of comprehensive chromosome screening (CCS) has enhanced embryo selection. However, some patients will have 100% of their cohort return as aneuploid. This varies from 5% in women in their late 20s to 55% in women >45. Beyond the initial disappointment, patients and clinicians are left with the difficult decision about whether or not to attempt another IVF cycle. Is a cohort which is 100% aneuploid indicative of future aneuploidy rates? The data presented here seek to answer those questions and assist in patient counseling and clinical decision making.

The accuracy of blastocoel fluid comprehensive chromosomal screening (CCS) is dependent on amplification yield and sequencing depth when using nextgen sequencing

The presence of DNA in the blastocoel provides an attractive target for less invasive aneuploidy screening. However, blastocoel fluid provides many challenges including DNA quantity and integrity. To date, a functional screening algorithm that overcomes these challenges and accurately identifies the karyotype of the blastocyst has not been validated. This study seeks to determine consistency of diagnosis between DNA isolated from trophectoderm and blastocoel fluid using NextGen based CCS.

Routinely distinguishing normal and balanced translocation carrier embryos using snp arrays

Many translocation carrier couples convey a strong interest in preventing transmission to their offspring, sparing them the reproductive challenges they faced. However, contemporary methods of comprehensive chromosome screening (CCS) cannot reliably distinguish between normal embryos and those that carry a balanced translocation. This study was conducted to develop a method that can concurrently identify unbalanced and aneuploid embryos, and, for the first time, distinguish translocation carrier and truly normal embryos all from the same embryonic trophectoderm biopsy SNP array data.

Blastocyst aneuploidy rates, a comparison of different comprehensive chromosome screening platforms: results from a single center

Blastocyst aneuploidy rates, a comparison of different comprehensive chromosome screening platforms: results from a single center

Proportion of aneuploidy does not impact live birth rate or pregnancy loss rate in patients with recurrent pregnancy loss (RPL) undergoing comprehensive chromosome screening

There is limited evidence that aneuploidy disproportionately effects outcomes in patients with RPL as compared to the general infertility population. In an effort to investigate the potential that patients with RPL without balanced translocations have a biologic mechanism related to aneuploidy behind their poor pregnancy outcomes, we sought to evaluate if the proportion of aneuploidy within a cohort of embryos impacts pregnancy outcomes. If an intrinsic biologic deficit exists, we suspect that patients with high aneuploidyrates in the RPL population would be disproportionately helped by CCS.

Rapid and concurrent comprehensive chromosome screening (CCS) and single gene disorder (SGD) PGD

Combining accurate CCS with SGD diagnosis from the same biopsy has presented many challenges. Current methods can either fail to detect all origins of aneuploidy, require excessive workup times, involve long PGD turn-around times, or are expensive to perform. The present study develops a method which overcomes each of these limitations using quantitative real-time (q)PCR.

Blastocyst euploidy and implantation rates in a young (

To examine the relationship between blastocyst euploidy and implantation rates in a presumed fertile patient population. Retrospective analysis. Private IVF clinic. IVF patients undergoing comprehensive chromosome screening (CCS). Embryo biopsy at the blastocyst stage with preimplantation genetic screening using CCS. Euploidy, chemical pregnancy, and implantation rates. There was no significant difference in the number of euploid blastocysts between presumed fertile (68/118, 57.6%) and infertile (75/132, 56.8%) patients<35 years old. Likewise, there was no significant difference in the number of euploid blastocysts between presumed fertile (42/86, 48.8%) and infertile (97/206, 47.1%) patients≥35 years old. When those same patients underwent a corresponding frozen embryo transfer cycle, presumed fertile patients demonstrated a significantly higher chemical pregnancy rate when compared with infertile patients, 28/33 (84.8%) and 50/81 (61.7%), respectively. Moreover, presumed fertile patients exhibited significantly higher implantation rates compared with infertile patients, 36/42 (85.7%) and 54/109 (66.7%), respectively. When subdivided by maternal age, no significant difference was seen in blastocyst euploidy rates between presumed fertile and infertile patients; however, chemical pregnancy and implantation rates were significantly higher in a presumed fertile patient population even when transferring only euploid blastocysts. This would indicate that infertility, as a disease, may encompass other aspects such as uterine or other unknown embryological factors that can influence outcomes.

Clinical utilisation of a rapid low-pass whole genome sequencing technique for the diagnosis of aneuploidy in human embryos prior to implantation

BackgroundThe majority of human embryos created using in vitro fertilisation (IVF) techniques are aneuploid. Comprehensive chromosome screening methods, applicable to single cells biopsied from preimplantation embryos, allow reliable identification and...

