Detection Of Trisomy Research Articles

Clinical Performance of Cell-Free Fetal DNA Testing for Fetal Aneuploidies and Subchromosomal Deletions/Duplications in a Cohort of 19,531 Pregnancies

Objective: We aim to assess the clinical performance of cell-free fetal DNA (cffDNA) testing for detecting common fetal aneuploidies as well as subchromosomal deletions/duplications and explore the pregnancy decisions in screen-positive cases. Methods: A cohort of 19,531 pregnant women was offered cffDNA testing for detection of trisomies 21, 18, and 13 (T21, T18, and T13); sex chromosome aneuploidies (SCAs); and subchromosomal deletions/duplications. Screen-positive cases were confirmed by karyotyping and single-nucleotide polymorphism array analysis. Results: A total of 47 cases failed the test. The overall screen-positive rate of chromosomal abnormalities was 1.07% (208/19,484), including 57 cases with T21, 18 cases with T18, 7 cases with T13, 106 cases with SCAs, and 20 cases of subchromosomal deletions/duplications. Positive predictive values were 91.30% (42/46), 38.46% (5/13), 33.33% (2/6), 41.33% (31/75), and 27.78% (5/18), respectively. There was no significant difference in the screening of fetal chromosomal aneuploidies in the high-risk group compared with the low-risk group (P > 0.05). All of the pregnant women who had confirmed fetal T21, T18, or T13 terminated their pregnancies, except for a case of T13 mosaic, whereas 45.16% (14/31) of women with fetal SCAs continued their pregnancies. Furthermore, 17 pregnant women with positive screens for T21, T18, or T13 without a subsequent diagnosis chose to terminate their pregnancy, whereas 29 of 31 women with SCAs chose to continue their pregnancies. Conclusions: CffDNA testing exhibited good screening accuracy for T21, T18, and T13 and also contributed to detecting fetal SCAs and subchromosomal deletions/duplications. Pregnant women with fetal 47, XXX or 47, XYY were more willing to terminate their pregnancy than those with fetal 45, X or 47, XXY.

Fetal fraction of cell-free DNA in pregnancies after fresh or frozen embryo transfer following assisted reproductive technologies.

Is the fetal fraction (FF) of circulating cell-free DNA (cfDNA) affected in pregnancies following ART treatment with either fresh or frozen embryo transfer (ET) compared with natural conception? This study shows a significant reduction in the FF in ART patients compared with naturally conceived pregnancies, which seems to be more pronounced after fresh ET compared with frozen ET. Non-invasive prenatal testing (NIPT) is based on cfDNA in maternal blood, of which about 10% is of placental origin and thus represents the fetal karyotype. Validation studies have demonstrated a high sensitivity, specificity and positive predictive value of NIPT for the detection of fetal trisomy 21, 18 and 13. Nevertheless, the FF of cfDNA is an important factor for NIPT test accuracy. Several studies have found a reduction in FF for pregnancies following ART in comparison with natural conception. However, knowledge on how the FF is affected in ART pregnancies after fresh ET compared with frozen ET is very limited. The study was designed as a case-control study. A total of 54 women with an ongoing pregnancy following ART treatment were included. After exclusion for different reasons, statistical analyses were based on 23 NIPT samples from pregnant women treated with fresh ET and 26 NIPT samples from pregnant women treated with frozen-thawed ET in a modified natural cycle. Women were included between February 2018 and November 2018. The results were compared with a control group of 238 naturally conceived pregnancies with a high-risk result from the combined first trimester screening (cFTS). The study included women from the Fertility Clinics at Copenhagen University Hospital Hvidovre and Copenhagen University Hospital Rigshospitalet. Blood samples for NIPT analysis were drawn between 11 + 0 and 14 + 2 weeks of gestation and were all analyzed at the NIPT Center at Copenhagen University Hospital Hvidovre. The NIPT-test was performed by massive-parallel whole-genome sequencing. The FF was determined using the SeqFF algorithm. We found a reduction in FF in ART patients compared with naturally conceived pregnancies, and the reduction was more pronounced for ART pregnancies after fresh ET (mean FF = 0.049) compared with frozen ET (mean FF = 0.063) (multivariate analysis adjusted for maternal BMI, P = 0.02). Another multivariate analysis, adjusted for BMI and multiples of median (MoM) values for pregnancy-associated plasma protein-A (PAPP-A), demonstrated a significantly reduced FF for ART pregnancies (mean FF = 0.056) compared with naturally conceived pregnancies (mean FF = 0.072) (P < 0.0001). We found that FF was significantly reduced with increasing maternal BMI (P < 0.0001) and with decreasing MoM values of PAPP-A (P = 0.003). A limitation of our study design was the relatively small sample size. Another limitation was that the control group was not matched with the ART-treated women. The majority of the women from the control group had a high risk from cFTS, thereby their biochemical markers were diverging. However, the biochemical markers for the ART-treated women with fresh or frozen ET were not divergent within the subgroups. Concurrent with other studies demonstrating a reduced FF for singleton pregnancies after ART treatment compared with naturally conceived pregnancies, we found a reduction in FF between the two groups. This is one of the first studies to examine FF in ART pregnancies after fresh ET compared with frozen ET, hence the existing knowledge is limited. We find that FF is even more reduced in pregnancies following fresh ET compared with frozen ET, which might possibly reflect the predisposition of being small for gestational age after fresh ET compared with natural cycle frozen ET. The study was supported by the A.P. Møller og Hustru Chastine Mc-Kinney Møllers Fond til almene Formaal (the A.P. Møller Foundation for General Purposes). All authors declare no conflicts of interest. NA.

