Mosaic Chromosome Abnormalities Research Articles

Diagnostic and prognostic role of soft ultrasound markers in prenatal detection and assessment of foetal abnormalities.

Various soft markers can be detected in the ultrasonography of foetuses, which can be related to chromosomal abnormalities and increases the risk of abnormalities, or they can be considered as normal variations that can disappear due to the pregnancy progress. There are different tools to detect chromosomal abnormalities like conventional karyotyping, chromosomal microarray analysis (CMA), single nucleotide polymorphism (SNP) array, non-invasive prenatal test (NIPT), and non-invasive prenatal screening (NIPS). Therefore, in the present study, we aim to assess the accuracy of ultrasonic soft markers in the diagnosis of chromosomal abnormalities such as chromosomal structural abnormalities, aneuploidy, and triploidy, especially Trisomy 21 and Trisomy 18. A systemic literature search was performed using PubMed, Scopus, Google Scholar, and Web of Science. We gathered all articles published before August 2023. We selected English studies such as retrospective and cross-sectional ones that assessed the relationship between ultrasonic soft markers and foetal chromosomal abnormalities. A total of 10 articles with 18,580 cases were included in our systematic review article that assessed the foetal abnormalities and aneuploidies by using conventional karyotyping, SNP array, CMA, and NIPT (or NIPS). Trisomy 21, Trisomy 18, and chromosomal structural abnormalities were the most common abnormalities related to ultrasonic soft markers by karyotyping; however, Trisomy 13, 47, XXY, 45, X, and mosaic chromosomal abnormalities were other abnormalities detected. Results by CMA showed Trisomy 21 and Trisomy 18 as the most common abnormalities in the foetuses also with ultrasonic soft markers, and other abnormalities were pathogenic copy-number variations, Turner (XO), polyploidy, 22q11.2deletion, and Trisomy13, respectively. It was discovered that there is a greater possibility of having pathogenic copy number variations (CNVs) in the groups with multiple ultrasonic soft markers, while foetuses with ultrasonic soft markers have a decreased prevalence of CMA abnormality compared to those who had significant abnormalities or abnormal nuchal translucency. Trisomy 21 was the only abnormality found by NIPT in the groups with 1 and 2 soft markers, while groups with multiple soft markers were all normal. By using SNP array, it was identified that the rate of chromosomal abnormalities such as aneuploidy and triploidy, LOH, and CNVs was lower in the group with a single ultrasonic soft marker compared to the group with structural abnormalities in multiple systems. Trisomy 21, Trisomy 18, and chromosomal structural abnormalities were the most common chromosomal abnormalities that ultrasound soft markers could diagnose. Therefore, it is recommended to employ soft markers besides CMA, SNP array, and NIPS (or NIPT) for greater accuracy in detecting foetus abnormalities.

Prevalence of chromosomal alterations in first-trimester spontaneous pregnancy loss

Pregnancy loss is often caused by chromosomal abnormalities of the conceptus. The prevalence of these abnormalities and the allocation of (ab)normal cells in embryonic and placental lineages during intrauterine development remain elusive. In this study, we analyzed 1,745 spontaneous pregnancy losses and found that roughly half (50.4%) of the products of conception (POCs) were karyotypically abnormal, with maternal and paternal age independently contributing to the increased genomic aberration rate. We applied genome haplarithmisis to a subset of 94 pregnancy losses with normal parental and POC karyotypes. Genotyping of parental DNA as well as POC extra-embryonic mesoderm and chorionic villi DNA, representing embryonic and trophoblastic tissues, enabled characterization of the genomic landscape of both lineages. Of these pregnancy losses, 35.1% had chromosomal aberrations not previously detected by karyotyping, increasing the rate of aberrations of pregnancy losses to 67.8% by extrapolation. In contrast to viable pregnancies where mosaic chromosomal abnormalities are often restricted to chorionic villi, such as confined placental mosaicism, we found a higher degree of mosaic chromosomal imbalances in extra-embryonic mesoderm rather than chorionic villi. Our results stress the importance of scrutinizing the full allelic architecture of genomic abnormalities in pregnancy loss to improve clinical management and basic research of this devastating condition.

O-078 Mosaic embryos – are they accurately detected by PGT-A?