Comparison of aneuploidy, pregnancy and live birth rates between day 5 and day 6 blastocysts

Comprehensive chromosome screening is typically used for aneuploidy analysis of blastocysts. It is believed that either day of blastocyst development is acceptable. Euploidy rates and outcomes were examined between day 5 and day 6 blastocysts in two studies. First, euploidy rates of day 5 and day 6 blastocysts were examined on a per-embryo and per-patient basis. Second, outcomes were compared when only euploid day 5 or day 6 blastocysts were transferred in a cryopreserved embryo transfer cycle. In cycles (n = 70) that had blastocysts biopsied on both day 5 and day 6, day 5 blastocysts had a higher chance of being euploid than day 6 blastocysts (125/229 [54.6%]) and (77/180 [42.8%]), respectively (P = 0.0231). Similarly, euploid rates in blastocysts from patients (n = 193) with day 5 biopsy, day 6 biopsy, or both, were significantly higher in day 5 (235/421 [55.8%]) compared with day 6 (184/413 [44.6%]) blastocysts (P = 0.0014). In the second study, 50 women (36.1 ± 4.3 years) and 39 women (35.1 ± 3.8 years) with only euploid day 5 or euploid day 6 blastocysts transferred during a cryopreserved embryo transfer had similar cycle outcomes. Although underpowered, these data suggest that euploid day 6 blastocysts are as capable of positive outcomes as their euploid day 5 counterparts.

Obstetrical and Neonatal Outcomes From the BEST Trial

Nearly 50% of all US babies born after in vitro fertilization are multiple gestations. Because nearly all deliveries after elective single-embryo transfer (eSET) are singletons, the authors hypothesized that obstetrical and neonatal outcomes could be improved in patients randomized to euploid eSET compared with those undergoing transfer of 2 untested embryos. The Blastocyst Euploid Selective Transfer (BEST) trial was a randomized, noninferiority study comparing SET after real-time quantitative polymerase chain reaction–based comprehensive chromosome screening with transfer of 2 untested embryos. When at least 2 embryos reached the blastocyst stage of development, patients were randomized to the screened euploid eSET or untested 2-embryo transfer group. The primary outcome was the ongoing pregnancy rate to a viable gestation after the first embryo transfer, fresh or frozen. This current study is an analysis of the final obstetrical and neonatal delivery outcomes of patients randomized in the earlier trial after the initial fresh cycle and up to 1 frozen transfer. Patients provided data on gestational age at delivery, mode of delivery, birth weight, sex, pregnancy complications, and number of days their newborns were in the neonatal intensive care unit (NICU). Eighty-nine and 86 patients were randomized to the euploid eSET and untested 2-embryo transfer group, respectively. The 2 groups were similar in all demographic characteristics; the mean (SD) ages were 35.1 (3.9) and 34.5 (4.7) years (P = 0.5), respectively. The mean number of blastocysts suitable for transfer was 5.8 (range, 2–22) for eSET and 5.3 (range, 2–18) for 2-embryo transfer. In the eSET group, a biopsy of 521 blastocysts was performed for comprehensive chromosome screening, with an aneuploidy rate of 31% (n = 162). The proportion of aneuploid embryos increased with increasing age (21%, 34%, 56%, and 56% for ages <35 years, 35–37 years, 38–40 years, and 41–42 years, respectively; P < 0.001). The cumulative delivery rate after up to 1 frozen transfer was 69% after eSET and 72% after 2-embryo transfer (P = 0.6). Interestingly, 58% of the transferred embryos resulted in live births after eSET of genetically screened embryos; and 46%, after 2-embryo transfer of untested embryos (P = 0.048). Rates of multiple deliveries were 47% in the 2-embryo transfer group and 1.6% in the eSET group (relative risk [RR], 28.5; 95% confidence interval, 4.01–202.9). Per patient randomized, the chance of having a term singleton delivery was 60% after eSET and 31% after 2-embryo transfer (P < 0.001). The risk for preterm delivery was 29% after 2-embryo transfer and 13% for eSET (RR, 2.21; 95% confidence interval, 1.04–4.70; P = 0.03). The risk for a newborn being less than 2500 g was 11% in the eSET group and 33% in the 2-embryo transfer group (P = 0.002). The risk for a newborn being admitted to the NICU was 26% and 11% in the 2-embryo transfer and eSET group, respectively (P = 0.04). Culturing embryos to the blastocyst stage, performing a trophectoderm biopsy, and amplifying DNA with quantitative polymerase chain reaction assays on each chromosome will allow selection of a single euploid blastocyst with high reproductive potential. This strategy dramatically reduces the risk for multiple gestation and increases the chance for a healthy, term singleton delivery without requiring patients to undergo many failed cycles. The improved obstetrical and neonatal outcomes suggest that this approach may become the standard of care for couples requiring in vitro fertilization.

