Abstract
Noninvasive prenatal testing by massive parallel sequencing of maternal plasma DNA has rapidly been adopted as a mainstream method for detection of fetal trisomy 21, 18 and 13. Despite the relative high accuracy of current NIPT testing, a substantial number of false-positive and false-negative test results remain. Here, we present an analysis pipeline, which addresses some of the technical as well as the biologically derived causes of error. Most importantly, it differentiates high z-scores due to fetal trisomies from those due to local maternal CNVs causing false positives. This pipeline was retrospectively validated for trisomy 18 and 21 detection on 296 samples demonstrating a sensitivity and specificity of 100%, and applied prospectively to 1350 pregnant women in the clinical diagnostic setting with a result reported in 99.9% of cases. In addition, values indicative for trisomy were observed two times for chromosome 7 and once each for chromosomes 15 and 16, and once for a segmental trisomy 18. Two of the trisomies were confirmed to be mosaic, one of which contained a uniparental disomy cell line. As placental trisomies pose a risk for low-grade fetal mosaicism as well as uniparental disomy, genome-wide noninvasive aneuploidy detection is improving prenatal management.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Lo YM, Corbetta N, Chamberlain PF et al: Presence of fetal DNA in maternal plasma and serum. Lancet 1997; 350: 485–487.
Chiu RW, Chan KC, Gao Y et al: Noninvasive prenatal diagnosis of fetal chromosomal aneuploidy by massively parallel genomic sequencing of DNA in maternal plasma. Proc Natl Acad Sci USA 2008; 105: 20458–20463.
Fan HC, Blumenfeld YJ, Chitkara U, Hudgins L, Quake SR : Noninvasive diagnosis of fetal aneuploidy by shotgun sequencing DNA from maternal blood. Proc Natl Acad Sci USA 2008; 105: 16266–16271.
Ashoor G, Syngelaki A, Wagner M, Birdir C, Nicolaides KH : Chromosome-selective sequencing of maternal plasma cell-free DNA for first-trimester detection of trisomy 21 and trisomy 18. Am J Obstet Gynecol 2012; 206: 322–325.
Bianchi DW, Platt LD, Goldberg JD, Abuhamad AZ, Sehnert AJ, Rava RP : Genome-wide fetal aneuploidy detection by maternal plasma DNA sequencing. Obstet Gynecol 2012; 119: 890–901.
Chiu RW, Akolekar R, Zheng YW et al: Non-invasive prenatal assessment of trisomy 21 by multiplexed maternal plasma DNA sequencing: large scale validity study. BMJ 2011; 342: c7401.
Dan S, Wang W, Ren J et al: Clinical application of massively parallel sequencing-based prenatal noninvasive fetal trisomy test for trisomies 21 and 18 in 11,105 pregnancies with mixed risk factors. Prenat Diagn 2012; 32: 1225–1232.
Ehrich M, Deciu C, Zwiefelhofer T et al: Noninvasive detection of fetal trisomy 21 by sequencing of DNA in maternal blood: a study in a clinical setting. Am J Obstet Gynecol 2011; 204: 205–211.
Nicolaides KH, Syngelaki A, Ashoor G, Birdir C, Touzet G : Noninvasive prenatal testing for fetal trisomies in a routinely screened first-trimester population. Am J Obstet Gynecol 2012; 207: 374–376.
Norton ME, Brar H, Weiss J et al: Non-Invasive Chromosomal Evaluation (NICE) Study: results of a multicenter prospective cohort study for detection of fetal trisomy 21 and trisomy 18. Am J Obstet Gynecol 2012; 207: 137–138.
Palomaki GE, Deciu C, Kloza EM et al: DNA sequencing of maternal plasma reliably identifies trisomy 18 and trisomy 13 as well as Down syndrome: an international collaborative study. Genet Med 2012; 14: 296–305.
Palomaki GE, Kloza EM, Lambert-Messerlian GM et al: DNA sequencing of maternal plasma to detect Down syndrome: an international clinical validation study. Genet Med 2011; 13: 913–920.
Sehnert AJ, Rhees B, Comstock D et al: Optimal detection of fetal chromosomal abnormalities by massively parallel DNA sequencing of cell-free fetal DNA from maternal blood. Clin Chem 2011; 57: 1042–1049.
Sparks AB, Struble CA, Wang ET, Song K, Oliphant A : Noninvasive prenatal detection and selective analysis of cell-free DNA obtained from maternal blood: evaluation for trisomy 21 and trisomy 18. Am J Obstet Gynecol 2012; 206: 319.
Sparks AB, Wang ET, Struble CA et al: Selective analysis of cell-free DNA in maternal blood for evaluation of fetal trisomy. Prenat Diagn 2012; 32: 3–9.
