[Show abstract][Hide abstract] ABSTRACT: Purpose:
The purpose of this study was to compare the confirmation rate of day-3 embryo biopsy (blastomere) and trophectoderm biopsy using array-comparative genomic hybridization (array-CGH) technology.
A blinded study was conducted to re-analyse 109 embryos previously diagnosed as chromosomally abnormal by array-CGH. Preimplantation genetic screening (PGS) was performed using array-CGH on day 3 (n = 50) or day 5 (n = 59). Partial chromosome gains or losses were excluded (n=6), and only whole chromosome aneuploidies were considered. Re-analysis of whole blastocysts was carried out following the same array-CGH protocol used for PGS.
The PGS result was confirmed in the whole blastocyst in (a) 49/50 (98 %) abnormal embryos after day-3 biopsy and (b) 57/59 (96.6 %) abnormal embryos after trophectoderm biopsy. One embryo (1/50; 2 %) was diagnosed as abnormal, with monosomy 18, on day 3, and software analysis of the whole blastocyst gave a euploid result; however, a mosaic pattern was observed for monosomy 18 in the whole blastocyst. Two trophectoderm biopsy cases (3.4 %) did not have the abnormalities (trisomy 7, and trisomy 1 and 4, respectively) verified in the whole embryo. Concordance rates for both biopsy strategies and for individual chromosomes were evaluated by Fisher's exact test and showed no significant differences.
Both types of biopsies showed similar high concordance rates with whole blastocyst results. Therefore, regarding the confirmation rates shown in this work, day-3 embryo biopsies can be representative of the whole embryo and both types of biopsy can be used for clinical analysis in PGS following the described array-CGH protocol.
Full-text · Article · Nov 2015 · Journal of Assisted Reproduction and Genetics
[Show abstract][Hide abstract] ABSTRACT: Study question
Is blastulation rate associated with euploid embryo status?
Euploid embryos appear to be three times more likely to undergo blastulation than aneuploid embryos among six to eight cell embryos undergoing blastomere biopsy and array CGH for Pregestational Genetic Screening (PGS).
What is known already
Recent studies on trophoectoderm biopsies show a higher rate of euploid embryos at the blastocyst stage than the euploid rate previously reported from cleavage stage blastomere biopsies. However, it has historically been thought that aneuploid embryos are just as likely to undergo blastulation as euploid embryos.
Study design, size, duration
Retrospective cohort study was performed on 44 cycles between January 2011 and December 2012 that underwent IVF and day three single cell blastomere biopsy with array CGH and PGS.
Participants/materials, setting, methods
Subjects underwent IVF-ICSI and CGH at a university hospital based IVF center. All cleavage-stage embryos underwent single cell blastomere biopsy and fixation for microarray CGH and results were reported for all embryos on day 5. The percent of both euploid and aneuploid embryos were compared utilizing Fisher's exact test.
Main results and the role of chance
Mean patient age among the 44 cycles was 37.2 (range 28-42). A total of 463 embryos were produced in 44 cycles from which 382 (82.5%) six to eight embryos were biopsied. Overall blastulation rate for all biopsied embryos was 32.0% which was not significantly different from a matched control group that did not undergo biopsy. The number of euploid and aneuploid embryos after biopsy was 106 (27.7%) and 276 (72.3%), respectively. 84 (79.2%) of the euploid embryos and 66 (23.9%) of the aneuploid embryos progressed to blastocyst stage. (p < 0.0001). However only 58/106 (54.7%) of the euploid embryos formed fully expanded or hatching blastocysts by day 5, and therefore would not have been able to undergo trophoectoderm biopsy.
Limitations, reason for caution
Limitations include retrospective design and somewhat small study group. Additionally mosaicism in single cell biopsy could affect our outcomes.
Wider implications of the findings
Our results suggest that euploid embryos are far more likely to undergo blastulation than aneuploid embryos. However there are still many aneuploid blastocysts that would be transferred in cycles without PGS. One advantage of day three biopsy is that only half of the euploid embryos in this study advanced enough for trophoectoderm biopsy on day five.
Study funding/competing interest(s)
The study had no funding.
Trial registration number
Full-text · Article · Jun 2013 · Human Reproduction
[Show abstract][Hide abstract] ABSTRACT: In our routine programme of preimplantation genetic aneuploidy screening (PGS) by fluorescence in situ hybridization (FISH), nine chromosomes (13, 15, 16, 17, 18, 21, 22, X and Y) are analysed in two consecutive hybridization rounds. We also perform additional hybridization rounds for these chromosomes, using probes that bind to different loci, for non-conclusive results and for confirmation of certain aneuploidies. The aim of this study was to evaluate the impact of additional hybridization rounds on FISH accuracy.
This is a retrospective analysis of our FISH data from 1000 PGS cycles performed from December 2007 to December 2008 for various indications. In addition to the hybridization rounds described above, 132 of the embryos diagnosed as chromosomally abnormal were re-analysed on Day 5.
A total of 2477 embryos were re-hybridized, 1496 due to non-conclusive results and 981 to confirm observed aneuploidies. After re-hybridization, 882 embryos (59%) were then diagnosed as normal, 600 embryos (40.1%) had a clear abnormality and only 14 embryos (0.9%) remained non-informative. From the 981 embryos in the latter group, 890 embryos had monosomies and, after re-hybridization 174 embryos (19.6%) were normal and 716 (80.5%) had confirmed monosomies. In contrast, re-hybridization confirmed 90 (98.9%) of the 91 observed trisomies. In addition, Day-5 re-analysis of abnormal embryos showed a higher rate of concordant diagnosis between Day 3 and Day 5 when re-hybridizations had been included on Day-3 (95 versus 82.7%; P= 0.0443), especially for the confirmation of monosomies (82.8 versus 61.0%; P = 0.0087).