Human blastocysts exhibit unique microrna profiles in relation to maternal age and chromosome constitution.

To determine microRNA (miRNA) expression in human blastocysts relative to advanced maternal age and chromosome constitution. Cryopreserved human blastocysts were warmed and underwent a trophectoderm biopsy for comprehensive chromosomal screening. Select blastocysts were then lysed, reverse transcribed, and pre-amplified prior to running real-time PCR. Statistical analysis was performed using an internal constant housekeeping miRNA. Significant microRNA's of interest were then analyzed for their predicted genes and biological pathways. Additional cryopreserved blastocysts were warmed and stained for the SIRT1 protein for validation. Human blastocysts exhibit unique miRNA expression profiles in relation to maternal age and chromosome constitution. miR-93 was exclusively expressed in blastocysts from women in their forties and further up-regulated with an abnormal chromosome complement. Up-regulated miR-93 resulted in an inverse down-regulation of targets like SIRT1, resulting in reduced oxidative defense. MiRNAs play an important role in aging as well as chromosome constitution and have downstream effects that regulate proteins which can compromise embryonic development.

A greater number of euploid blastocysts in a given cohort predicts excellent outcomes in single embryo transfer cycles

This multicentered retrospective study analyzed whether the quantity of euploid blastocysts in a given cohort after comprehensive chromosomal screening can be used to identify candidates for single embryo transfer. Blastocysts from 437 patients underwent trophectoderm biopsy followed by array comparative genomic hybridization. Embryos were then selected for single or double embryo transfer. The number of euploid blastocysts produced and transferred for each patient was recorded, as was clinical pregnancy rate and multiple gestation rate. In patients with ≤ 3 euploid blastocysts, clinical pregnancy rate was higher in double, compared to single embryo transfers. However, in patients with ≥ 4 euploid blastocysts, clinical pregnancy rate was not reduced with single embryo transfer was performed, whereas the multiple gestation rate was greatly reduced. Size of the euploid embryo cohort is a marker for success in single embryo transfer cycles. Patients who produce at least four euploid blastocysts are outstanding candidates for single embryo transer.

Outcomes of blastocysts biopsied and vitrified once versus those cryopreserved twice for euploid blastocyst transfer

Trophectoderm biopsy with comprehensive chromosome screening (CCS) has been shown to increase implantation and pregnancy rates. Some patients desire CCS on previously cryopreserved blastocysts, resulting in blastocysts that are thawed/warmed, biopsied, vitrified and then warmed again. The effect of two cryopreservation procedures and two thawing/warming procedures on outcomes has not been effectively studied. Cycles were divided into two groups: group 1 patients underwent a cryopreserved embryo transfer with euploid blastocysts that were vitrified and warmed once; group 2 patients had a cryopreserved embryo transfer of a euploid blastocyst that was cryopreserved, thawed/warmed, biopsied, vitrified and warmed. Groups 1 and 2 included 85 and 17 women aged 35.6±3.9 and 35.3±4.9years, respectively (not significantly different). Blastocyst survival in group 1 (114/116, 98.3%) and survival of second warming in group 2 (21/24, 87.5%) was significantly different (P=0.0354). There was no difference between biochemical (68.2% and 62.5%) and clinical (61.2% and 56.3%) pregnancy rates, implantation rate (58.4% and 52.4%) and live birth/ongoing pregnancy rate (54.0% and 47.6%) between groups 1 and 2, respectively. Although it is unconventional to thaw/warm, biopsy, revitrify and rewarm blastocysts for cryopreserved embryo transfer, the results indicate that outcomes are not compromised.Trophectoderm biopsy and screening the embryos for chromosomal abnormalities has been reported to increase implantation and pregnancy rates. There is a category of patients requesting chromosomal screening on previously cryopreserved blastocysts. This scenario requires blastocysts to be thawed/warmed, biopsied, cryopreserved, and thawed/warmed again. The effect of double cryopreservation procedures and double thawing/warming procedures on pregnancy is unknown. Patients were divided into two groups, group 1 underwent a cryopreserved embryo transfer with a chromosomally normal blastocyst that was vitrified and warmed once and group 2 included patients that had a cryopreserved embryo transfer of a chromosomally normal blastocyst that was cryopreserved, thawed/warmed, biopsied, vitrified, and rewarmed. A total of 85 and 17 women aged 35.6±3.9 and 35.3±4.9years were included in groups 1 and 2, respectively. The survival rate for group 1 (114 of 116, 98.3%) compared with the second warming for group 2 (21 of 24, 87.5%) was significantly higher. There was no difference between biochemical (68.2% and 62.5%), and clinical pregnancies (61.2% and 56.3%), implantation (58.4% and 52.4%), and live birth/ongoing rates (54.0% and 47.6%) between groups 1 and 2. Although it is unconventional to twice cryopreserve and twice thaw/warm a blastocyst, our results indicate that outcomes are not compromised.