The prognostic significance of POD24 in 106 cases with splenic marginal lymphoma with bone marrow invasion

目的探讨24个月内疾病进展（POD24）对伴骨髓侵犯的脾边缘区淋巴瘤（SMZL）患者总生存的影响，比较POD24与非POD24患者的临床特征。方法回顾性分析2002年1月至2017年1月中国医学科学院血液病医院收治的有治疗指征且经过正规治疗的伴骨髓侵犯的SMZL患者，选取随访时间足以对POD24进行判断的患者（排除因非进展因素发生死亡的患者），进行预后评估及临床特征比较。结果共入组患者106例，中位年龄57（25～79）岁。①临床特征：全部患者均有骨髓侵犯和脾肿大，其中巨脾59.4％（63/106），肝大14.8％（15/101）；复杂核型22.7％（18/79），13q缺失5.1％（4/78），11q缺失1.3％（1/72），17p缺失2.5％（2/80），12号染色体三体（CEP12）7.5％（4/53）。②生存分析：单因素分析提示POD24、HGB<100 g/L以及CEP12为与总生存相关的不良预后因素；多因素分析提示仅POD24有独立预后意义［HR＝20.116（95％CI 2.226～181.820），P＝0.008］。③亚组分析：POD24患者较非POD24患者起病时纵隔淋巴结肿大的发生率（63.6％对18.9％，P＝0.005）及复杂核型发生率（50.0％对17.9％，P＝0.024）明显增高；腹腔淋巴结肿大、贫血、血小板减少、白蛋白下降以及乳酸脱氢酶增高的发生率在POD24患者中更高，与非POD24患者相比差异无统计学意义（P>0.05）。结论POD24为影响伴骨髓侵犯的SMZL患者总生存的独立预后不良因素，起病时伴纵隔淋巴结肿大及复杂核型的患者发生POD24比例更高。

Rapid prenatal aneuploidy detection of BACs-on-Beads assay in 4961 cases of amniotic fluid samples

Objective To evaluate the diagnostic value of the BACs-on-Beads (BoBs) assay for the rapid diagnosis of common aneuploidies and microdeletions. Methods A total of 4961 pregnant women admitted to the Wuxi Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University from January 2017 to March 2019 were enrolled. BoBs assay and conventional karyotyping were applied to detect amniotic fluid samples with various indications for prenatal diagnosis. Chromosomal microarray analysis (CMA) and maternal cell contamination (MCC) tests were used for further validation. Results The overall abnormality detection rates (BoBs associated with karyotyping) were 4.25% (211/4961). The prenatal diagnosis success rate of karyotyping was 99.4% (4933/4961), compared to 100% (4961/4961) using the BoBs assays. The BoBs assay was similar to karyotyping for the detection of trisomy 21 (1.01%, 50/4961), trisomy 18 (0.40%, 20/4961), trisomy 13 (0.04%, 2/4961), and sex chromosomal aneuploidies (0.15%, 12/4961). The BoBs assay also identified sex chromosomal microduplications/microdeletions (1.73%, 86/4961), 22q11.2 microdeletions/microduplications (0.1%, 5/4961), and Cri du Chat syndrome (0.02%, 1/4961) which were missed by karyotyping. The sensitivity for the detection of numerical chromosomal abnormalities of the BoBs assay and karyotyping analysis was 100% (95/95, 95% CI: 1.0–1.0) and 98.9% (94/95, 95% CI: 0.969–1.010), respectively. The sensitivity of detecting structural chromosomal abnormalities in the BoBs assay was significantly higher than those of karyotyping (79.3%, 92/116, 95% CI: 0.718–0.868) versus 21.6% (25/116, 95% CI: 0.140–0.291) (p < .01). Conclusions The BoBs assay is a reliable and rapid test for the detection of common aneuploidies and nine microdeletion syndromes with high sensitivity and accuracy in prenatal diagnosis. The assay can compensate for the limitations of karyotyping analysis.

