Search

Pre-Implantation genetic diagnosis is available to couples at risk of conceiving a pregnancy affected with a known genetic disorder. Assisted reproductive techniques are used in combination with micromolecular diagnostic technologies to recognise at-risk embryos with pathogenic genetic variants at the pre-implantation stage using polar body, blastomere or trophectoderm biopsy. This review will discuss the varying genetic disorders diagnosed by Pre-Implantation Genetic Diagnosis, as well as the ethical, legal and safety implications of the process. Pioneering advances in molecular biology and cytogenomics have been utilised to expand the spectrum of genetic disorders detected.

Background Although preimplantation genetic test (PGT) has been used worldwide, few studies investigated the effect of trophectoderm biopsy of blastocysts on early embryo development. This study aimed to investigate whether trophectoderm (TE) biopsy of blastocysts for a PGT affected serum β-human chorionic gonadotropin (hCG) levels 14 days after transfer. Methods This was a retrospective cohort study conducted at the Third Affiliated Hospital of Guangzhou Medical University. The study population comprised pregnant women undergoing the transfer of single vitrified-warmed blastocysts after PGT between January 1, 2018, and July 30, 2020. The control group had non-PGT cycles with other inclusion criteria identical to those for the study group. Propensity score matching was used to screen a group of patients so that the baseline characteristics were similar between the two groups. Serum β-hCG levels were compared between the PGT and non-PGT cycles. Multiple linear regression was used to analyze the influence of PGT on serum β-hCG levels, while receiver operating characteristic curves (ROC curves) were plotted to predict pregnancy outcomes using serum β-hCG levels. Results Serum β-hCG levels were comparable between the PGT and non-PGT patients: live birth: 2503 ± 1702 mIU/mL vs 2266 ± 1289 mIU/mL (P = 0.219); clinical pregnancy: 2261 ± 1564 mIU/mL vs 2148 ± 1348 mIU/mL (P = 0.461); and ongoing pregnancy: 2412 ± 1589 mIU/mL vs 2278 ± 1308 mIU/mL (P = 0.422). Multiple linear regression analysis indicated no impact of PGT on the serum β-hCG level (standardized coefficient = − 0.001, P = 0.989). For clinical pregnancy, the cutoff value was 482 mIU/mL and 302 mIU/mL for PGT and non-PGT patients, respectively. The threshold to predict live birth was 1345 mIU/mL and 1621 mIU/mL in the PGT and non-PGT cycles, respectively. Conclusion Trophectoderm biopsy of blastocysts for PGT did not affect the serum β-hCG level 14 days after transfer.

Preimplantation genetic testing for monogenic/single-gene defects (PGT-M) is a well established tool in assisted reproduction. A couple, where the male was carrier of the c.550G>A (p.Gly184Arg) mutation of the SERPING 1 gene causing hereditary angioedema (HAE) attended our clinic for PGT-M. Employing a strategy of preserving embryos after blastocyst trophectoderm biopsy by vitrification cryopreservation, we managed to screen 10 embryos collected from a single IVF cycle. Three embryos were found unaffected after preimplantation genetic analysis for the paternal mutation. The transfer of one normal blastocyst post-thaw resulted in a healthy and uneventful pregnancy and in the live birth of a male neonate on the 38th week of pregnancy. This is the first report of a live birth in Greece after blastocyst trophectoderm biopsy and preimplantation genetic analysis for hereditary angioedema.

Background Spent culture medium (SCM) as a source of DNA for preimplantation genetic tests aneuploidy (PGT-A) has been widely discussed. Methods Seventy-five blastocysts that were donated for research provided a unique possibility in which multiple specimens, including trophectoderm (TE) biopsy, SCM, and paired corresponding whole blastocyst (WB) specimens from the same blastocyst source, could be utilized for the purpose of this preclinical validation. Results To conduct a validation ploidy concordance assessment, we evaluated the full chromosomal concordance rates between SCM and WB (SCM-to-WB), and between TE and WB (TE-to-WB) as well as sensitivity, specificity and overall diagnostic accuracy. 78.67% (59/75) of NGS results in the SCM group were interpretable, a significantly lower percentage than their corresponding TE and WB groups. This discrepancy manifests itself in intrinsically low quantity and poor integrity DNA from SCM. Subsequently, remarkable differences in full concordance rates (including mosaicism, and segmental aneuploidies) are seen as follows: 32.2% (SCM-to-WB, 19/59) and 69.33% (TE-to-WB, 52/75), (p . In such cases, full concordance rates were 27.27% (15/55) in SCM-to-WB, and, 76% (57/75) in TE-to-WB (p . Collectively, the NGS data from SCM also translated into lower sensitivities, Positive Predictive Value (PPV), Negative Predictive Value (NPV), overall diagnostic accuracies, and higher Negative Likelihood Ratio (NLR). Conclusions Our study reveals that DNA is detectable in the majority of SCM samples. Individual chromosomal aberration, such as segmental aneuploidy and mosaicism, can be quantitatively and qualitatively measured. However, TE still provides a more accurate and reliable high-throughput methodology for PGT-A. Meanwhile, cell-free DNA in SCM reporting lacks uniform diagnostic interpretations. Considering that this test is meant to determine which embryos are relegated to be discarded, PGT-A with cell-free DNA in SCM should not be permitted to be applied in routine clinical settings for diagnosis purpose.

Objective To evaluate the diagnostic efficiency of spent blastocyst culture medium (BCM) in noninvasive preimplantation genetic testing (niPGT) by comparing the karyotype concordance with corresponding inner cell mass (ICM) among initial trophectoderm (TE) biopsy, TE re-biopsy, and BCM sampling. Design Re-analysis aneuploid/mosaic blastocysts donated for research by couples. Setting Institutional in vitro fertilization center. Patient(s) A total of 12 couples donated their blastocysts, which had previously been diagnosed as aneuploid or mosaic by initial TE-biopsy preimplantation genetic testing for aneuploidy (PGT-A) for research. Intervention(s) A total of 26 frozen−thawed blastocysts were re-analyzed by TE re-biopsy, ICM biopsy, and the collection of spent BCM. Main Outcome Measure(s) Karyotype concordance rates. Result(s) For 23 embryos diagnosed as aneuploid by initial TE biopsy, 78.3% of initial TE samples, 87.0% of TE re-biopsies samples, and 78.3% of BCM samples were concordant with corresponding ICM samples, and for three mosaic embryos, the concordance rates with ICM of these three groups were 0%, 100%, and 100%, respectively. With the corresponding ICM result as the true result, sensitivity of both niPGT-A and initial TE were 100%; however, the false-positive rate (FPR) of initial TE was higher than that of niPGT-A (100% vs. 0). Conclusion(s) niPGT-A using BCM had diagnostic efficiency similar to that of TE-biopsy PGT-A. In the case of mosaic embryos, niPGT-A using BCM may be more reliable for predicting karyotypes of ICM than initial TE biopsy.