Abstract Study question Following quantitative NGS for PGT-A, do current thresholds for mosaicism calling give a true reflection of the presumed mitotic origin of mosaic errors? Summary answer A large proportion of embryo mosaicism diagnosed following quantitative NGS-based PGT-A does not correlate with a true and expected mitotic origin, showing instead meiotic signatures. What is known already Mosaicism is a true biological phenomenon that can affect ongoing embryo development. However, the reproductive potential of mosaic embryos in assisted reproduction varies greatly across publications and is not fully understood. While the transfer of mosaic embryos could result in healthy live births, embryos containing meiotic errors almost never result in healthy live births. Since the clinical implications of meiotic and mitotic errors are very different, it is critical to accurately differentiate between the two. However, diagnosing mosaicism using current NGS-based PGT-A is not always accurate due to technical and biological limitations which may misclassify mosaic embryos. Study design, size, duration Follow-up analysis was carried out on 159 embryos previously categorised as mosaic by NGS-based PGT-A. To identify the origin of mosaic calls, SNP genotyping and karyomapping were performed on the same amplified DNA used for the clinical NGS analysis. Trophectoderm biopsy samples were obtained from embryos on days 5,6 or 7 of development from a single IVF centre between 2018-2022. For all PGT-A tested embryos, the reported mosaicism rate at the IVF centre was 12%. Participants/materials, setting, methods Samples were analysed using PGTai from CooperSurgical with a 30-80% CNV mosaicism range. Patients with mosaic embryos consented to karyomapping follow-up using sibling embryo reference DNA. The origin of mosaic events was defined by SNP inheritance patterns for the affected chromosome. Meiotic errors were defined by the presence of both haplotypes from one parent for trisomy, and the absence of a parental haplotype for monosomy. Mitotic errors were inferred following a result of biparental inheritance. Main results and the role of chance A total of 154 diploid embryos identified as mosaic by NGS-based PGT-A were re-analysed by SNP-genotyping. Of these, 30.5% (n = 47) displayed at least one error of meiotic origin following SNP genotyping. Among the 154 embryos, 189 mosaic chromosomal abnormalities were identified. After SNP-genotyping, 30.7% of chromosomal abnormalities (n = 58) showed evidence of meiotic signatures: 52 whole-chromosome (24 gains, 28 losses), and 6 segmental errors (5 gains and 1 loss). Of 73 segmental errors detected in total, only 8.2% of them (n = 6) were identified to be of meiotic origin. Following PGT-A, 7.8% of embryos (n = 12/154) displayed ≥3 chromosomal abnormalities in the mosaic range. Of these embryos, 58.3% (n = 7/12) displayed one or more meiotic aneuploidies. Overall, embryos showing multiple mosaic events were significantly more likely to have a least one error of meiotic origin than embryos containing one abnormality (McNemar’s test p-value= 1.171e-14). Among mosaic calls above 50% CNV, 56.7% (n = 55/97) showed evidence of meiotic origin. In contrast, but importantly, in the low-moderate mosaicism range <50% CNV, 3.3% of errors (3/92) also displayed meiotic signatures, including two whole chromosome errors (chr21 gain at 32% CNV and chr13 loss at 46% CNV), and one segmental error (dup(6)(pter-p12.1) at 33% CNV). Limitations, reasons for caution Since karyomapping relies on recombination events to identify meiotic trisomy, the true origin of intermediate copy-number changes cannot always be identified. Therefore, errors without recombination may be missed, potentially underestimating meiotic gains, especially segmentals. The absence of a parental haplotype cannot entirely rule-out a mitotic error from a biopsy sample. Wider implications of the findings Quantitative NGS methodologies are unable to accurately distinguish the origin of mosaic errors in PGT-A samples. Since the clinical impact of meiotic and mitotic errors differs greatly, this limitation could explain mosaic embryos’ reduced viability. Reliable identification of meiotic aneuploidies in presumed mosaic embryos is likely to improve clinical outcomes. Trial registration number not applicable

P-066 The impact of male factor infertility on the chromosomal status of the pre-implantation embryo