Is there still a role for nuchal translucency measurement in the changing paradigm of first trimester screening?

ObjectivesTo give an overview of the genetic and structural abnormalities occurring in fetuses with nuchal translucency (NT) measurement exceeding the 95th percentile at first‐trimester screening and to investigate which of these abnormalities would be missed if cell‐free fetal DNA (cfDNA) were used as a first‐tier screening test for chromosomal abnormalities.MethodsThis is a national study including 1901 pregnancies with NT≥95th percentile referred to seven university hospitals in the Netherlands between 1 January 2010 and 1 January 2016. All cases with unknown pregnancy outcome were excluded. Results of detailed ultrasound examinations, karyotyping, genotyping, pregnancy and neonatal outcomes, investigation by a clinical geneticist and post‐mortem investigations were collected.ResultsIn total, 821 (43%) pregnancies had at least one abnormality. The rate of abnormalities was 21% for fetuses with NT between 95th and 99th percentile and 62% for fetuses with NT≥99th percentile. Prevalence of single‐gene disorders, submicroscopic, chromosomal and structural abnormalities was 2%, 2%, 30% and 9%, respectively.ConclusionAlthough cfDNA is superior to the combined test, especially for the detection of trisomy 21, 34% of the congenital abnormalities occurring in fetuses with increased NT may remain undetected in the first trimester of pregnancy, unless cfDNA is used in combination with fetal sonographic assessment, including NT measurement.

Detection of fetal trisomy and single gene disease by massively parallel sequencing of extracellular vesicle DNA in maternal plasma: a proof-of-concept validation

BackgroundDuring human pregnancy, placental trophectoderm cells release extracellular vesicles (EVs) into maternal circulation. Trophoblasts also give rise to cell-free DNA (cfDNA) in maternal blood, and has been used for noninvasive prenatal screening for chromosomal aneuploidy. We intended to prove the existence of DNA in the EVs (evDNA) of maternal blood, and compared evDNA with plasma cfDNA in terms of genome distribution, fragment length, and the possibility of detecting genetic diseases.MethodsMaternal blood from 20 euploid pregnancies, 9 T21 pregnancies, 3 T18 pregnancies, 1 T13 pregnancy, and 2 pregnancies with FGFR3 mutations were obtained. EVs were separated from maternal plasma, and confirmed by transmission electronic microscopy (TEM), western blotting, and flow cytometry (FACS). evDNA was extracted and its fetal origin was confirmed by quantitative PCR (qPCR). Pair-end (PE) whole genome sequencing was performed to characterize evDNA, and the results were compared with that of cfDNA. The fetal risk of aneuploidy and monogenic diseases was analyzed using the evDNA sequencing data.ResultsEVs separated from maternal plasma were confirmed with morphology by TEM, and protein markers of CD9, CD63, CD81 as well as the placental specific protein placental alkaline phosphatase (PLAP) were confirmed by western blotting or flow cytometry. EvDNA could be successfully extracted for qPCR and sequencing from the plasma EVs. Sequencing data showed that evDNA span on all 23 pairs of chromosomes and mitochondria, sharing a similar distribution pattern and higher GC content comparing with cfDNA. EvDNA showed shorter fragments yet lower fetal fraction than cfDNA. EvDNA could be used to correctly determine fetal gender, trisomies, and de novo FGFR3 mutations.ConclusionsWe proved that fetal DNA could be detected in EVs separated from maternal plasma. EvDNA shared some similar features to plasma cfDNA, and could potentially be used to detect genetic diseases in fetus.