Recurrent implantation failure (RIF) is an intrigue condition during in vitro fertilization (IVF) cycles or intracytoplasmic sperm injection (ICSI) treatments. The purpose of this retrospective study is to explore the value of next-generation sequencing (NGS)-based preimplantation genetic testing for aneuploidy (PGT-A) of trophectoderm biopsy in the clinical outcomes for RIF patients with advanced age. A total of 265 RIF patients, who underwent 346 oocyte retrieval cycles and 250 PGT-A cycles, were classified as two groups according to the female age, including < 38 and ≥ 38 years old groups. The two groups were statistically comparable in baseline characteristics. The component of aneuploid embryos was significantly higher in advanced age group than in younger age group (68.9 vs 39.9%, P < 0.001). But there were no statistically significant differences in pregnancy rate (43.5 vs 64.7%), clinical pregnancy rate (39.1 vs 48.0%), implantation rate (39.1 vs 51.0%), and miscarriage rate (4.3 vs 7.8%) per embryo transfer (ET) between the two groups. Results suggest that the embryo-related factor plays a crucial role in RIF. Maternal age does not influence the implantation potential of euploid blastocysts. The NGS-based PGT-A involving trophectoderm biopsy is valuable for RIF patients of advanced age by improving their clinical outcomes. In conclusion, the NGS-based PGT-A involving trophectoderm biopsy may represent a valuable supplement to the current RIF management. Nonetheless, these findings should be further validated in a well-designed randomized controlled trial.

PURPOSE: To determine whether the blastocyst mitochondrial DNA (mtDNA) content is related to the miscarriage rate in patients undergoing single euploid frozen embryo transfer (SEFET). METHODS: A total of 355 single euploid frozen embryo transfer cycles were studied retrospectively between April 2017 and December 2018. A trophectoderm biopsy was performed on day 5/6 blastocysts. Post next-generation sequencing (NGS), the mtDNA content was calculated as the ratio of mitochondrial DNA over nuclear DNA, and the association between blastocyst mtDNA content and miscarriage rate was evaluated. RESULT(S): Three hundred fifty-five euploid blastocysts were selected for SEFET in 314 patients with an average age of 33.7 ± 5.6 years; 255 were biopsied on day 5 (71.8%) and 100 on day 6 (28.2%). Frozen embryo transfer (FET) was performed either in a hormone replacement therapy (HRT) cycle (71.8%; n = 255) or in a natural cycle (NC) (28.2%; n = 100). A pregnancy rate of 66.2% (235/355) was obtained with clinical pregnancy and miscarriage rates of 52.4% (n = 186) and 5.6% (n = 20), respectively. There was no significant difference neither between the blastocyst mtDNA content of pregnant and nonpregnant patients (27.7 ± 9.2 vs. 29.4 ± 8.6, P = 0.095) nor between patients with a clinical pregnancy and miscarriage (30.5 ± 9.3 vs. 27.3 ± 9.2, P = 0.136). Multivariate logistic regression analysis showed the same nonsignificant relationship, except for the miscarriage rate and BMI (OR 1.149, 95% CI 1.03–1.28; P = 0.012). CONCLUSION(S): Mitochondrial DNA content is unable to predict the miscarriage of implanted human euploid blastocysts. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10815-020-02050-8.

Purpose The purpose of this study is to summarize the clinical outcomes of apparently balanced chromosome rearrangement (ABCR) carriers in preimplantation genetic testing (PGT) cycles by next-generation sequencing following microdissecting junction region (MicroSeq) to distinguish non-carrier embryos from balanced carriers. Methods A retrospective study of 762 ABCR carrier couples who requested PGT for structural rearrangements combined with MicroSeq at the Reproductive and Genetic Hospital of CITIC-Xiangya was conducted between October 2014 and October 2019. Results Trophectoderm biopsy was performed in 4122 blastocysts derived from 917 PGT-SR cycles and 3781 blastocysts were detected. Among the 3781 blastocysts diagnosed, 1433 (37.9%, 1433/3781) were balanced, of which 739 blastocysts were carriers (51.57%, 739/1433) and 694 blastocysts were normal (48.43%, 694/1433). Approximately 26.39% of cycles had both carrier and normal embryo transfer, and the average number of biopsied blastocysts was 6.7. In the cumulative 223 biopsied cycles with normal embryo transfer, all couples chose to transfer the normal embryos. In the 225 cycles with only carrier embryos, the couples chose to transfer the carrier embryos in 169/225 (75.11%) cycles. A total of 732 frozen embryo transfer cycles were performed, resulting in 502 clinical pregnancies. Cumulatively, 326 babies were born; all of these babies were healthy and free of any developmental issues. Conclusion Our study provides the first evaluation of the clinical outcomes of a large sample with ABCR carrier couples undergoing the MicroSeq-PGT technique and reveals its powerful ability to distinguish between carrier and non-carrier balanced embryos.

Background Recently, the Japan Society of Obstetrics and Gynecology initiated a clinical study of preimplantation genetic test for aneuploidy. There will be a great need for a standardized embryo biopsy technique in Japan. However, the gold standard trophectoderm (TE) biopsy procedure has not been established, and this review outlines the clinical use of TE biopsy. Methods Based on literature, the method and associated techniques for TE biopsy, a dissection method of TE cells from blastocysts, were investigated. Main findings Two TE biopsy methods are used, namely assisted hatching (herniating) and non‐assisted hatching (direct suction); however, it is not clear which of these methods is superior. It is critical to understand whether the flicking or pulling method is beneficial. Conclusion Non‐assisted hatching biopsy method may cause blastocyst collapse with a higher probability, and it may extend the biopsy time. The biopsy procedure should be performed within 3 minutes, and thus direct TE suction may have greater disadvantages. It is a fact that pulling method of TE dissection with laser pulse is simple; however, excess laser shots may induce a higher frequency of mosaicism. It is important to understand that each technique of TE biopsy has benefits and disadvantages.