Our data indicate that additional hybridization rounds improve the accuracy of the diagnosis, increasing the number of chromosomally normal embryos available for transfer. Re-hybridization with additional probes as a standard approach to PGS could enhance the potential benefits of the technique.
[Show abstract][Hide abstract] ABSTRACT: The Balkan Peninsula is a complex cultural mosaic comprising populations speaking languages from several branches of the Indo-European family and Altaic, as well as culturally-defined minorities such as the Aromuns who speak a Romance language. The current cultural and linguistic landscape is a palimpsest in which different peoples have contributed their cultures in a historical succession. We have sought to find any evidence of genetic stratification related to those cultural layers by typing both mtDNA and Y chromosomes, in Albanians, Romanians, Macedonians, Greeks, and five Aromun populations. We have paid special attention to the Aromuns, and sought to test genetically various hypotheses on their origins. MtDNA and Y-chromosome haplogroup frequencies in the Balkans were found to be similar to those elsewhere in Europe. MtDNA sequences and Y-chromosome STR haplotypes revealed decreased variation in some Aromun populations. Variation within Aromun populations was the primary source of genetic differentiation. Y-chromosome haplotypes tended to be shared across Aromuns, but not across non-Aromun populations. These results point to a possible common origin of the Aromuns, with drift acting to differentiate the separate Aromun communities. The homogeneity of Balkan populations prevented testing for the origin of the Aromuns, although a significant Roman contribution can be ruled out.
Full-text · Article · Aug 2006 · Annals of Human Genetics
[Show abstract][Hide abstract] ABSTRACT: Previous observations have addressed a decreased male:female ratio associated with smoking. Our aim was to assess whether this effect is observed at the spermatozoa or at the early embryo development.
We retrospectively assessed smoking intake habits of 56 couples included in our preimplantation genetic diagnosis (PGD) program. Three groups were established according to male or female cigarette consumption per day: non-smokers, smokers (1-19 cigarettes per day) and heavy smokers (> or =20 cigarettes per day). Fluorescence in-situ hybridization (FISH) was performed on ejaculated sperm samples to analyse chromosomes X and Y. On day 3, embryos were also analysed. Additionally, sperm samples from four heavy smoking and four non-smoking donors were prospectively analysed before and after capacitation.
FISH on spermatozoa revealed no statistical differences in the Y:X ratio between the three groups. However, in the PGD study, in male heavy smokers, the XY:XX embryo ratio was decreased compared with non-smokers (22:47 versus 80:71; P = 0.0057). The smoking condition of the female partner had no significant effect on embryo XY:XX ratio, but for non-smoking females with a heavy smoking partner, the ratio was decreased (P = 0.0018) compared with non-smoking males. In heavy smoking donors a decreased of Y:X ratio was observed after swim-up with a statistically significant difference of ratios (P = 0.021).
Smoking habits of males do not have an effect on the percentage of X- and Y-bearing spermatozoa on ejaculated samples. However, male heavy smokers produce an increased incidence of female embryos that could be related to an enrichment of X spermatozoa after swim-up in patients with high tobacco consumption.
Full-text · Article · Oct 2005 · Human Reproduction
[Show abstract][Hide abstract] ABSTRACT: An increased incidence of numerical chromosomal abnormalities has been reported in the ejaculated spermatozoa of infertile patients. However, there are few cytogenetic studies of testicular and epididymal spermatozoa, and their results are still controversial.
Fluorescence in-situ hybridization (FISH) analysis of chromosomes 13, 18, 21, X and Y was performed on seven testicular samples and two epididymal samples from patients with obstructive azoospermia (OA), and on 13 testicular samples from patients with non-obstructive azoospermia (NOA). Five ejaculated sperm samples from normozoospermic fertile donors were evaluated as a control group.
Both epididymal sperm samples showed normal FISH results for the parameters analysed when compared with those of the control group. FISH results were abnormal in 29% (two of seven) of testicular samples from OA patients and in 54% (seven of 13) of those from NOA patients, although this difference was not statistically significant. Testicular samples from OA patients showed a significant increase of disomy for sex chromosomes (P<0.01), whereas NOA patients displayed significantly higher rates of diploidy (P<0.0001) and disomy for chromosomes 13 (P<0.0001), 21 (P<0.001) and sex chromosomes (P<0.0001) than the control group.
Testicular spermatozoa from azoospermic patients present increased rates of chromosomal abnormalities, mainly of the sex chromosomes, which are particularly high in NOA patients.
Full-text · Article · Feb 2004 · Human Reproduction
[Show abstract][Hide abstract] ABSTRACT: Cystic fibrosis (CF) is the most frequent severe recessive disorder in European populations. We have analyzed its mutation frequency spectrum in 94 European, North African and SW Asian populations taken from the literature. Most major mutations as well as the incidence of CF mutations showed clinals patterns as demonstrated by autocorrelogram analysis. More importantly, measures of mutation diversity did also show clinal patterns, with mutation spectra being more diverse in southern than in northern Europe. This increased diversity would imply roughly a three-fold long-term effective population size in southern than in northern Europe. Distances were computed among populations based on their CF mutation frequencies and compared with distances based on other genic regions. CF-based distances correlated with mtDNA but not with Y-chromosome-based distances, which may be a consequence of the relatively homogeneous CF mutation frequencies in European populations.
Full-text · Article · Jun 2003 · European Journal of HumanGenetics