Abstract Study question Do poor sperm parameters and/or advance male age increase the rate of aneuploid blastocysts in PGT-A cycles? Summary answer In our study neither impaired semen quality nor advanced male age increased the rate of aneuploid blastocysts after PGT-A. What is known already The vast majority of aneuploidies in pre-implantation embryos arise from failures during the meiotic division of the oocyte. The extend of the paternal contribution to aneuploidies in the embryo is still discussed. It is known that that the rates of aneuploid spermatozoa increase from around 4.5% in normozoospermia by 2-6-fold in severe male factor infertility. Despite this fact, several studies reported no effect of sperm quality or male age on aneuploidy rates in IVF-embryos, however reports are controversial. Actually, severe male factor is still rated as an indication for PGT-A in several countries. Study design, size, duration In this retrospective multi-center study (2016-2018) we analyzed 298 PGT-A cycles from 179 patients. Calculations was performed for two groups according to female age (group 1 women ≤ 35 years; group 2 women ≥ 36 years). Sperm parameter were classified according to the WHO 2010 criteria and the motile sperm organelle morphology examination (MSOME) classification. Participants/materials, setting, methods Blastocyst culture was performed in a single step culture medium in a time-lapse incubator. Biopsy of 5-10 TE-cells was performed on culture day 5-6 on 1012 blastocysts. For whole genome amplification (WGA) PicoPlex (Rubicon Genomics) or SurPlex (BlueGenome/ Illumina) platforms were used. Next generation sequencing (NGS) was processed using the VeriSeq platform and data analysis by BlueFuse Multi (Illumina). Main results and the role of chance Comparing men with normozoospermia to men showing reduced sperm sperm parameters, we found in the group of women ≤ 35 years (group 1) a rate of 40.3% vs. 45.2% euploid blastocysts after TE biopsy. In the group of women ≥ 36 years (group 2) the rate of euploid blastocysts was 19.3% vs. 23.1% in normozoospermia vs. non-normozoospermia men. No difference in the number of blastocysts with uniform or mosaic chromosomal abnormalities were found between both groups. In a further analysis, no impact of sperm parameters on the number embryos with of sex chromosome aberrations was observed. With application of the MSOME-classification we found a similar picture. There was no difference in the rate of aneuploid blastocysts with o%, 1-3% or min. 4% normal forms in both groups. When focusing on male age by comparing men ≤ 49 years and ≥ 50 years, we found no difference in the rate of euploid blastocysts. In group the rate of euploid blastocysts was 42.1% vs. 47.5% and in group 2 the rates for euploid blastocysts were 21.8% vs. 19.8% in men ≤ 49 years and ≥ 50 years, respectively. Limitations, reasons for caution Chromosomal screening of all chromosomes using PGT-A will not detect small chromosomal aberrations such as small deletions, insertions or mutations. These small aberration could play an important role in male factor infertility. Focus if the present study was to test the efficiency of PGT-A in reduced sperm parameters. Wider implications of the findings In line with previous studies we found neither an impact of poor sperm quality, nor of advanced male age on the rate of euploid blastocysts after PGT-A. However, results published are controversial and male factor as indication for PGT-A has to be further evaluated. Trial registration number EK-2-8/2020

Clonal myelopoiesis promotes adverse outcomes in chronic kidney disease

We sought to determine the relationship between age-related clonal hematopoiesis (CH) and chronic kidney disease (CKD). CH, defined as mosaic chromosome abnormalities (mCA) and/or driver mutations was identified in 5449 (2.9%) eligible UK Biobank participants (n = 190,487 median age = 58 years). CH was negatively associated with glomerular filtration rate estimated from cystatin-C (eGFR.cys; β = −0.75, P = 2.37 × 10–4), but not with eGFR estimated from creatinine, and was specifically associated with CKD defined by eGFR.cys < 60 (OR = 1.02, P = 8.44 × 10–8). In participants without prevalent myeloid neoplasms, eGFR.cys was associated with myeloid mCA (n = 148, β = −3.36, P = 0.01) and somatic driver mutations (n = 3241, β = −1.08, P = 6.25 × 10–5) associated with myeloid neoplasia (myeloid CH), specifically mutations in CBL, TET2, JAK2, PPM1D and GNB1 but not DNMT3A or ASXL1. In participants with no history of cardiovascular disease or myeloid neoplasms, myeloid CH increased the risk of adverse outcomes in CKD (HR = 1.6, P = 0.002) compared to those without myeloid CH. Mendelian randomisation analysis provided suggestive evidence for a causal relationship between CH and CKD (P = 0.03). We conclude that CH, and specifically myeloid CH, is associated with CKD defined by eGFR.cys. Myeloid CH promotes adverse outcomes in CKD, highlighting the importance of the interaction between intrinsic and extrinsic factors to define the health risk associated with CH.