Embryo aneuploidies may be responsible for implantation failures, miscarriages and affects IVF outcomes. A variety of technologies have been implemented to individuate euploid embryos in IVF treatments, which is named preimplantation genetic testing for aneuploidies (PGT-A). According to this strategy, a better embryo selection should increase IVF results. In reality, several issues remain unaddressed including the sampling strategy, involving the test outcomes, and the frequent occurrence of embryo mosaicism, affecting the criteria for selection of supposed viable embryos and possibly posing an ethical dilemma. Safety issues are in place, including perinatal and postnatal consequences of embryo sampling and the epigenetic weaknesses from a prolonged in vitro culture, necessary for trophectoderm biopsy. On the other side, chromosome number mistakes are progressively recognized as physiologic events in the early pre-implantation embryo with many corrective mechanisms in place and their destiny in the post-implantation development is unclear. Accordingly, the increasing precision of the diagnostic tools should be used to investigate the effect of such interventions within rigorous research programs in the sake of improved clinical outcomes. Meantime the diagnosis of embryo aneuploidies in IVF cycles should be considered as a research tool and systematic implementation in clinical practice may appear unjustified.

This study was conducted to assess the effectiveness of a new trophectoderm (TE) biopsy method that does not require prior opening of the zona pellucida at the blastocyst stage.TE biopsy was conducted using a modified extrusion method for embryos during the cleavage stage. In this method, culture medium was injected into the perivitelline space to help extrude TE cells from the zona pellucida before TE biopsy.Our extrusion method preserves the embryo culture environment until immediately before biopsy because it does not require opening of the zona pellucida prior to TE biopsy. Furthermore, this method does not require a waiting time for blastocyst hatching after laser irradiation, thereby minimizing damage to the embryos and maintaining the time schedule of culture operations.TE biopsy using this novel extrusion method may be useful in various applications, including the collection of TE cells for next-generation sequencing analysis.

Objective To study whether embryonic cell-free DNA (cfDNA) in spent blastocyst media is representative of the chromosomal constitution of a blastocyst. Design Pilot prospective blinded study. Setting In vitro fertilization center and genetics laboratory. Patient(s) A total of 115 trophectoderm (TE) biopsies and spent blastocyst media (SBM) from 46 patients with ages ranging from 32 to 46 years, whose indications for preimplantation genetic testing of aneuploidy (PGT-A) were advanced maternal age, recurrent miscarriage, or recurrent implantation failure. Interventions(s) Spent blastocyst media collection and TE biopsy. Main Outcome Measure(s) Concordance rates, sensitivity, and specificity between TE biopsies and SBM. Clinical outcomes in cases with euploid TE biopsies and euploid SBM compared with cases with euploid TE and aneuploid SBM. Result(s) In general, the total concordance rate for ploidy and sex was 78.7%, and sensitivity and specificity were 94.5% and 71.7%, respectively. A significant increase for all parameters was observed for day 6/7 samples compared with day 5 samples, with day 6/7 samples showing total concordance for ploidy and sex of 84%, and sensitivity and specificity of 95.2% and 82.1%, respectively. Ongoing implantation rates in euploid TE/euploid SBM showed a threefold increase compared with euploid TE/aneuploid SBM (52.9% vs. 16.7%, respectively), without reaching significant differences. Interestingly, no miscarriages were observed when TE and SBM were euploidy concordant. Conclusion(s) These results offer a better understanding of the dynamics of cfDNA during embryo development and despite more basic research being needed, they are reassuring to consider in the future this noninvasive approach as an alternative to TE biopsy for PGT-A.

A systematic review of the literature showed that trophectoderm biopsy could assist in the selection of healthy embryos for uterine transfer without affecting implantation rates. However, previous studies attempting to establish the relationship between trophectoderm gene expression profiles and implantation competency using either microarrays or RNA sequencing strategies, were not sufficiently optimized to handle the exceptionally low RNA inputs available from biopsied material. In this pilot study, we report that differential gene expression in human trophectoderm biopsies assayed by an ultra-sensitive next generation RNA sequencing strategy could predict blastocyst implantation competence. RNA expression profiles from isolated human trophectoderm cells were analysed with established clinical pregnancy being the primary endpoint. Following RNA sequencing, a total of 47 transcripts were found to be significantly differentially expressed between the trophectoderm cells from successfully implanted (competent) versus unsuccessful (incompetent) blastocysts. Of these, 36 transcripts were significantly down-regulated in the incompetent blastocysts, including Hydroxysteroid 17-Beta Dehydrogenase 1 (HSD17B1) and Cytochrome P450 Family 11 Subfamily A Member 1 (CYP11A1), while the remaining 11 transcripts were significantly up-regulated, including BCL2 Antagonist/Killer 1 (BAK1) and KH Domain Containing 1 Pseudogene 1 (KHDC1P1) of which the latter was always detected in the incompetent and absent in all competent blastocysts. Ontological analysis of differentially expressed RNAs revealed pathways involved in steroidogenic processes with high confidence. Novel differentially expressed transcripts were also noted by reference to a de novo sequence assembly. The selection of the blastocyst with the best potential to support full-term pregnancy following single embryo transfer could reduce the need for multiple treatment cycles and embryo transfers. The main limitation was the low sample size (N = 8). Despite this shortcoming, the pilot suggests that trophectoderm biopsy could assist with the selection of healthy embryos for embryo transfer. A larger cohort of samples is needed to confirm these findings. Abbreviations: AMA: advanced maternal age; ART: assisted reproductive technology; CP: clinical pregnancy; DE: differential expression; FDR: false discovery rate; IVF: in vitro fertilization; LD PCR: long distance PCR; qRT-PCR: quantitative real-time PCR; SET: single embryo transfer; TE: trophectoderm