Isolated Echogenic Cardiac Focus: Assessing Association with Trisomy 21 by Combining Results from a Prenatal Center with a Bayesian Meta-Analysis.

Objective To investigate the clinical relevance of an isolated echogenic cardiac focus (iECF) as a marker for trisomy 21 using a large second-trimester collective including a low-risk subgroup. Materials and Methods We retrospectively evaluated 1 25 211 pregnancies from 2000–2016 and analyzed all iECF cases with regard to chromosomal anomalies. It consisted of an early second-trimester collective from 14+0−17+6 weeks (n=34 791) and a second-trimester anomaly scan collective from 18+0–21+6 weeks. Two a priori risk subgroups (high and low risk) of the latter were built based on maternal age and previous screening test results using a cut-off of 1:300. Likelihood ratios (LR) of iECF for the detection of trisomy 21, trisomy 13, trisomy 18 and structural chromosomal anomalies were estimated. Results In total, 1 04 001 patients were included. An iECF was found in 4416 of 1 02 847 euploid fetuses (4.29%) and in 64 of 557 cases with trisomy 21 (11.49%) giving a positive LR of 2.68 (CI: 2.12–3.2). The sensitivity was 11.5% at a false-positive rate of 4.29% (CI:4.17–4.42) with p≤0.01%. In the high-and low-risk subgroups, the prevalence of iECF was comparable: 5.08% vs. 5.05%. The frequency of trisomy 21 was 0.39%, 98/24 979 vs 0.16%, 69/44 103. LR+was 3.86 (2.43–5.14) and 2.59 (1.05–4). For both subgroups the association of iECF with trisomy 21 was statistically significant. The prevalence of structural chromosomal anomalies in the second-trimester anomaly scan collective was 0.08% (52/68 967), of which 2 showed an iECF. Conclusion The detection of an iECF at the time of 14+0–21+6 weeks significantly increases the risk for trisomy 21 in the high-risk and in the low-risk subgroups and does not statistically change the risks for trisomy 13/18 or structural abnormalitie.

Result of Prospective Validation of the Trisomy Test® for the Detection of Chromosomal Trisomies.

Noninvasive prenatal testing (NIPT) is one of the most common prenatal screening tests used worldwide. Trisomy Test® belongs to NIPT tests based on low-coverage whole-genome sequencing. In our prospective study, 7279 samples of pregnant women collected during approximately two years were analyzed. In this cohort, 117 positive cases for trisomies 21, 18, and 13 were reported. An in-house designed bioinformatic pipeline and proprietary biostatistical approach was used for the detection of trisomies. The pooled sensitivity and specificity of our test reached 99.12% and 99.94%, respectively. The proportion of repeatedly uninformative results after repeated blood draws was 1.11%. Based on the presented results, we can confirm that the Trisomy Test® is fully comparable with other commercial NIPT tests available worldwide.

Noninvasive Prenatal Testing Using Next Generation Sequencing: Pilot Experience of the D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology

In recent years, noninvasive prenatal testing (NIPT) for fetal chromosomal abnormalities has come into wide use. NIPT allows detection of fetal chromosomal abnormalities without invasive sampling of fetal material: by analyzing cell-free fetal DNA in maternal blood. Here, we report on the pilot results of using NIPT at the D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology (St. Petersburg, Russia). The results obtained in 149 blood samples from pregnant women proved the clinical value of NIPT for detection of fetal trisomies 21, 18, and 13. Aneuploidy was detected in 20 out of 149 samples. Among these 20 aneuploid cases, trisomy 21 amounted to 60%, trisomy 18 – to 25%, and trisomy 13 to 5% of cases. In one sample, double aneuploidy involving chromosomes 13 and 21 was detected, and in one case, trisomy X was identified. In 100% of cases, aneuploidies were confirmed by prenatal karyotyping advocating for the absence of false positive NIPT results. The observed high specificity was consistent with the declared test specificity level of >99.9%. The sensitivity of the method was 100%. To summarize, NIPT is a new reliable technique that will probably soon replace the conventional first trimester combined screening. The only disadvantages of implementing NIPT in Russia are its relatively high cost and the lack of coverage by insurance companies. NGS-based NIPT is suitable for clinical practice and will be widely deployed due to a further cost decrease.