O-020 Aneuploidy and mosaicism in human embryos: How correct detection may improve IVF clinical outcomes

Abstract text Study question Can new next-generation sequencing (NGS) based strategies for preimplantation genetic testing of aneuploidy (PGT-A) improve clinical outcomes after assisted reproductive technology (ART)? Summary answer Recent randomised controlled trials (RCTs) suggest that NGS-based PGT-A strategies can improve clinical outcomes for older women. The clinical management of mosaic embryos remains controversial. What is known already There are two types of chromosome abnormalities present in embryos, meiotic arising mostly during oogenesis, and mitotic arising after fertilisation. Meiotic aneuploidies are present in all of the embryonic cells and in their vast majority are lethal. Conversely, mitotic abnormalities are present in only part of the embryonic cells with the remaining cells having a different cytogenetic constitution. This phenomenon is known as mosaicism. The sensitivity of NGS meant that mosaic aneuploidy became readily detectable in trophectoderm (TE) samples during PGT-A. The viability and clinical management of mosaic embryos has led to debates and controversies in the reproductive medicine field. Study design, size, duration The study involved an assessment of the impact of mosaic chromosome abnormalities to embryonic viability and clinical outcomes after ART cycles using PGT-A via NGS. A large number of embryos generated in IVF clinics in Europe and the USA was examined. Participants/materials, setting, methods Embryos were generated by couples referred for PGT-A due to various indications. All embryos were cultured to the blastocyst stage, and underwent a TE biopsy, followed by vitrification. TE samples were shipped to 6 reference PGT laboratories and analysed via the use of the same NGS platform. Mosaic chromosome abnormalities were scored according to validated thresholds set by the reference laboratories. The clinical management of mosaic embryos took place according to published guidelines. Main results and the role of chance Comparison of clinical outcomes seen after the transfer of mosaic embryos with those seen after the transfer of euploid embryos led to the following findings: Mosaic embryos with &amp;lt;40% abnormal cells in the TE sample had an OIR of 50% compared to 27% for mosaics with 40–80% abnormal cells in the TE, and 9% for complex mosaic embryos. Karyotyping of ongoing pregnancies resulting after the transfer of mosaic embryos demonstrated a normal chromosome constitution of the resulting foetuses. Limitations, reasons for caution - Cytogenetic classification was based on TE samples removed from blastocysts during PGT-A analysis. As only a fraction of the cells from each embryo are tested, inevitably some mosaic embryos will be incorrectly classified fully euploid or aneuploid. However, this misclassification is expected to have little impact on the results. Wider implications of the findings - The transfer of NGS-classified mosaic embryos was associated with poorer clinical outcomes compared to euploid embryos. However, the ongoing pregnancies resulting from mosaic transfers were euploid. NGS’s ability to identify embryos of reduced viability has the potential to improve IVF clinical outcomes.

Clonal myelopoiesis in the UK Biobank cohort: ASXL1 mutations are strongly associated with smoking.

We sought to determine the significance of myeloid clonal hematopoiesis (CH) in the UK Biobank cohort (n = 502,524, median age = 58 years). Utilizing SNP array (n = 486,941) and whole exome sequencing data (n = 49,956), we identified 1166 participants with myeloid CH, defined by myeloid-associated mosaic chromosome abnormalities (mCA) and/or likely somatic driver mutations in DNMT3A, TET2, ASXL1, JAK2, SRSF2, or PPM1D. Myeloid CH increased by 1.1-fold per annum (myeloid mCA, P = 1.57 × 10-38; driver mutations, P = 5.89 × 10-47). Genome-wide association analysis identified two distinct signals within TERT that predisposed to myeloid CH, plus a weaker signal corresponding to the JAK2 46/1 haplotype. Specific subtypes of myeloid CH were associated with several blood features and clinical phenotypes, including TET2 mutations and chronic obstructive pulmonary disease. Smoking history was significantly associated with myeloid CH: 53% of myeloid CH cases were smokers compared to 44% of controls (P = 3.38 × 10-6), a difference principally due to current (OR = 1.10; P = 6.14 × 10-6) rather than past smoking (P = 0.08). Breakdown of CH by specific mutation type revealed that ASXL1 loss of function mutations were most strongly associated with current smoking status (OR = 1.07; P = 1.92 × 10-5), and the only abnormality associated with past smoking (OR = 1.04; P = 0.0026). We suggest that the inflammatory environment induced by smoking may promote the outgrowth of ASXL1-mutant clones.