Study question To explore the effect of chromosomal mosaicism detected in preimplantation genetic testing (PGT-A) on prenatal and postnatal outcome of mosaic embryo pregnancies Summary answer No significant difference between euploid and mosaic embryos was observed in terms of weeks of gestation, average weight, and developmental defect of the babies born What is known already Mosaic embryos have the potential to implant and develop into healthy babies. Transfer of these embryos is now offered as an option for women who undergo IVF resulting in no euploid embryos. While, prenatal diagnosis has shown the depletion of chromosomal mosaicism in mosaic embryos, several concerns remain. For instance, the direct effects of different kind of mosaicism on prenatal/postnatal outcome and the possibility that intra-biopsy mosaicism in the TE is a poor predictor of the ploidy status of the ICM. Thus, there is certainly a need for comprehensive analyses of obstetrical and neonatal outcome data of transferred mosaic embryos. Study design, size, duration Compiled analysis from multicenter data on transfers of mosaic embryos (n = 1,000) and their outcome, with comparison to a euploid control group (n = 5,561). To explore the effect of embryonic mosaicism on newborns, we matched mosaic embryos resulting in a birth with a euploid embryo by a series of parameters (maternal age, embryo morphology, and indication for PGT-A). Prenatal tests and birth characteristics of > 200 neonates from mosaic embryo transfers were compared to > 200 euploid embryos. Participants/materials, setting, methods PGT-A was performed on blastocyst-stage embryos with 24-Chromosome whole genome amplification (WGA)-based Next Generation Sequencing (NGS). In accordance with established guidelines, embryos were categorized as mosaic when PGT-A results indicated 20-80% aneuploid content. Prenatal testing where performed in 30% of pregnancies with amniocentesis, 4% did an extra analysis for potential UPD for the suspected mosaic chromosome, and an additional 16% performed chorionic villus sampling (CVS) and 9.5% performed noninvasive prenatal testing (NIPT). Main results and the role of chance Of the 465 mosaic embryos that implanted, about 20% miscarried, and out of those, 75% were early spontaneous abortions. Of the pregnancies, 3 out of 368 were stillborn (2 out of them were twins that were extremely premature at 23 weeks, and the other died during pregnancy from a heart defect). The remaining 99% of those have been born or are late ongoing pregnancies at the time of analysis. Prenatal tests were performed in > 200 pregnancies and the vast majority tested normal. All 5 abnormal cases were amniocentesis tests showing microdeletions or insertions of sizes smaller than the resolution used during PGT-A, so they were unrelated to the mosaicism detected with PGT-A. In fact, in none of the cases did the prenatal test reflect the mosaicism detected at the embryonic stage. Matching each of the 162 mosaic embryos resulting in a birth with a euploid embryo, we found that the length of gestation was similar on average, and so was the average weight of the babies at birth. We also gathered information on the routine physical examination performed on babies at birth, and of those 162 babies from mosaic embryo transfers, none had obvious developmental defects or gross abnormalities. Limitations, reasons for caution Even though newborns resulting from mosaic embryo transfers in this study invariably appeared healthy by routine examination, concerns for long-term health cannot yet be entirely dispelled. The question must therefore be carefully considered by each clinic and patient situation. Wider implications of the findings Prenatal testing of > 200 pregnancies from mosaic embryo transfers showed no incidence of mosaicism that matched the PGT-A findings, indicating the involvement of self-corrective mechanisms. Pregnancy and obstetric data indicates that mosaic embryos prevailing through gestation and birth have similar chromosomal and physiological health compared to euploid embryos. Trial registration number none

Study question The objective of this study is to evaluate the effect of trophectoderm (TE) biopsy on different blastocyst stages and its clinical outcome. Summary answer Our results showed that TE biopsy significantly reduced the clinical outcome of fully hatched blastocyst. What is known already: TE biopsy is a method widely practiced to harvest cells to determine the chromosomal constitution of a blastocyst, ensuring higher implantation and healthy pregnancies. The effect on clinical outcome after transferring blastocysts biopsied at different blastocysts stages has not been extensively studied. Study design, size, duration This retrospective study was conducted from January 2017 until July 2019 at Alpha IVF & Women’s Specialists. Following laser assisted hatching on day 3, TE biopsy was performed on unhatched, hatching and fully hatched day–5 blastocysts. A total of 1,020 single euploid blastocysts transfer (SBT) were performed. The average maternal age was 31.7. Implantation rates (IR) were evaluated for all stages of hatching (Unhatched: BG3 & 4; hatching: BG5; fully hatched: BG6). Participants/materials, setting, methods Laser assisted hatching (Hamilton Thorne Bioscience, USA) was performed on day–3 and subsequently cultured to blastocyst-stage. Different hatching stages were observed using embryoscope time-lapse system (Vitrolife, Sweden) and were recorded. Day–5 blastocysts with at least BG3BB grade (Gardner’s System) were selected for TE biopsy and the biopsied cells were sent for preimplantation genetic testing for aneuploidy (PGT-A) using Next-Generation Sequencing (Life Technologies, USA). All blastocysts were vitrified and warmed using the Cryotec Method (Cryotech, Japan). Main results and the role of chance: All 1,020 blastocysts survived post-warmed (post-warm survival rate= 100%) and were transferred in frozen transfer cycles. TE biopsy performed on unhatched blastocysts showed a comparable IR to hatching blastocysts (60.0% [15/25] and 65.2% [627/961]). While fully hatched blastocysts (44.12% [15/34]) show a significantly lower IR when compared to hatching blastocysts (65.2% [627/961]), no significant difference was seen when comparing unhatched blastocysts to fully hatched blastocysts (60.0% [15/25] and 44.12% [15/34]; p = 0.2949). Limitations, reasons for caution The sample size was comparatively smaller in the unhatched and fully hatched group than the hatching group. Further studies with a larger sample size is recommended to ascertain the clinical outcome. Since this is a retrospective study and biopsy was done by different embryologists, the biopsy technique was not controlled. Wider implications of the findings: To achieve higher clinical pregnancy, it is recommended to perform TE biopsy before the blastocysts is fully hatched. Trial registration number Not applicable

Study question Is it useful or beneficial to perform Assisted Hatching (AH) on D + 3 previously to biopsy for PGT-A on blastocyst stage on D + 5? Summary answer The routine use of AH on D + 3 to facilitate the embryo biopsy on D + 5 could negatively influence the development of the embryos to blastocyst stage. What is known already The blastocyst stage is the optimal stage for performing biopsies for PGT-A, which has been reported as a key factor determining the growing clinical application of this strategy worldwide. For trophectoderm (TE) biopsy, laser-assisted drilling is used to create a zona opening on D + 3 or D + 5 of development. The method of zona opening on D + 3 allows some of the TE cells to herniate during blastocyst formation and expansion, which facilitates the biopsy process. However, this method may result in herniation of inner cell mass cells instead of TE or maybe could affect the development of the embryo to blastocyst stage. Study design, size, duration A total of 100 PGT-A cycles were performed in 2019 and 2020. In 78 of them laser-assisted drilling was used to create a zona opening on D + 5 only in those embryos which arrived to blastocyst stage for TE biopsy (Group No-AH). In 22 cycles the same drilling was achieved on D + 3 in all embryos, independently of their quality (Group AH). The average of embryos per cycle in each group was 5 and 4.3 respectively. Participants/materials, setting, methods A total of 100 PGT-A cycles coming from 65 patients were studied. The average of the age of the patients was 40.83 (SD 3.45) in the group No-AH vs 42.18 (SD 3.42) in the Group AH (p = 0.108), so the age was not a determining factor for the development of the embryos. We analyzed by χ 2 test differences between groups on fertilization rates, number of embryos, development to blastocyst stage, euploidy and pregnancy rates. Main results and the role of chance The fertilization rate was 74.79% (No-AH group) and 68.53% (AH group) with no significative statistical differences (p = 0.12). In the No-AH group, the TE biopsy was performed on D + 5 in 63 cycles (81%). In the AH group, 41% of cycles didn’t reach the blastocyst stage, obtaining statistical differences between groups (p = 0.035). We found also significant differences in the number of cycles with biopsied blastocyst when we had 1 to 6 embryos/cycle on D + 3 between groups (p = 0.002), without obtaining any blastocyst to be diagnosed in 53% of the cycles in AH group vs 27% in No-AH group. When the number of embryos on D + 3 per cycle was > 6, at least 1 embryo reached the blastocyst stage in both groups, although this number was higher in No-AH group. The rate of biopsied blastocysts was significantly higher in the No-AH group compared to the AH group (46.61 vs 34.69) with a p = 0.031. The rate of euploid embryos analyzed was 23.30% in the No-AH group compared to 29.41% in the AH group, although no significant differences were found (p = 0.44) between groups. In the No-AH group, a clinical pregnancy rate of 52.94% was obtained (n = 34) vs 50% in the AH group (n = 4) (p = 0.91). Limitations, reasons for caution We have recently started to perform AH on D + 3, so the number of cases is smaller than No-AH group. We use a time lapse incubator in all cases, so in the No-AH the culture dish is changed, disturbing the stable incubation environment, while in the other group it is not. Wider implications of the findings: The use of AH on D + 3 in order to facilitate the TE biopsy on D + 5 could affect negatively the development of the embryos to blastocyst stage. Its routine use should be avoided based on laboratory workload, mainly if the patient has less than 7 embryos at D + 3. Trial registration number Not applicable

