A short fluorescence in-situ hybridization (FISH) procedure using fluorochrome and digoxigenin labelled DNA probes was developed for application in human preimplantation embryos in order to analyse the five chromosomes most involved in human aneuploidy (X, Y, 18, 13 and 21). The chromosomes were fluorescent-stained and detected simultaneously in 157 blastomeres from 30 human embryos. Successful FISH analysis was achieved in 93% of the blastomeres. Aberrations for these chromosomes were found in 70% of abnormally developing monospermic embryos. The majority of normally developing monospermic embryos obtained from older patients were also chromosomally abnormal. By analysing all or most of the cells from these embryos, true mosaicism was distinguished from technique failure. Mosaic embryos, polyploid embryos with ploidies as high as 8n, haploid embryos, embryos monosomic for 13/21 and for X, and embryos trisomic for 13/21 and 18, were common in abnormally developing embryos. In contrast, aneuploidy was the main chromosome abnormality found in normally developing monospermic embryos.

To determine the pregnancy outcome potential of mosaic embryos, detected by means of preimplantation genetic screening (PGS) with the use of next-generation sequencing (NGS). Retrospective study. Genetics laboratories. PGS cycles during which either mosaic or euploid embryos were replaced. Blastocysts were biopsied and processed with the use of NGS, followed by frozen embryo transfer. Trophectoderm (TE) biopsies were classified as mosaic if they had 20%-80% abnormal cells. Implantation, miscarriage rates, and ongoing implantation rates (OIRs) were compared between euploid and types of mosaic blastocysts. Complex mosaic embryos had a significantly lower OIR (10%) than aneuploidy mosaic (50%), double aneuploidy mosaic (45%), and segmental mosaic (41%). There was a tendency for mosaics with 40%-80% abnormal cells to have a lower OIR than those with 40% abnormal cells and those with multiple mosaic abnormalities (chaotic mosaics) are likely to have lower OIRs and should be given low transfer priority.

Background The 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 transfer of euploid embryos. Recently, randomised trials using such methods have indicated that aneuploidy screening improves IVF success rates. However, the high cost of testing has restricted the availability of this potentially beneficial strategy. This study aimed to harness next-generation sequencing (NGS) technology, with the intention of lowering the costs of preimplantation aneuploidy screening. Methods Embryo biopsy, whole genome amplification and semiconductor sequencing. Results A rapid (<15 h) NGS protocol was developed, with consumable cost only two-thirds that of the most widely used method for embryo aneuploidy detection. Validation involved blinded analysis of 54 cells from cell lines or biopsies from human embryos. Sensitivity and specificity were 100%. The method was applied clinically, assisting in the selection of euploid embryos in two IVF cycles, producing healthy children in both cases. The NGS approach was also able to reveal specified mutations in the nuclear or mitochondrial genomes in parallel with chromosome assessment. Interestingly, elevated mitochondrial DNA content was associated with aneuploidy (p<0.05), a finding suggestive of a link between mitochondria and chromosomal malsegregation. Conclusions This study demonstrates that NGS provides highly accurate, low-cost diagnosis of aneuploidy in cells from human preimplantation embryos and is rapid enough to allow testing without embryo cryopreservation. The method described also has the potential to shed light on other aspects of embryo genetics of relevance to health and viability.