Thanatophoric Dysplasia

Objective: Report a case of thanatophoric dysplasiaMethod: Case report Result: Case of a 25-year-old woman, with a diagnosis of gravid preterm G4P2A1H2 31-32 weeks + polyhydramnios + fetal hydrops, a single intrauterine live fetus with thanatophoric dysplasia. On ultrasound examination found fetal biometry; BPD: 7.78 cm, FL: 3.58 cm, HL: 3.11 cm, AC: 30.90 cm, HC: 28.48 cm AFI: 33.27 cm, a frontal bossing (+) picture appears, claver leaf skull (+) and micromelia (proximal, distal, phalanges). The ultrasound examination suggested Severe skeletal dysplasia (thanatophoric dysplasia), polyhydramnios, + single intrauterine live fetus + SC 1x scars. Then an amnioinfusion is performed and results are obtained. Chromosome analysis is carried out using the G-banding technique. Chromosomes have been studied from 20 cells from 3 different cell culture preparations and obtained the number of chromosomes in each cell studied is 46, XY which means the number of chromosomes 46 pieces with fetal sex chromosome XY. Mosaic chromosome abnormalities generally occur due to non-disjuntion in the mitotic phase after conception. At 33-34 weeks gestation, an infant was born by SC with birth weight: 1900 g, baby’s length: 31 cm, A / S 2/3.Conclusion : Thanatophoric dysplasia is a "lethal" skeletal dysplasia. A careful prenatal examination is needed in the diagnosis and termination of pregnancy.Keywords: Thanatophoric dysplasia, prenatal diagnosis

Mosaicism: detection and clinical outcomes

Introduction It has been proposed that selecting euploid blastocysts to transfer via preimplantation genetic testing for aneuploidy (PGT-A) with the use of comprehensive chromosome screening (CCS) methods, could lead to significant improvements in IVF outcomes. However, questions have been raised related to the implantation potential of mosaic embryos, composed of a mixture of normal and chromosomally abnormal cells. The main aim of this study was to examine the developmental ability of mosaic blastocysts. Materials and methods 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. Results Different aneuploidy types, including some complex forms, were capable of persisting to the blastocyst stage. Euploidy rates decreased dramatically with advancing female age, whereas uniform aneuploidy increased. Moreover, the transfer of mosaic diploid aneuploid blastocysts was associated with higher miscarriage and lower implantation rates, compared to embryos in which only euploid cells were detected in the TE biopsy. Conclusions Extensive analysis of the chromosome constitution of human blastocysts suggested that a significant proportion are chromosomally normal in every cell. These findings do not support the notion that mosaic chromosome abnormalities are a natural part of embryo development. Blastocysts with uniform aneuploidy present in the biopsied TE sample and, to a lesser extent, those with mosaic abnormalities were associated with poorer clinical outcomes, compared to euploid embryos.

Cytogenetic Analysis of Spontaneous Miscarriages Using Long-Term Culturing of Chorionic Villi

Spontaneous miscarriages are strongly associated with fetal chromosomal abnormalities. We sought to validate the occurrence of chromosomal abnormalities in Japanese patients with early spontaneous miscarriages using a conventional long-term culture method. One hundred and thirty-two patients were studied after excluding 7 cases with culture failure. Chromosomal analysis of 132 cases was performed using a long-term culture method of the villous placental tissue. Patients’ age, number of previous spontaneous abortions, fetal data from ultrasonography, and the method of conception (ART or spontaneous pregnancy) were obtained from the electronic medical records. The association between the patient’s background and the cytogenetic data was analyzed using a Chi squared test. The average maternal age of the 132 cases of early spontaneous abortion was 34.2 ± 4.7 year (range: 24–43). Chromosomal abnormalities were detected in 89 cases (67.4%), among which 52.8% represented trisomy, 1.1% represented double trisomy, 10.1% represented polyploidy, 7.9% represented monosomy, 12.4% represented structural chromosomal abnormality, and 15.7% represented mosaic chromosome abnormalities. Women older than 35 year had a higher rate of chromosomal abnormalities than younger women (76.6% vs. 58.8%; P = 0.015). Twenty-five of the 132 pregnancies were achieved with ART and were associated with a higher rate of trisomy (64.0% vs. 37.4%; P = 0.008). G-banding identified a rare form of polyploidy 94,XXYYYY. Our results confirmed previous findings that older Japanese women with spontaneous miscarriages have a higher rate of chromosomal abnormalities.