15. THE INCIDENCE AND ORIGIN OF CHROMOSOME ANEUPLOIDIES IN HIGH QUALITY KARYOMAPPING SNP PROFILES

Introduction Karyomapping serves as the universal PGT-M platform based on Mendelian inheritance of SNP markers which assist in identifying haplotypes linked to a mutated gene. Moreover, karyomapping enables detection of aneuploidies by interrogating genome-wide SNP data in embryo samples and assists in a selection of euploid embryos for a transfer. We have found out that high quality SNP profiles, characterized by overall call rate ≥95%, might be reliably used for the identification of mitotic chromosome errors, aneuploidies in mosaic or segmental chromosome aberrations. Since karyomapping utilizes parental DNA samples, the origin of identified aneuploidies might be tracked in embryo samples. Our study describes the incidence and origin of aneuploidy, aiming at segmental chromosomes errors and mosaic aneuploidies which represents hot topic in respect to their clinical significance. Material & methods SNP data obtained by HumanKaryomap-12 BeadChips, Illumina SNP arrays were analysed in 2085 trophectoderm (TE) samples derived from 319 couples requesting PGT-M. Mean maternal age was 32.4 ± 4.5. Based on the karyomapping SNP data QC metrics, mean and median call rate was 90.4% and 95%, respectivelly. In total, 936 embryos fulfilled criteria of high quality SNP profiles (call rate ≥95%). A subset of twenty TE samples detected with mitotic trisomies, mosaic aneuploidies and segmental chromosome abnormalities by karyomapping were reanalysed using PG-Seq™ kit, which is the fully validated PGT-A platform in our laboratory. We have observed full concordance between both platforms. Results Analysing 936 high quality TE samples, we have observed that 617 (65.9%) of them were euploid. The remaining 319 TE samples were detected with the following aberrations: a) only aneuploid 234 (73.4%) samples; b) only mosaic 43 (13.5%) samples; c) aneuploid + mosaic 42 (13.2%) samples. In other words, overall mosaicism rate in 936 TE samples was 9.1% (85/936) in our study. This number tend to be underestimated, since we did not report low level mosaicism due to a lack of proper sensitivity validation of the karyomapping in last years. We have detected segmental aneuploidies in 12.6% (118/936) of TE samples, 56 (47.4%) of them were present in mosaic form. Regarding the origin of aneuploidies, 87% (220/253) of whole chromosome aneuploidies affected maternal chromosomes, the ratio difference was even more highlighted in whole chromosome trisomies (93%; 91 out of 98). In contrast, segmental chromosome abnormalities affected more frequently paternal chromosomes (61,3%; 38 out of 62). Notably, this trend towards paternally related segmental aneuploidies were also followed in segmental chromosome abnormalities in mosaic (60,7%; 34 out of 56). Conclusions 1. According to our validation, high quality SNP profiles (overall SNP call rate ≥95%) enables detection of mitotic trisomies, mosaicism at 25% sensitivity level and segmental chromosome abnormalities (in 5Mb resolution). 2. Meiotic whole chromosome trisomies prevail over mitotic whole chromosome trisomies (in full or mosaic state) in the ratio 4:1. 3. Segmental chromosome abnormalities (in full or mosaic state) affects more frequently paternal chromosomes (P value 4. Whole chromosome aneuploidies in mosaic affect equally maternal and paternal chromosomes.

GENOME-WIDE PROFILING AND HAPLOTYPING OF CELL-FREE DNA ENABLING COMBINED NON-INVASIVE PRENATAL DIAGNOSIS OF INHERITED MONOGENIC DISEASES AND ANEUPLOIDY

Non-invasive prenatal diagnosis for the common trisomies 21, 18 and 13 has become standard of care in many countries world-wide. We developed methods that enable not only the detection of those common trisomies but leverage the identification of all fetal and maternal genomic and chromosomal disorders. As of July 2017 NIPT is reimbursed for all pregnant women in Belgium, so the outcome of this screening program will be presented, as well as our innovative NIPT pipeline. Non-invasive prenatal diagnosis (NIPD) for monogenic diseases has only been tested on a small number of genes using a locus-specific approach and has not become widely implemented. Here, we present a new method that enables the deduction of fetal haplotype to detect inherited monogenic diseases and aneuploidy. Using targeted sequencing in a genome-wide level of resolution, parental haplotype inheritance in the fetus was determined non-invasively with high accuracy based on cfDNA haplotyping analysis. In all clinical cases, the haplotypes blocks linked with the wild-type allele were correctly identified, and a maternally inherited trisomy case was properly deduced. The method is generic and can deduce the presence or absence of any inherited disease allele. Moreover, it leverage the genome wide fetal haplotype at relatively low cost. Since genome wide haplotyping of embryos (karyomapping or OnePGT) enables the reconstruction of genome-wide haplotypes, copy-number profiles and are able to determine the segregational origin of haplotypes, those methodologies are being implemented as a generic method for preimplantation genetic testing (PGT) in many IVF laboratories. This non-invasive prenatal haplotyping approach can also confirm that the fetus is derived from the embryo transferred and will replace invasive prenatal confirmation testing.