Correlation between standard blastocyst morphology, euploidy and implantation: an observational study in two centers involving 956 screened blastocysts

Does conventional blastocyst morphological evaluation correlate with euploidy (as assessed by comprehensive chromosome screening (CCS) of trophectoderm (TE) biopsies) and implantation potential? A moderate relation between blastocyst morphology and CCS data was observed but the ability to implant seems to be mainly determined by the chromosomal complement of preimplantation embryos rather than developmental and morphological parameters conventionally used for blastocyst evaluation. Combined with improving methods for cryopreservation and blastocyst culture, TE biopsy and CCS is considered to be a promising approach to select euploid embryos for transfer. Understanding the role of morphology in blastocyst stage preimplantation genetic screening (PGS) cycles may help in further optimizing the cycle management and clinical outcomes. This is a multicenter retrospective observational study performed between January 2009 and August 2013. The study includes the data analysis of 956 blastocysts with conclusive CCS results obtained from 213 patients following 223 PGS cycles. Single frozen embryo transfer (FET) cycles of 215 euploid blastocysts were performed where it was possible to track the implantation outcome of each embryo transferred. PGS was offered to infertile patients of advanced maternal age (>35 years) and/or with a history of unsuccessful IVF treatments (more than two failed IVF cycles) and/or previous spontaneous abortion (more than two spontaneous miscarriages). Prior to TE biopsy for CCS, blastocyst morphology was assessed and categorized in four groups (excellent, good, average and poor quality). The developmental rate of each embryo reaching the expanded blastocyst stage was defined according to the day of biopsy post-fertilization. Day 5 and Day 6 biopsied blastocysts were defined as faster and slower growing embryos, respectively. A novel blastocyst biopsy method, not requiring the opening of the zona pellucida at the cleavage stage of embryo development, was used. Linear regression models were used to test the relationship between blastocyst morphology and developmental rate CCS data and FET cycle outcomes of euploid blastocysts. Among the embryological variables assessed (morphology and developmental rate), only blastocyst morphology was predictive of the CCS data. The euploidy rate was 56.4, 39.1, 42.8 and 25.5% in the excellent, good, average and poor blastocyst morphology groups, respectively. A diagnosis of complex aneuploidy was also associated with blastocyst morphology (P < 0.01) with 6.8, 15.2, 17.4 and 27.5% of excellent, good, average and poor quality embryos, respectively, showing multiple chromosome errors. Faster and slower growing embryos showed a similar aneuploidy rate. Regression logistic analysis showed that none of the parameters used for conventional blastocyst evaluation (morphology and developmental rate) was predictive of the implantation potential of euploid embryos. The implantation potential of euploid embryos was the same, despite different morphologies and developmental rates. The study is limited by its retrospective nature. A higher sample size or a prospective randomized design could be used in future studies to corroborate the current findings. This study provides knowledge for a better laboratory and clinical management of blastocyst stage PGS cycles suggesting that the commonly used parameters of blastocyst evaluation are not good enough indicators to improve the selection among euploid embryos. Accordingly, all poor morphology and slower growing expanded blastocysts should be biopsied and similarly considered for FET cycles. This knowledge will be of critical importance to achieve similar cumulative live birth rates in PGS programs compared with conventional IVF, avoiding the potential for exclusion of low quality but viable embryos from the biopsy and transfer procedures. Future research to identify non-invasive biomarkers of reproductive potential may further enhance selection among euploid blastocysts. No funding was obtained for the study. All authors have no conflicts to declare. None.