The cytogenetic constitution of human blastocysts: insights from comprehensive chromosome screening strategies.

Embryos that are able to form blastocysts have succeeded in activating their genome and differentiating into two cell types-an external layer of trophectoderm cells, which will go on to form extra-embryonic tissues such as the placenta, and the inner cell mass, which will give rise to the embryo proper. Culturing embryos to the blastocyst stage has become an increasingly popular IVF practice over the past decade. Additionally, it has been proposed that the identification and transfer of euploid blastocysts could significantly improve IVF outcomes. Toward this end, comprehensive molecular cytogenetic methods have been developed. The application of such methods in both clinical and research contexts has yielded cytogenetic data from large numbers of blastocysts. Questions have been raised concerning the implantation potential of blastocysts diagnosed 'euploid' or 'aneuploid', and there has been particular debate over the detection and viability of embryos categorized as 'mosaic'-composed of a mixture of normal and aneuploid cells. This review aims to summarize data from studies using comprehensive molecular cytogenetic methods to examine blastocyst-stage embryos, describing current knowledge related to rates of euploidy, uniform aneuploidy and mosaicism. Issues associated with the developmental capacity of blastocysts of different cytogenetic constitutions will also be addressed. Guidelines on the clinical management of blastocysts with varying chromosome complements will be considered. Rates of euploidy, uniform aneuploidy (in which all cells have the same abnormal karyotype) and mosaicism were determined via a thorough literature search (PubMed). The keywords used in the search were as follows: preimplantation embryo development, blastocyst stage, embryonic aneuploidy, meiotic chromosome malsegregation, post-zygotic chromosome malsegregation, comprehensive chromosome screening, array comparative genomic hybridization, single-nucleotide polymorphism array, next-generation sequencing, embryo mosaicism and implantation of mosaic embryos. Relevant articles written in English and published up to March 2018 were reviewed. Different types of aneuploidy, including some complex forms, are capable of persisting to the blastocyst stage. As expected, euploidy rates decreased with advancing female age, whereas uniform aneuploidy increased. Analysis of multiple cells from individual blastocysts revealed that most of those classified 'abnormal' contained no euploid cells (due to meiotic errors arising in the gametes and therefore present in every cell), some having additional mosaic (post-fertilization, mitotic) errors. Blastocysts with a mix of normal and aneuploid cells were observed less frequently than other classes of embryo. The transfer of embryos with diploid-aneuploid mosaic biopsy specimens is reportedly associated with higher miscarriage and lower implantation rates, compared to embryos in which only euploid cells are detected. Detailed investigations into the chromosome constitution of human blastocysts suggest that a significant proportion is euploid in every cell, although the exact fraction is strongly influenced by female age. These findings do not support the notion that mosaic chromosome abnormalities are a natural part of embryo development. Mosaic aneuploidies, arising post-zygotically, were detected by various different comprehensive molecular cytogenetic methods, suggesting that the majority of these represent genuine findings. However, it remains possible that certain comprehensive molecular cytogenetic methods may carry a risk of mosaicism being incorrectly assigned, in a minority of samples, as a result of technical artifact. This may be a consequence of degraded DNA in the trophectoderm biopsy or other technical issues. According to published studies, blastocysts considered to have uniform aneuploidy and, to a lesser extent, those with mosaic abnormalities were associated with poorer clinical outcomes in comparison with euploid embryos.