Computational framework for targeted high-coverage sequencing based NIPT.

Non-invasive prenatal testing (NIPT) enables accurate detection of fetal chromosomal trisomies. The majority of publicly available computational methods for sequencing-based NIPT analyses rely on low-coverage whole-genome sequencing (WGS) data and are not applicable for targeted high-coverage sequencing data from cell-free DNA samples. Here, we present a novel computational framework for a targeted high-coverage sequencing-based NIPT analysis. The developed framework uses a hidden Markov model (HMM) in conjunction with a supplemental machine learning model, such as decision tree (DT) or support vector machine (SVM), to detect fetal trisomy and parental origin of additional fetal chromosomes. These models were developed using simulated datasets covering a wide range of biologically relevant scenarios with various chromosomal quantities, parental origins of extra chromosomes, fetal DNA fractions, and sequencing read depths. Developed models were tested on simulated and experimental targeted sequencing datasets. Consequently, we determined the functional feasibility and limitations of each proposed approach and demonstrated that read count-based HMM achieved the best overall classification accuracy of 0.89 for detecting fetal euploidies and trisomies on simulated dataset. Furthermore, we show that by using the DT and SVM on the HMM classification results, it was possible to increase the final trisomy classification accuracy to 0.98 and 0.99, respectively. We demonstrate that read count and allelic ratio-based models can achieve a high accuracy (up to 0.98) for detecting fetal trisomy even if the fetal fraction is as low as 2%. Currently, existing commercial NIPT analysis requires at least 4% of fetal fraction, which can be possibly a challenge in case of early gestational age (<10 weeks) or high maternal body mass index (>35 kg/m2). More accurate detection can be achieved at higher sequencing depth using HMM in conjunction with supplemental models, which significantly improve the trisomy detection especially in borderline scenarios (e.g., very low fetal fraction) and enables to perform NIPT even earlier than 10 weeks of pregnancy.

The contingent use of cell-free fetal DNA for prenatal screening of trisomies 21, 18, 13 in pregnant women within a national health service: A budget impact analysis.

ObjectiveNon-invasive prenatal testing (NIPT) based on cell-free fetal DNA (cffDNA) is highly accurate in the detection of common fetal autosomal trisomies. Aim of this project was to investigate short-term costs and clinical outcomes of the contingent use of cffDNA for prenatal screening of trisomies 21, 18, 13 within a national health service (NHS).MethodsAn economic analysis was developed from the perspective of the Italian NHS to compare two possible scenarios for managing pregnant women: women managed according to the Standard of Care screening (SoC) vs a cffDNA scenario, where Harmony Prenatal Test was introduced as a second line screening choice for women with an “at risk” result from SoC screening.ResultsThe introduction of cffDNA as a second line screening test, conditional to a risk ≥ 1:1,000 from SoC screening, showed a 3% increase in the detection of trisomies, with a 71% decrease in the number of invasive tests performed. Total short-term costs (pregnancy management until childbirth) decreased by € 19 million (from € 84.5 to 65.5 million).ConclusionThe adoption of the Harmony Prenatal Test in women resulting at risk from SoC screening, implied a greater number of trisomies detection, together with a reduction of the healthcare costs.

Non-invasive prenatal testing for detection of trisomy 13, 18, 21 and sex chromosome aneuploidies in 8594 cases.