Study question Does laser use during trophectoderm biopsy affect biopsy results on prehatching embryos with regard to mosaicism ? Summary answer According to our findings laser usage during trophectoderm biopsy increases mosaic results on next generation sequencing (NGS) irrespective of embryo quality. What is known already Chromosomal mosaicism, which is a result of mitotic errors after fertilization, is defined as the presence of karyotypically distinct cell lines within an embryo. The introduction of NGS made it possible to detect chromosomal mosaicism at levels as low as 20%. The incidence of mosaicism is highly variable between clinics which reported the incidences between 4-32%. Apart from the biological reasons, there are also various technical factors that may impact the incidence of mosaicism. One of the most emphasized factors is the trophoectoderm biopsy technique. Laser usage and number of laser pulses may cause excessive heat during the procedure Study design, size, duration The mosaicism ratio in embryos in which trophectoderm biopsy was performed with or without laser, between January 2017 December 2020 in Istanbul Memorial Hospital (IMH) were examined retrospectively. A total of 13002 embryos were analyzed. A subgroup analysis was also performed regarding mosaicism ratios in different embryo qualities. Blastocysts were classified according to Gardner’s classification and classified as follows: top quality-TQ (4AA,5AA,6AA), good quality-GQ (3AA, 4,5,6AB,BA) and moderate quality-MQ (3,4,5 BB). Participants/materials, setting, methods The biopsy samples of the cases who had PGT-A in IMH between 2017-2020 were evaluated by NGS method. This method enables the identification of embryos with 20% to 80% mosaicism. The study assessed whether there was an increase in the embryos with mosaic results due to the use of laser during biopsy. The effects of laser use among the TQ (4AA,5AA,6AA), GQ (3AA, 4,5,6AB,BA) and MQ (3,4,5 BB) groups according to Garder classification were analyzed. Main results and the role of chance Trophectoderm biopsy was applied on 13002 embryos within the specified period. During biopsy in 5088 embryos laser was used and in 7843 embryos laser was not used, and biopsy was performed mechanically (flicking method). After observing the biopsy results, 945/5088 (18.5%) of the laser applied embryos; and 1087/7914 (13.7%) of laser not applied embryos were defined as mosaic(p 0.05) Limitations, reasons for caution The retrospective nature of the data is the main limitation of the study. On the other hand, the large number of NGS based PGT-A tested TQ and GQ embryos from a single center and resuts from single laboratory. However, further studies are required to corroborate our findings. Wider implications of the findings Laser dependent heat effect may increase mosaicism. To reduce the cell damage, teasing of cells should be avoided and a minimum number of laser pulses should be used in order to avoid excessive heat and contact points should be preferably confined to cell junctions Trial registration number None

BackgroundChromosomal mosaicism and aneuploidies are routine phenomena throughout human pre- and post-implantation development. The benefit of implanting mosaicism or aneuploidies is still controversial. The purposes of the study are to investigate the developmental potential of embryos with chromosomally segmental or mosaic abnormalities, and whether precise Next Generation Sequencing (NGS) resolution would reduce the development of an abnormal embryo in preimplantation genetic testing (PGT) cycles.MethodsThe peripheral blood of 17 PGT babies were collected for single nucleotide polymorphism (SNP) array and were compared with trophectoderm (TE) biopsy results at different NGS resolutions.Results76.5% (13/17) of babies’ peripheral blood chromosome analysis was consistent with 10Mb TE biopsies and 58.8% (10/17) of babies’ analysis was consistent with 4Mb TE biopsies. 2 babies who had euploid TE showed abnormal peripheral blood chromosome analysis. 17.6% (3/17) embryos with aberrant TE biopsies produced healthy babies. Although the sensitivity of 10Mb was lower than 4Mb (25% vs. 50%), the specificity (100% vs. 76.9%), PPV (100% vs. 40%) and diagnostic accuracy (82.4% vs. 70.6%) of 10Mb showed better results than 4Mb.Conclusion(s)The chromosomal results between peripheral blood samples and TE biopsies of born babies are not completely congruent. Aneuploid and mosaic embryos have potential to produce healthy babies, whereas normal embryos also have chance to produce babies with chromosomal abnormalities. In spite of low sensitivity of both resolutions, 10Mb has higher specificity, PPV and diagnostic accuracy than 4Mb. It is suggested that TE biopsy be analyzed in both 10Mb and 4Mb resolutions to uncover severely adverse chromosomal aberrations but use 10Mb resolution to guide transfer.Trial registrationThe study was retrospectively registered in the Chinese Clinical Trial Registry (ChiCTR2100042522).

To review the cumulative outcome of pre-implantation genetic diagnosis (PGD) cycles performed for prevention of sickle cell disease (SCD). Couples referred for PGD for SCD between April 2012 and October 2017 were included. Ovarian stimulation was performed using a short gonadotrophin-releasing hormone (GnRH) antagonist protocol and follicle-stimulating hormone injections. The GnRH agonist was used to trigger oocyte maturation. Oocytes were fertilised using intracytoplasmic sperm injection. Trophectoderm biopsy was performed on day 5 or 6 followed by vitrification. Genetic testing was done using pre-implantation genetic haplotyping. A total of 60 couples started 70 fresh PGD cycles (mean 1·2 cycles/couple) and underwent a total of 74 frozen-embryo-transfer (FET) cycles (mean 1·3 FET/couple). The mean (SD) female age was 33 (4·4) years and the mean (SD) anti-mullerian hormone level was 22·9 (2·8) pmol/l. The cumulative live-birth rate was 54%/PGD cycle started and 63%/couple embarking on PGD. The rate of multiple births was 8%. The cumulative outcome of PGD treatment for prevention of SCD transmission is high and PGD treatment should be offered to all at-risk couples.