Bianchi DW, Parker RL, Wentworth J et al: DNA sequencing versus standard prenatal aneuploidy screening. N Engl J Med 2014; 370: 799–808.
Chiu RW, Cantor CR, Lo YM : Non-invasive prenatal diagnosis by single molecule counting technologies. Trends Genet 2009; 25: 324–331.
Fan HC, Quake SR : Sensitivity of noninvasive prenatal detection of fetal aneuploidy from maternal plasma using shotgun sequencing is limited only by counting statistics. PLoS One 2010; 5: e10439.
Yeang CH, Ma GC, Hsu HW et al: Genome-wide normalized score: a novel algorithm to detect fetal trisomy 21 during non-invasive prenatal testing. Ultrasound Obstet Gynecol 2014; 44: 25–30.
Straver R, Sistermans EA, Holstege H, Visser A, Oudejans CB, Reinders MJ : WISECONDOR: detection of fetal aberrations from shallow sequencing maternal plasma based on a within-sample comparison scheme. Nucleic Acids Res 2014; 42: e31.
Yao H, Zhang L, Zhang H et al: Noninvasive prenatal genetic testing for fetal aneuploidy detects maternal trisomy X. Prenat Diagn 2012; 32: 1114–1116.
Osborne CM, Hardisty E, Devers P et al: Discordant noninvasive prenatal testing results in a patient subsequently diagnosed with metastatic disease. Prenat Diagn 2013; 33: 609–611.
Choi H, Lau TK, Jiang FM et al: Fetal aneuploidy screening by maternal plasma DNA sequencing: 'false positive' due to confined placental mosaicism. Prenat Diagn 2013; 33: 198–200.
Futch T, Spinosa J, Bhatt S, de FE, Rava RP, Sehnert AJ : Initial clinical laboratory experience in noninvasive prenatal testing for fetal aneuploidy from maternal plasma DNA samples. Prenat Diagn 2013; 33: 569–574.
Gao Y, Stejskal D, Jiang F, Wang W : False-negative trisomy 18 non-invasive prenatal test result due to 48,XXX,+18 placental mosaicism. Ultrasound Obstet Gynecol 2014; 43: 477–478.
Hahnemann JM, Vejerslev LO : Accuracy of cytogenetic findings on chorionic villus sampling (CVS) – diagnostic consequences of CVS mosaicism and non-mosaic discrepancy in centres contributing to EUCROMIC 1986-1992. Prenat Diagn 1997; 17: 801–820.
Hahnemann JM, Vejerslev LO : European collaborative research on mosaicism in CVS (EUCROMIC) – fetal and extrafetal cell lineages in 192 gestations with CVS mosaicism involving single autosomal trisomy. Am J Med Genet 1997; 70: 179–187.
Hall AL, Drendel HM, Verbrugge JL et al: Positive cell-free fetal DNA testing for trisomy 13 reveals confined placental mosaicism. Genet Med 2013; 15: 729–732.
Lau TK, Jiang FM, Stevenson RJ et al: Secondary findings from non-invasive prenatal testing for common fetal aneuploidies by whole genome sequencing as a clinical service. Prenat Diagn 2013; 33: 602–608.
Mao J, Wang T, Wang BJ et al: Confined placental origin of the circulating cell free fetal DNA revealed by a discordant non-invasive prenatal test result in a trisomy 18 pregnancy. Clin Chim Acta 2014; 433C: 190–193.
Mennuti MT, Cherry AM, Morrissette JJ, Dugoff L : Is it time to sound an alarm about false-positive cell-free DNA testing for fetal aneuploidy? Am J Obstet Gynecol 2013; 209: 415–419.
Pan M, Li FT, Li Y et al: Discordant results between fetal karyotyping and non-invasive prenatal testing by maternal plasma sequencing in a case of uniparental disomy 21 due to trisomic rescue. Prenat Diagn 2013; 33: 598–601.
Srebniak MI, Diderich KE, Noomen P, Dijkman A, de VF, Opstal DV : Abnormal NIPT results concordant with the karyotype of the cytotrophoblast, but not reflecting the abnormal fetal karyotype. Ultrasound Obstet Gynecol. 2014; 44: 109–111.
Wang Y, Zhu J, Chen Y et al: Two cases of placental T21 mosaicism: challenging the detection limits of non-invasive prenatal testing. Prenat Diagn 2013; 33: 1207–1210.
Brady PD, Delle CB, Christenhusz G et al: A prospective study of the clinical utility of prenatal chromosomal microarray analysis in fetuses with ultrasound abnormalities and an exploration of a framework for reporting unclassified variants and risk factors. Genet Med 2013; 16: 469–476.