Cell-free fetal DNA has been widely used in prenatal genetic testing during recent years. We explored the feasibility of non-invasive prenatal testing (NIPT) for analysis of common fetal aneuploidies among pregnancies in northwest China. A total of 8594 maternal blood samples were collected from October 2014 to December 2017 in the Department of Obstetrics and Gynecology at the First Affiliated Hospital of the Air Force Medical University. Cases with positive screening results by NIPT detection were validated using karyotype analysis. Of 8594 clinical pregnancies, 88 had positive NIPT results and 78 of 88 (88.6%) positive NIPT results were shown to be false-positive by amniotic fluid puncture and chromosome karyotyping analysis. There were 44 cases (49.44%) with trisomy 21, 18, and 13 syndromes (30 cases of trisomy 21, 9 cases of trisomy 18, and 5 cases of trisomy 13). There were 44 cases (50.56%) with sex chromosome abnormalities, including 11 cases with Turner syndrome (45, X), 17 cases with Triple X syndrome (47, XXX), 2 cases with Klinefelter syndrome (47, XXY), and 14 cases with 47, XYY syndrome (47, XYY). The accuracy, specificity, high efficiency, and acceptance of NIPT can effectively avoid birth defects and improve the quality of the birth population. We should deepen mining and analysis of the clinical data and explore ways to use NIPT. It is recommended that the NIPT guidelines be extended to low-risk patients to further explore the impact of a significant increase in screening.

Association Between Fetal Fraction on Cell-Free DNA Testing and First-Trimester Markers for Pre-eclampsia

(Abstracted from Ultrasound Obstet Gynecol 2018;52:722–727) Screening for chromosomal abnormalities based on cell-free fetal DNA (cfDNA) has been clinically available since 2011. While highly accurate for detection of trisomy 21, the accuracy of the test depends on the proportion of the total cfDNA that contains fetal genetic material (ie, the fetal fraction [FF]), with accuracy being higher when the FF is high and lower when the FF is low.

The significance of trisomy 7 mosaicism in noninvasive prenatal screening

BackgroundThis study was an evaluation of the role of noninvasive prenatal testing (NIPT) in the detection of trisomy 7 in prenatal diagnosis.MethodA total of 35 consecutive cases underwent screening for trisomies by cell-free DNA testing between April 2015 and November 2017 due to suspicious NIPT results; these cases represented 0.11% of patients (35/31,250) with similar frequencies of abnormal results among the laboratories performing the tests. NIPT was offered to further screen for common fetal chromosomal abnormalities. Karyotype analysis, chromosomal microarray analysis (CMA), and next-generation sequencing (NGS) were used to detect 20, 14, and 25 patients, respectively, who accepted confirmatory diagnostic testing.ResultsHigh-risk results by NIPT were recorded for trisomy 7 alone in 29 women: dual aneuploidy in 4 patients and multiple aneuploidy in 2 patients. Karyotype analysis of amniotic fluid cells was normal in all 20 pregnancies, suggesting a probability of confined placental mosaicism. Further CMA data were obtained in 14 of the cases mentioned above, and 2 fetuses were detected with positive results with copy number variation. The NGS results suggested that all these samples were placental chimerisms of chromosome 7, except for one sample that was found to be an additional chimerism of chromosome 2, which was also consistent with the NIPT result.ConclusionOur results may be useful for the counseling of pregnant women in the detection of trisomy 7 by NIPT.

Application of Differentially Methylated Loci in Clinical Diagnosis of Trisomy 21 Syndrome.

To determine the diagnostic precision of using different sets of fetal-specific methylation markers with methylation-sensitive restriction enzyme-quantitative polymerase chain reaction (MSRE-qPCR) for detection of trisomy 21 (T21). The diagnostic value for trisomy 21 of differential methylation of HLCS, C21orf25, and RASSF1A (a fetal-specific internal control) was examined by MSRE-qPCR. The combined marker set of HLCS and RASSF1A achieved accurate quantification of fetal-specific chromosome 21 and was an excellent marker for detecting the presence of three copies of chromosome 21. MSRE-qPCR correctly identified three cases of fetal T21 from 11 clinical samples, which were 100% consistent with karyotyping results. In addition, this method was able to detect fetal-specific, T21-derived, cell-free fetal DNA at concentrations as low as 0.1%. Evaluation of the HLCS and RASSF1A fetal-specific methylation marker set by MSRE-qPCR could be a highly sensitive, specific, cost-effective, and noninvasive prenatal screening method for T21. This MSRE-qPCR testable marker should be considered as an alternative to next generation sequencing technology for diagnosing fetal T21.

Employ ductus venous blood flow in the early detection of trisomy 21, trisomy 18, and trisomy 13: A meta-analysis.