Our objective was to evaluate and characterize the extent and patterns of worldwide usage of preimplantation genetic screening (PGS) among the assisted reproductive technique community. A prospective, web-based questionnaire with questions relating to practices of, and views on, PGS was directed to users and non-users of PGS. A total of 386 IVF units from 70 countries conducting 342,600 IVF cycles annually responded to the survey. A total of 77% of respondents routinely carry out PGS in their clinics for a variety of indications: advanced maternal age (27%), recurrent implantation failure (32%) and recurrent pregnancy loss (31%). Few (6%) offer PGS to all their patients. In most cycles (72%), trophectoderm biopsy is carried out and either array-comparative genomic hybridization (59%) or next-generation sequencing (16%) are used for genetic analysis. Only 30% of respondents regard PGS as clearly evidenced-based, and most (84%) believe that more randomized controlled trials are needed to support the use of PGS. Despite ongoing debate and lack of robust evidence, most respondents support the use of PGS, and believe that it may aid in transferring only euploid embryos, thereby reducing miscarriage rates and multiple pregnancies, increasing live birth rates and reducing the risk of aneuploid pregnancies and births.

STUDY QUESTION What are the trends and developments in preimplantation genetic testing (PGT) in 2013–2015 as compared to previous years? SUMMARY ANSWER The main trends observed in the retrospective data collections 2013–2015, representing valuable data on PGT activity in (mainly) Europe, are the increased application of trophectoderm biopsy at the cost of cleavage stage biopsy and the continuing expansion of comprehensive testing technology in PGT for chromosomal structural rearrangements and for aneuploidies (PGT-SR and PGT-A). WHAT IS KNOWN ALREADY Since it was established in 1997, the ESHRE PGT Consortium has been collecting data from international PGT centres. To date, 15 data sets and an overview of the first 10 years of data collections have been published. STUDY DESIGN, SIZE, DURATION Collection of (mainly) European data by the PGT Consortium for ESHRE. The data for PGT cycles performed between 1 January 2013 and 31 December 2015 were provided by participating centres on a voluntary basis. For the collection of cycle, pregnancy and baby data, separate, pre-designed MS Excel tables were used. PARTICIPANTS/MATERIALS, SETTING, METHODS Data were submitted by 59, 60 and 59 centres respectively for 2013, 2014 and 2015 (full PGT Consortium members). Records with incomplete or inconsistent data were excluded from the calculations. Corrections, calculations, figures and tables were made by expert co-authors. MAIN RESULTS AND THE ROLE OF CHANCE For data collection XVI/XVII/XVIII, 59/60/59 centres reported data on 8164/9769/11 120 cycles with oocyte retrieval: 5020/6278/7155 cycles for PGT-A, 2026/2243/2661 cycles for PGT for monogenic/single gene defects, 1039/1189/1231 cycles for PGT-SR and 79/59/73 cycles for sexing for X-linked diseases. From 2013 until 2015, the uptake of biopsy at the blastocyst stage was mainly observed in cycles for PGT-A (from 23% to 36%) and PGT-SR (from 22% to 36%), alongside the increased application of comprehensive testing technology (from 66% to 75% in PGT-A and from 36% to 58% in PGT-SR). LIMITATIONS, REASONS FOR CAUTION The findings apply to the 59/60/59 participating centres and may not represent worldwide trends in PGT. Data were collected retrospectively and no details of the follow-up on PGT pregnancies and babies born were provided. WIDER IMPLICATIONS OF THE FINDINGS Being the largest data collection on PGT worldwide, detailed information about ongoing developments in the field is provided. STUDY FUNDING/COMPETING INTEREST(S) The study has no external funding and all costs are covered by ESHRE. There are no competing interests declared. TRIAL REGISTRATION NUMBER N/A.

Preimplantation genetic diagnosis technique has been changed in the past few years from a cytogenetic technique that analyzes a limited number of chromosomes targeting only one or two points to a molecular diagnostic technique that permits to analyze a complete comprehensive chromosome screening targeting thousands of points in each chromosome. That, together with a huge increase in culture conditions on the laboratory (low pressure oxygen incubators, single culture medium, vitrification protocols, and time-lapse), has change the rules of the game. In this chapter, we analyze new issues to face: what to do with mosaicism, fresh or frozen embryo transfer, segmental aneuploidies, etc.

Preimplantation genetic testing for monogenic diseases (PGT-M) was introduced in assisted reproductive programs as a preventive test for patients identified as having high risk for inherited Mendelian conditions. PGT-M is considered an early prenatal diagnosis for monogenic conditions with the advantage of avoiding or limiting the decisions of termination of pregnancy as only genetically normal embryos are transferred to the uterus. In recent years, the implementation of new methods and improvements to in vitro fertilization programs, such as vitrification and trophectoderm biopsy, has enabled the achievement of better clinical results after embryo testing. In parallel, advancements in genetic knowledge, technology, and molecular methods have allowed for the possibility of offering an ever-expanded list of conditions to be tested in embryo biopsies. In this chapter, this evolution is described and current PGT strategies where testing for monogenic and aneuploidy screening preimplantation genetic testing for aneuploidies from the same embryo sample is the standard for all PGT-M indications are explained.

The transition in biopsy timing from blastomere to trophectoderm biopsy has led to a remarkable decrease in the percentage of undiagnosed blastocysts. However, patients with few or no euploid blastocysts can be affected by this residual percentage of diagnosis failure. The aim of this study is to assess whether blastocyst rebiopsy and revitrification is an efficient and safe procedure to be applied in cases of no results after analysis. Fifty-three patients agreed to the warming of 61 blastocysts to perform a second biopsy and PGT-A by aCGH. Only 75.4% of the blastocysts survived, reexpanded, and could be rebiopsied. After the second biopsy and analysis, 95.6% of the blastocysts were successfully diagnosed with an euploidy rate of 65.9%. Eighteen euploid blastocysts were warmed and transferred to 18 patients with a 100% survival and reexpansion rate. Seven clinical pregnancies have been achieved with 4 live births, 1 ongoing pregnancy, and 2 miscarriages. Thus, although few transfers of rebiopsied and revitrified blastocysts have been performed till date, our preliminary results show that this approach is efficient and safe to be applied for undiagnosed blastocysts, as it ultimately allows the transfer of euploid blastocysts and good clinical outcomes.