Sensitive and specific detection of mosaic chromosomal abnormalities using the Parent-of-Origin-based Detection (POD) method

BackgroundMosaic somatic alterations are present in all multi-cellular organisms, but the physiological effects of low-level mosaicism are largely unknown. Most mosaic alterations remain undetectable with current analytical approaches, although the presence of such alterations is increasingly implicated as causative for disease.ResultsHere, we present the Parent-of-Origin-based Detection (POD) method for chromosomal abnormality detection in trio-based SNP microarray data. Our software implementation, triPOD, was benchmarked using a simulated dataset, outperformed comparable software for sensitivity of abnormality detection, and displayed substantial improvement in the detection of low-level mosaicism while maintaining comparable specificity. Examples of low-level mosaic abnormalities from a large autism dataset demonstrate the benefits of the increased sensitivity provided by triPOD. The triPOD analyses showed robustness across multiple types of Illumina microarray chips. Two large, clinically-relevant datasets were characterized and compared.ConclusionsOur method and software provide a significant advancement in the ability to detect low-level mosaic abnormalities, thereby opening new avenues for research into the implications of mosaicism in pathogenic and non-pathogenic processes.

Detectable clonal mosaicism and its relationship to aging and cancer.

In an analysis of 31,717 cancer cases and 26,136 cancer-free controls drawn from 13 genome-wide association studies (GWAS), we observed large chromosomal abnormalities in a subset of clones from DNA obtained from blood or buccal samples. Mosaic chromosomal abnormalities, either aneuploidy or copy-neutral loss of heterozygosity, of size >2 Mb were observed in autosomes of 517 individuals (0.89%) with abnormal cell proportions between 7% and 95%. In cancer-free individuals, the frequency increased with age; 0.23% under 50 and 1.91% between 75 and 79 (p=4.8×10−8). Mosaic abnormalities were more frequent in individuals with solid-tumors (0.97% versus 0.74% in cancer-free individuals, OR=1.25, p=0.016), with a stronger association for cases who had DNA collected prior to diagnosis or treatment (OR=1.45, p=0.0005). Detectable clonal mosaicism was common in individuals for whom DNA was collected at least one year prior to diagnosis of leukemia compared to cancer-free individuals (OR=35.4, p=3.8×10−11). These findings underscore the importance of the role and time-dependent nature of somatic events in the etiology of cancer and other late-onset diseases.

Somatic Chromosome Abnormalities in the Lungs of Patients with Pulmonary Arterial Hypertension

Vascular remodeling in pulmonary arterial hypertension (PAH) involves proliferation and migration of endothelial and smooth muscle cells, leading to obliterative vascular lesions. Previous studies have indicated that the endothelial cell proliferation is quasineoplastic, with evidence of monoclonality and instability of short DNA microsatellite sequences. To assess whether there is larger-scale genomic instability. We performed genome-wide microarray copy number analysis on pulmonary artery endothelial cells and smooth muscle cells isolated from the lungs of patients with PAH. Mosaic chromosomal abnormalities were detected in PAEC cultures from five of nine PAH lungs but not in normal (n = 8) or disease control subjects (n = 5). Fluorescent in situ hybridization analysis confirmed the presence of these abnormalities in vivo in two of three cases. One patient harbored a germline mutation of BMPR2, the primary genetic cause of PAH, and somatic loss of chromosome-13, which constitutes a second hit in the same pathway by deleting Smad-8. In two female subjects with mosaic loss of the X chromosome, methylation analysis showed that the active X was deleted. One subject also showed completely skewed X-inactivation in the nondeleted cells, suggesting the pulmonary artery endothelial cell population was clonal before the acquisition of the chromosome abnormality. Our data indicate a high frequency of genetically abnormal subclones within PAH lung vessels and provide the first definitive evidence of a second genetic hit in a patient with a germline BMPR2 mutation. We propose that these chromosome abnormalities may confer a growth advantage and thus contribute to the progression of PAH.

Low‐level mosaicism of trisomy 14: Phenotypic and molecular characterization

Trisomy 14 mosaicism is a rare cytogenetic abnormality with a defined and recognizable clinical phenotype. We present a detailed clinical history and physical findings of five patients with low-level mosaicism of trisomy 14 detected by array-based comparative genomic hybridization (array-CGH) analysis or by routine chromosome analysis. These patients exhibited growth and developmental delays with variable severity, congenital anomalies, pigmentary skin lesions, and dysmorphic features. The phenotype of our patients was compared with previously described cases. This report suggests that trisomy 14 mosaicism may be more common than has been previously appreciated and also illustrates the important application of array-CGH to detect low-level mosaic chromosome abnormalities. We predict that a wider application of the array-CGH technology will significantly increase the detection rate of low-level mosaicism and will subsequently improve our ability to provide a diagnosis for patients with dysmorphic features, congenital anomalies, and developmental delay.

