[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.
Journal of Assisted Reproduction and Genetics 11/2015; DOI:10.1007/s10815-015-0605-0 · 1.72 Impact Factor
[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.
Human Reproduction 07/2010; 25(7):1812-7. DOI:10.1093/humrep/deq122 · 4.57 Impact Factor
[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.
Annals of Human Genetics 08/2006; 70(Pt 4):459-87. DOI:10.1111/j.1469-1809.2005.00251.x · 2.21 Impact Factor
[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.
Human Reproduction 10/2005; 20(9):2517-22. DOI:10.1093/humrep/dei087 · 4.57 Impact Factor
[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.
Human Reproduction 02/2004; 19(1):118-23. DOI:10.1093/humrep/deh012 · 4.57 Impact Factor
[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.
European Journal of HumanGenetics 06/2003; 11(5):385-94. DOI:10.1038/sj.ejhg.5200970 · 4.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Haplotype diversity in a genomic region of approximately 70 kb in 1q21 between genes PKLR and GBA was characterized by typing one single nucleotide polymorphism (SNP) in PKLR, two SNPs in GBA and one short tandem repeat polymorphism (STRP) in PKLR in 1792 chromosomes from 17 worldwide populations. Two other SNPs in GBA were typed in three African populations. Most chromosomes carried one of either two phylogenetically distinct haplotypes with different alleles at each site. Allele diversity at the STRP was tightly linked to haplotype background. Linkage disequilibrium (LD) was highly significant for all SNP pairs in all populations, although it was, on average, slightly higher in non-African populations than in sub-Saharan Africans. Variation at PKLR-GBA was also tightly linked to that at the GBA pseudogene, 16 kb downstream from GBA. Thus, a 90 kb-long LD block was observed, which points to a low recombination rate in this region. Detailed haplotype phylogeny suggests that the chimpanzee GBA haplotype is not one of the two most frequent haplotypes. Based on variability at the PKLR STRP and on the geographical distribution of LD, the expansion of the two main haplotypes may have predated the "Out of Africa" expansion of anatomically modern humans. LD and STRP variability in non-Africans are approximately 87% of those in Africans, in contrast with other loci; this implies that the "out of Africa" bottleneck may have had a broad distribution of effects across loci.
Human Genetics 07/2002; 110(6):532-44. DOI:10.1007/s00439-002-0734-2 · 4.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The genetic background of the mutations that most often cause cystic fibrosis (CF) is different from that of non-CF chromosomes in populations of European origin. It is not known whether these haplotype backgrounds could be found at high frequencies in populations in which CF is, at present, not common; such populations would be candidates for the place of origin of CF mutations. An analysis of haplotypes of CF transmembrane conductance regulator, together with their variation in specific CF chromosomes, in a worldwide survey of normal chromosomes shows (1) a very low frequency or absence of the most common CF haplotypes in all populations analyzed and (2) a strong genetic variability and divergence, among various populations, of the chromosomes that carry disease-causing mutations. The depth of the gene genealogy associated with disease-causing mutations may be greater than that of the evolutionary process that gave rise to present-day human populations. The concept of "population of origin" lacks either spatial or temporal meaning for mutations that are likely to have been present in Europeans before the ethnogenesis of present populations; subsequent population processes may have erased the traces of their geographic origin.
The American Journal of Human Genetics 02/2002; 70(1):257-64. DOI:10.1086/338243 · 10.93 Impact Factor