Objective: The goal of the present study was to investigate the rate of chromosomal aneuploidies in surplus embryos after sex determination at the cleavage stage. Then, the same chromosomal aneuploidies were evaluated in blastocysts after extended culture.Methods: Sixty-eight surplus embryos were biopsied at the cleavage stage and incubated for an additional 3 days to allow them to reach the blastocyst stage. The embryos were reanalyzed via fluorescence in situ hybridization (FISH) to examine eight chromosomes (13, 15, 16, 18, 21, 22, X, and Y) in both cleavage-stage embryos and blastocysts.Results: Although the total abnormality rate was lower in blastocysts (32.35%) than in cleavage-stage embryos (45.58%), the difference was not significant (p=0.113). However, when we restricted the analysis to autosomal abnormalities, we observed a significant difference in the abnormality rate between the cleavage-stage embryos (44.11%) and the blastocysts (17.64%, p=0.008). A higher rate of sex chromosomal abnormalities was also observed in cleavage-stage embryos (29.4%) than in blastocysts (14.70%, p=0.038). Conclusion: The data indicated that embryo biopsy should be conducted at the blastocyst stage rather than the cleavage stage. The results also emphasized that examination of common chromosomal aneuploidies apart from sex selection cycles can be conducted in the blastocyst stage with the FISH method.

The majority of embryos created through in vitro fertilization (IVF) do not implant. It seems plausible that rates of implantation would improve if we had a better understanding of molecular factors affecting embryo competence. Currently, the process of selecting an embryo for uterine transfer uses an ad hoc combination of morphological criteria, the kinetics of development, and genetic testing for aneuploidy. However, no single criterion can ensure selection of a viable embryo. In contrast, RNA-sequencing (RNA-seq) of embryos could yield high-dimensional data, which may provide additional insight and illuminate the discrepancies among current selection criteria. Recent advances enabling the production of RNA-seq libraries from single cells have facilitated the application of this technique to the study of transcriptional events in early human development. However, these studies have not assessed the quality of their constituent embryos relative to commonly used embryological criteria. Here, we perform proof-of-principle advancement to embryo selection procedures by generating RNA-seq libraries from a trophectoderm biopsy as well as the remaining whole embryo. We combine state-of-the-art embryological methods with low-input RNA-seq to develop the first transcriptome-wide approach for assessing embryo competence. Specifically, we show the capacity of RNA-seq as a promising tool in preimplantation screening by showing that biopsies of an embryo can capture valuable information available in the whole embryo from which they are derived. Furthermore, we show that this technique can be used to generate a RNA-based digital karyotype and to identify candidate competence-associated genes. Together, these data establish the foundation for a future RNA-based diagnostic in IVF.

Blastocyst biopsy is performed to obtain a reliable genetic diagnosis during IVF cycles with preimplantation genetic testing. Then, the ideal workflow entails a safe and efficient vitrification protocol, due to the turnaround time of the diagnostic techniques and to transfer the selected embryo(s) on a physiological endometrium in a following natural cycle. A biopsy approach encompassing the sequential opening of the zona pellucida and retrieval of 5-10 trophectoderm cells (ideally 7-8) limits both the number of manipulations required and the exposure of the embryo to sub-optimal environmental conditions. After proper training, the technique was reproducible across different operators in terms of timing of biopsy (~8 min, ranging 3-22 min based on the number of embryos to biopsy per dish), conclusive diagnoses obtained (~97.5%) and live birth rates after vitrified-warmed euploid blastocyst transfer (>40%). The survival rate after biopsy, vitrification and warming was as high as 99.8%. The re-expansion rate at 1.5 h from warming was as high as 97%, largely dependent on the timing between biopsy and vitrification (ideally ≤30 min), blastocyst morphological quality and day of biopsy. In general, it is better to vitrify a collapsed blastocyst; therefore, in non-PGT cycles, laser-assisted artificial shrinkage might be performed to induce embryo collapse prior to cryopreservation. The most promising future perspective is the non-invasive analysis of the IVF culture media after blastocyst culture as a putative source of embryonic DNA. However, this potential avant-garde is still under investigation and a reliable protocol yet needs to be defined and validated.

IntroductionTo report the first case of live birth after double trophectoderm biopsy before and after vitrification on the same cohort of blastocyst in our knowledge.DesignCase report.PatientA 36-year-old female with a history of 13 years of active married life for treatment of infertility.Main outcome measureLive birth after double trophectoderm biopsy.ResultsDouble biopsy pre- and postvitrification and its positive outcome.ConclusionPreimplantation genetic screening and diagnosis (PGS/PGD), though an invasive procedure on the embryos, when done meticulously would not dampen the implantation potential of the embryo and second biopsy could be a feasible option to salvage embryos with inconclusive or suspected false-positive PGS/PGD reports.How to cite this articleBhadarka H, Patel NH, Jadeja YD, Patel KB, Patel NH, Patel MN. First Case of Successful Implantation and Live Birth after Double Trophectoderm Biopsy before and after Vitrification on the Same Cohort of Blastocyst. Int J Infertil Fetal Med 2017;8(3):120-124.

Purpose The aim of this study is to investigate the minimum number of blastocysts for biopsy to increase the likelihood of obtaining at least one normal/balanced embryo in preimplantation genetic diagnosis (PGD) for translocation carriers. Methods This blinded retrospective study included 55 PGD cycles for Robertsonian translocation (RT) and 181 cycles for reciprocal translocation (rcp) to indicate when only one of the couples carried a translocation. Single-nucleotide polymorphism microarray after trophectoderm biopsy was performed. Results Reliable results were obtained for 355/379 (93.7 %) biopsied blastocysts in RT group and 986/1053 (93.6 %) in rcp group. Mean numbers of biopsied embryos per patient, normal/balanced embryos per patient, and mean normal/balanced embryo rate per patient were 7.4, 3.1, and 40.7 % in RT group and 8.0, 2.1, and 27.3 %, respectively, in rcp group. In a regression model, three factors significantly affected the number of genetically transferrable embryos: number of biopsied embryos (P = 0.001), basal FSH level (P = 0.040), and maternal age (P = 0.027). ROC analysis with a cutoff of 1.5 was calculated for the number of biopsied embryos required to obtain at least one normal/balanced embryo for RT carriers. For rcp carriers, the cutoff was 3.5. The clinical pregnancy rate per embryo transfer was 44.2 and 42.6 % in RT and rcp groups (P = 0.836). Conclusions The minimum numbers of blastocysts to obtain at least one normal/balanced embryo for RT and rcp were 2 and 4 under the conditions of female age

Blastocyst culture and transfer for human IVF has resulted in an increase in implantation and live birth rates and successfully facilitated the move toward single embryo transfer. Initially this was attained through the development and introduction of sequential media, designed to mirror the changing nutritional environments within the oviduct and uterus. Sequential media underwent years of development using several animal models and were validated by the transfer of thousands of embryos prior to its clinical evaluation and subsequent use in prospective randomized trials. Such media have proved themselves to be highly effective in clinical IVF and have facilitated the move to trophectoderm biopsy and vitrification of the embryo. However, successful embryo culture goes beyond the choice of media and requires an holistic approach, whereby all facets of the embryo culture system, especially the oxygen concentration, are optimized.