Detection of low‐level mosaicism by array CGH in routine diagnostic specimens

The advent of microarray-based comparative genomic hybridization (array CGH) promises to revolutionize clinical cytogenetics because of its ability to rapidly screen the genome at an unprecedented resolution. Yet, the ability of array CGH to detect and evaluate low-level mosaicism is not known. Our laboratory has analyzed over 3,600 clinical cases with the SignatureChip which we developed for the detection of microdeletions, microduplications, aneuploidy, unbalanced translocations, and subtelomeric and pericentromeric copy number alterations. Here, we report 18 cases of mosaicism detected by array CGH in a routine diagnostic setting, 14 of which were not known to us at the time of the analysis. These 14 cases represent approximately 8% of all abnormal cases identified in our laboratory. For each case, fluorescence in situ hybridization (FISH) analysis was performed on PHA-stimulated cultures after mosaic chromosome abnormalities were suspected by array CGH. In all cases, FISH confirmed the mosaic chromosome abnormalities which included a variety of marker chromosomes, autosomal trisomies, terminal and interstitial deletions, and derivative chromosomes. Interestingly, confirmatory FISH analyses on direct blood smears indicated that the percentage of abnormal cells in unstimulated cultures was in some cases different than that found in PHA-stimulated cells. We also report the detection of a previously unsuspected case of an isochromosome 12p (associated with Pallister-Killian syndrome) by array CGH using genomic DNA extracted from peripheral blood. These results support a growing body of data that suggests that stimulated peripheral blood cultures likely distort the percentage of abnormal cells and may, for some chromosome abnormalities, make their detection unlikely by conventional analysis. Thus, array CGH, which is based on genomic DNA extracted directly from uncultured peripheral blood, may be more likely to detect low-level mosaicism for unbalanced chromosome abnormalities than traditional cytogenetic techniques.

Confirmation of mosaicism and uniparental disomy in amniocytes, after detection of mosaic chromosome abnormalities in chorionic villi

Chromosome mosaicism is detected in about 1-2% of chorionic villi samples (CVS), and may be due to a postzygotic nondisjunction event generating a trisomic cell line in an initially normal conceptus (mitotic origin) or the postzygotic loss of one chromosome in an initially trisomic conceptus (meiotic origin and trisomy rescue). Depending on the distribution of the abnormal cell line, the mosaic can be confined to the placenta (CPM) or generalised to the fetus (TFM, true fetal mosaicism). Trisomy rescue could theoretically be associated with a 33.3% probability of uniparental disomy (UPD) in the fetus. The aim of this study was to determine the risk of fetal involvement in a cohort of numerical and structural chromosome mosaics revealed in chorionic villi by means of combined direct and long-term culture analyses; we also determined the incidence of UPD associated with mosaic aneuploidies and supernumerary markers involving imprinted chromosomes. A total of 273 of a consecutive series of 15,109 CVS evaluated during a period of 5 years showed a mosaic condition in direct preparations and/or long-term cultures; confirmatory amniocentesis was performed in 203 cases. The abnormal cell line was extended to the fetus in 12.8% cases in terms of structural and numerical abnormalities involving autosomes and sex chromosomes; the risk of TFM varied and depended on the placental tissue distribution of the abnormal cell line. One of the 51 cases in which the mosaic involved an imprinted chromosome showed UPD, thus indicating a risk of 1.96%.

Mosaic Turner's Syndrome With Unusual Manifestations

<h3>To the Editor.—</h3> Mosaic variants of Turner's syndrome have been reported to occur in about 20% of the cases and they show wide variations in their phenotypes.^1-4This report represents a case of Turner's syndrome with mosaic chromosomal abnormalities and unusual manifestations such as normal ovaries, multiple cardiac anomalies, and partial situs inversus. To our knowledge, the latter two manifestations have not been reported previously. <h3>Report of a Case.—</h3> An infant weighing 2,500 gm was born at 36 weeks gastation on Dec 19, 1969, to a 31-year-old gravida 7, para 2, white mother. Bradycardia (40 to 60 beats per minute) of the baby had been noted in prenatal examinations. Moderate cyanosis, hepatosplenomegaly, and a grade 4/6 systolic murmur were present at birth. The Apgar score was 5. The baby had low-set ears, a low hair line, a short and webbed neck, a shield chest with widely spaced nipples, and