Background and objective:Ductus venosus (DV) Doppler has been suggested as a biomarker for the early screening of trisomy diseases. However, results from different studies have been largely inconsistent. This study aimed to investigate the relationship between DV and top 3 fetal aneuploidies by a systematical meta-analysis: trisomy 21 (T21), trisomy 18 (T18), and trisomy 13 (T13).Methods:We performed a literature search covering articles from Medline, PubMed, RePORTER, and Elsevier publications. DV-T21/T18/T13 relation data were extracted from 9, 7, and 6 previous studies, respectively, including 31,053, 28,092 and 26,721 pregnant women worldwide. Both random-effects and fixed-effect model were used to study the log odds ratio (LOR) of T21, T18, and T13 in case of DV. Four potential influential factors were studied using a multiple linear regression (MLR) model, including maternal age, data age, sample size, and population region.Results:DV was significantly related to T21, T18, and T13 (LOR = 3.44, 3.89 and 3.46; P value <2.1E-13). Significant between-study variance was observed for T21 (P value <1.71E-14), but not for T18 (P value >.05) and T13 (P value >.87). MLR results suggested that significant influential factors could include population region (P value <.0021), but not sample size, data age, and maternal age (P value >.078).Conclusions:Integrating DV could help in the detection of trisomy. However, accuracy and validity may vary depending on the population regions, which need further study.

Screening of fetal chromosomal aneuploidy diseases using noninvasive prenatal testing in twin pregnancies

ABSTRACTObjectives: This study was aimed to report the clinical characteristics of fetal chromosomal aneuploidy diseases using noninvasive prenatal testing (NIPT) in twin pregnancies and analyze the results in terms of chorionicity, conception, and fetal fraction.Methods: A total of 1160 women with twin pregnancies were recruited from 1 October 2015, to 1 August 2017. Next-generation sequencing technology was used to detect fetal aneuploidies, such as trisomy 21, trisomy 18, trisomy 13 and trisomy X.Results: Aneuploidy was detected using NIPT in 26 fetuses, among which 18 fetal aneuploidies occurred in only one fetus of the twins. The rate of aneuploidy was 1.3% for dichorionic diamniotic twins and 0.5% for monochorionic diamniotic twins, respectively. The rate of aneuploidy was 1.2% for spontaneous pregnancy group and 1.1% for assisted reproductive technologies group.Conclusion: In this study, detection of trisomy 21, trisomy 18, trisomy 13, and X abnormality in twin pregnancies was confirmed to be accurate. The aneuploidies mostly occurred in only one fetus of the twins, and trisomy 21 was the most common type. The prenatal diagnostic standard for NIPT in singleton pregnancies could perform well in twin pregnancies, which means NIPT can be popularized as routine prenatal screening in twin pregnancies.

Implementation of cell-free DNA-based non-invasive prenatal testing in a National Health Service Regional Genetics Laboratory.

Non-invasive prenatal testing (NIPT) for the detection of foetal aneuploidy through analysis of cell-free DNA (cfDNA) in maternal blood is offered routinely by many healthcare providers across the developed world. This testing has recently been recommended for evaluative implementation in the UK National Health Service (NHS) foetal anomaly screening pathway as a contingent screen following an increased risk of trisomy 21, 18 or 13. In preparation for delivering a national service, we have implemented cfDNA-based NIPT in our Regional Genetics Laboratory. Here, we describe our validation and verification processes and initial experiences of the technology prior to rollout of a national screening service. Data are presented from more than 1000 patients (215 retrospective and 840 prospective) from 'high- and low-risk pregnancies' with outcome data following birth or confirmatory invasive prenatal sampling. NIPT was by the Illumina Verifi® test. Our data confirm a high-fidelity service with a failure rate of ~0.24% and a high sensitivity and specificity for the detection of foetal trisomy 13, 18 and 21. Secondly, the data show that a significant proportion of patients continue their pregnancies without prenatal invasive testing or intervention after receiving a high-risk cfDNA-based result. A total of 46.5% of patients referred to date were referred for reasons other than high screen risk. Ten percent (76/840 clinical service referrals) of patients were referred with ultrasonographic finding of a foetal structural anomaly, and data analysis indicates high- and low-risk scan indications for NIPT. NIPT can be successfully implemented into NHS regional genetics laboratories to provide high-quality services. NHS provision of NIPT in patients with high-risk screen results will allow for a reduction of invasive testing and partially improve equality of access to cfDNA-based NIPT in the pregnant population. Patients at low risk for a classic trisomy or with other clinical indications are likely to continue to access cfDNA-based NIPT as a private test.