(Abstracted from Hum Reprod 2019;34(4):758–769)The development of preimplantation genetic testing (PGT-A) in conjunction with assisted reproductive technology has led to valuable analysis of the high frequency of chromosomal abnormalities observed in human embryos. While mosaic embryos may lead to h

STUDY QUESTION: Does preimplantation genetic testing for aneuploidy (PGT-A) by comprehensive chromosome screening (CCS) of the first and second polar body to select embryos for transfer increase the likelihood of a live birth within 1 year in advanced maternal age women aged 36-40 years planning an ICSI cycle, compared to ICSI without chromosome analysis? SUMMARY ANSWER: PGT-A by CCS in the first and second polar body to select euploid embryos for transfer does not substantially increase the live birth rate in women aged 36-40 years. WHAT IS KNOWN ALREADY: PGT-A has been used widely to select embryos for transfer in ICSI treatment, with the aim of improving treatment effectiveness. Whether PGT-A improves ICSI outcomes and is beneficial to the patients has remained controversial. STUDY DESIGN, SIZE, DURATION: This is a multinational, multicentre, pragmatic, randomized clinical trial with intention-to-treat analysis. Of 396 women enroled between June 2012 and December 2016, 205 were allocated to CCS of the first and second polar body (study group) as part of their ICSI treatment cycle and 191 were allocated to ICSI treatment without chromosome screening (control group). Block randomization was performed stratified for centre and age group. Participants and clinicians were blinded at the time of enrolment until the day after intervention. PARTICIPANTS/MATERIALS, SETTING, METHODS: Infertile couples in which the female partner was 36-40 years old and who were scheduled to undergo ICSI treatment were eligible. In those assigned to PGT-A, array comparative genomic hybridization (aCGH) analysis of the first and second polar bodies of the fertilized oocytes was performed using the 24sure array of Illumina. If in the first treatment cycle all oocytes were aneuploid, a second treatment with PB array CGH was offered. Participants in the control arm were planned for ICSI without PGT-A. Main exclusion criteria were three or more previous unsuccessful IVF or ICSI cycles, three or more clinical miscarriages, poor response or low ovarian reserve. The primary outcome was the cumulative live birth rate after fresh or frozen embryo transfer recorded over 1 year after the start of the intervention. MAIN RESULTS AND THE ROLE OF CHANCE: Of the 205 participants in the chromosome screening group, 50 (24%) had a live birth with intervention within 1 year, compared to 45 of the 191 in the group without intervention (24%), a difference of 0.83% (95% CI: -7.60 to 9.18%). There were significantly fewer participants in the chromosome screening group with a transfer (relative risk (RR) = 0.81; 95% CI: 0.74-0.89) and fewer with a miscarriage (RR = 0.48; 95% CI: 0.26-0.90). LIMITATIONS, REASONS FOR CAUTION: The targeted sample size was not reached because of suboptimal recruitment; however, the included sample allowed a 90% power to detect the targeted increase. Cumulative outcome data were limited to 1 year. Only 11 patients out of 32 with exclusively aneuploid results underwent a second treatment cycle in the chromosome screening group. WIDER IMPLICATIONS OF THE FINDINGS: The observation that the similarity in birth rates was achieved with fewer transfers, less cryopreservation and fewer miscarriages points to a clinical benefit of PGT-A, and this form of embryo selection may, therefore, be considered to minimize the number of interventions while producing comparable outcomes. Whether these benefits outweigh drawbacks such as the cost for the patient, the higher workload for the IVF lab and the potential effect on the children born after prolonged culture and/or cryopreservation remains to be shown. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the European Society of Human Reproduction and Embryology. Illumina provided microarrays and other consumables necessary for aCGH testing of polar bodies. M.B.'s institution (UZBrussel) has received educational grants from IBSA, Ferring, Organon, Schering-Plough, Merck and Merck Belgium. M.B. has received consultancy and speakers' fees from Organon, Serono Symposia and Merck. G.G. has received personal fees and non-financial support from MSD, Ferring, Merck-Serono, Finox, TEVA, IBSA, Glycotope, Abbott and Gedeon-Richter as well as personal fees from VitroLife, NMC Healthcare, ReprodWissen, BioSilu and ZIVA. W.V., C.S., P.M.B., V.G., G.A., M.D.,T.E.G., L.G., G.Ka., G.Ko., J. L., M.C.M., M.P., A. S., M. T., K.V., J.G. and K.S. declare no conflict of interest. TRIAL REGISTRATION NUMBER: NCT01532284. TRIAL REGISTRATION DATE: 7 February 2012. DATE OF FIRST PATIENT'S ENROLMENT: 25 June 2012.

PGD for HLA typing is a procedure that can be performed when an affected child requires a transplant to treat a non-hereditary disorder related to the hematopoietic and/or immune system. Hematopoietic stem cell transplantation from an HLA-identical donor provides the best treatment option. Three conventional ovarian stimulation procedures for IVF were performed in a couple with a 10-year-old child diagnosed with T-cell acute lymphoblastic leukemia of high risk. Trophectoderm biopsy and aCGH examination were performed on 15 blastocysts, three on the first IVF procedure, four on the second cycle, and eight on the third. Three euploid blastocysts HLA-compatible with the genome of the affected child were identified. One euploid blastocyst HLA-compatible with the affected child was warmed and transferred, resulting in an HLA-matched live birth. In conclusion, combined aCGH for aneuploidy screening and Karyomapping may be performed in a single biopsy procedure.

This study investigates a case series of eight couples who underwent trophectoderm (TE) biopsy and comprehensive chromosomal screening (CCS) for routine aneuploidy screening and were found to have CCS results concerning for previously undetected parental balanced reciprocal translocations.In each case, controlled ovarian hyperstimulation and in vitro fertilization (IVF) yielded multiple blastocysts that each underwent CCS with high-density oligonucleotide microarray comparative genomic hybridization (aCGH).Parental translocations were suspected based on the finding of identical break point mutations in multiple embryos from each couple. Confirmation of these suspected translocations within blastocysts was performed with next-generation sequencing (NGS). Subsequent parental karyotypic evaluation resulted in a diagnosis of parental balanced reciprocal translocation in each case.We demonstrated that high-resolution aCGH and NGS on TE biopsies can accurately detect parental reciprocal translocations when previously unrecognized.