Significance: Chromosomal aneuploidies represent a major cause of fetal loss and birth defects. Current methods for the prenatal diagnosis of aneuploidy require invasive methods that are associated with a risk of miscarriage and other complications. Recently, noninvasive prenatal testing has been developed using cell-free fetal DNA in maternal plasma. In this study, we validated an effective method for noninvasive diagnosis of fetal aneuploidy using a semiconductor sequencer, which reduces the time and cost of sequencing. Our method is cost-effective and practical in a clinical setting with high sensitivity and specificity for the diagnosis of trisomy 13, 18, and 21 as well as sex chromosome aneuploidies.

Background: Exome sequencing (ES) is becoming more widely available in prenatal diagnosis. However, data on its clinical utility and integration into clinical management remain limited in practice. Herein, we report our experience implementing prenatal ES (pES) in a large cohort of fetuses with anomalies detected by ultrasonography using a hospital-based in-house multidisciplinary team (MDT) facilitated by a three-step genotype-driven followed by phenotype-driven analysis framework. Methods: We performed pES in 1618 fetal cases with positive ultrasound findings but negative for karyotyping and chromosome microarray analysis between January 2014 and October 2021, including both retrospective (n=565) and prospective (n=1053) cohorts. The diagnostic efficiency and its correlation to organ systems involved, phenotypic spectrum, and the clinical impacts of pES results on pregnancy outcomes were analyzed. Results: A genotype-driven followed by phenotype-driven three-step approach was carried out in all trio pES. Step 1, a genotype-driven analysis resulted in a diagnostic rate of 11.6% (187/1618). Step 2, a phenotype-driven comprehensive analysis yielded additional diagnostic findings for another 28 cases (1.7%; 28/1618). In the final step 3, data reanalyses based on new phenotypes and/or clinical requests found molecular diagnosis in 14 additional cases (0.9%; 14/1618). Altogether, 229 fetal cases (14.2%) received a molecular diagnosis, with a higher positive rate in the retrospective than the prospective cohort (17.3% vs. 12.4%, p<0.01). The diagnostic rates were highest in fetuses with skeletal anomalies (30.4%) and multiple organ involvements (25.9%), and lowest in fetuses with chest anomalies (0%). In addition, incidental and secondary findings with childhood-onset disorders were detected in 11 (0.7%) cases. Furthermore, we described the prenatal phenotypes for the first time for 27 gene-associated conditions (20.0%, 27/135) upon a systematic analysis of the diagnosed cases and expanded the phenotype spectrum for 26 (19.3%) genes where limited fetal phenotypic information was available. In the prospective cohort, the combined prenatal ultrasound and pES results had significantly impacted the clinical decisions (61.5%, 648/1053). Conclusions: The genotype-driven approach could identify about 81.7% positive cases (11.6% of the total cohort) with the initial limited fetal phenotype information considered. The following two steps of phenotype-driven analysis and data reanalyses helped us find the causative variants in an additional 2.6% of the entire cohort (18.3% of all positive findings). Our extensive phenotype analysis on a large number of molecularly confirmed prenatal cases had greatly enriched our current knowledge on fetal phenotype-genotype correlation, which may guide more focused prenatal ultrasound in the future. This is by far the largest pES cohort study that combines a robust trio sequence data analysis, systematic phenotype-genotype correlation, and well-established MDT in a single prenatal clinical setting. This work underlines the value of pES as an essential component in prenatal diagnosis in guiding medical management and parental decision making.

Objective: To evaluate the clinical performance of non‐invasive prenatal testing (NIPT) in detecting fetal copy number variants (CNVs).Methods: NIPT of cell‐free fetal DNA was performed retrospectively, using stored maternal plasma, at an average gestational age of 21.1 weeks in 117 pregnant women who had previously undergone invasive prenatal testing for chromosome microarray analysis (CMA). Among the fetal samples tested by CMA, 18 had positive results (CNVs > 1 megabase (Mb)) and 99 had negative results (CNVs < 1 Mb or without CNVs detected). The results of NIPT and CMA were then compared.Results: In the 11 CMA‐positive samples with CNVs > 5 Mb, the detection rate of CNVs was 90.9%; the one case missed by NIPT had a fetal fraction of 4.7%. For the other seven CMA‐positive samples with CNVs < 5 Mb, the detection rate was 14.3%; only one case with a 2.82‐Mb duplication was detected, with a fetal fraction of 26.7%. For 35.7% (5/14) of CNVs detected by both NIPT and CMA, the differences in fragment length were within ± 1 Mb. In this study, the overall sensitivity and specificity of NIPT for detecting CNVs > 1 Mb were 61.1% and 95.0%, respectively, with a false‐positive rate of 5.0%.Conclusions: Our results demonstrate that NIPT for common aneuploidies can detect fetal CNVs > 5 Mb with high sensitivity, provided that fetal fraction is high enough, without increasing sequencing depth. The detection power of NIPT is determined mostly by fetal fraction and CNV size. A positive NIPT screening result for CNVs must be interpreted with caution and validated by additional diagnostic study. Copyright © 2015 ISUOG. Published by John Wiley & Sons Ltd.