Li H, Durbin R : Fast and accurate short read alignment with Burrows–Wheeler transform. Bioinformatics 2009; 25: 1754–1760.
McKenna A, Hanna M, Banks E et al: The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res 2010; 20: 1297–1303.
Pickrell JK, Gaffney DJ, Gilad Y, Pritchard JK : False positive peaks in ChIP-seq and other sequencing-based functional assays caused by unannotated high copy number regions. Bioinformatics 2011; 27: 2144–2146.
Bernstein BE, Birney E, Dunham I, Green ED, Gunter C, Snyder M : An integrated encyclopedia of DNA elements in the human genome. Nature 2012; 489: 57–74.
Chen EZ, Chiu RW, Sun H et al: Noninvasive prenatal diagnosis of fetal trisomy 18 and trisomy 13 by maternal plasma DNA sequencing. PLoS One 2011; 6: e21791.
Rava RP, Srinivasan A, Sehnert AJ, Bianchi DW : Circulating fetal cell-free DNA fractions differ in autosomal aneuploidies and monosomy X. Clin Chem 2014; 60: 243–250.
Srinivasan A, Bianchi DW, Huang H, Sehnert AJ, Rava RP : Noninvasive detection of fetal subchromosome abnormalities via deep sequencing of maternal plasma. Am J Hum Genet. 2013; 92: 167–176.
Yu SC, Jiang P, Choy KW et al: Noninvasive prenatal molecular karyotyping from maternal plasma. PLoS One 2013; 8: e60968.
Chen S, Lau TK, Zhang C et al: A method for noninvasive detection of fetal large deletions/duplications by low coverage massively parallel sequencing. Prenat Diagn 2013; 33: 584–590.
Peters D, Chu T, Yatsenko SA et al: Noninvasive prenatal diagnosis of a fetal microdeletion syndrome. N Engl J Med 2011; 365: 1847–1848.
Jensen TJ, Dzakula Z, Deciu C, van den Boom D, Ehrich M : Detection of microdeletion 22q11.2 in a fetus by next-generation sequencing of maternal plasma. Clin Chem 2012; 58: 1148–1151.
Lau TK, Cheung SW, Lo PS et al: Non-invasive prenatal testing for fetal chromosomal abnormalities by low-coverage whole-genome sequencing of maternal plasma DNA: review of 1982 consecutive cases in a single center. Ultrasound Obstet Gynecol 2014; 43: 254–264.
Chu T, Yeniterzi S, Rajkovic A et al: High resolution non-invasive detection of a fetal microdeletion using the GCREM algorithm. Prenat Diagn 2014; 34: 469–477.
Verweij EJ, de Boer MA, Oepkes D : Non-invasive prenatal testing for trisomy 13; more harm than good? Ultrasound Obstet Gynecol. 2014; 44: 112–114.
Acknowledgements
This work has been made possible by University of Leuven (KU Leuven) SymBioSys (PFV/10/016 and GOA/12/015 to JRV and KD) and by funding from the Belgian Science Policy Office Interuniversity Attraction Poles (BELSPO-IAP) programme through the project IAP P7/43-BeMGI. HVE and KD are ‘Clinical Investigators’ of the Fonds voor Wetenschappelijk Onderzoek Vlaanderen (FWO). We thank Elly Pijkels and Kristien Philippe for their contribution to the validation study. We are grateful to all women who consented to participate in our validation study.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no conflict of interest.
Additional information
Supplementary Information accompanies this paper on European Journal of Human Genetics website
Supplementary information
Rights and permissions
About this article
Cite this article
Bayindir, B., Dehaspe, L., Brison, N. et al. Noninvasive prenatal testing using a novel analysis pipeline to screen for all autosomal fetal aneuploidies improves pregnancy management. Eur J Hum Genet 23, 1286–1293 (2015). https://doi.org/10.1038/ejhg.2014.282
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/ejhg.2014.282
This article is cited by
-
Population screening for 15q11-q13 duplications: corroboration of the difference in impact between maternally and paternally inherited alleles
European Journal of Human Genetics (2024)
-
Prenatal diagnosis of a 46,XY karyotype female fetus with an SRY-associated gonadal dysgenesis, conceived through an intracytoplasmic sperm injection: a case report
BMC Pregnancy and Childbirth (2022)
-
Rare autosomal trisomies detected by non-invasive prenatal testing: an overview of current knowledge
European Journal of Human Genetics (2022)
-
Machine learning-based detection of immune-mediated diseases from genome-wide cell-free DNA sequencing datasets
npj Genomic Medicine (2022)
-
Estimation of cell-free fetal DNA fraction from maternal plasma based on linkage disequilibrium information
npj Genomic Medicine (2021)
