Targeted fetal cell‐free DNA screening for aneuploidies in 4,594 pregnancies: Single center study

Abstract

Background Next‐generation sequencing (NGS) and discovery of fetal cell‐free DNA (cfDNA) in the maternal circulation render possible prenatal screening for trisomy 21 (Down syndrome), trisomy 18, trisomy 13, and sex chromosome aneuploidies. The approach is called “fetal cfDNA screening” and in contrast to noninvasive conventional serum screening, it provides the identification of 98%–99% of fetuses with Down syndrome. Methods Retrospective analysis of targeted noninvasive prenatal testing (NIPT) (Clarigo Test) pregnancies with moderate risk, which we have reported between 2016 and 2018 years is presented. Two separate laboratory workflows and NGS platforms are used for the same targeted NIPT analysis. Results In total, 4,594 pregnant women were investigated. Initial 3,594 cases are studied by MiSeq platform, the last 1,000 cases by NextSeq. Failure rate for MiSeq platform is 10.9% and for NextSeq is 8.7%. Automatically reported cases constitute 75% of the MiSeq group and 87% of the NextSeq group. Conclusions Targeted NIPT results suggest that MiSeq platform could be used for NIPT which would be an essential option particularly for laboratories with low sample flow. And, the NextSeq platform has easier wet lab process and also increased success rate in automatic reporting which is suitable for centers with high number of NIPT cases.

Systematic evaluation of NIPT aneuploidy detection software tools with clinically validated NIPT samples

“…Furthermore, our data demonstrated that a well-chosen computational NIPT software employed in combination with appropriate Z-score thresholds can be used at lower sequencing coverages, even at sequencing depths below 5M RPS. This is in line with the current knowledge, as NIPT usage potential at lower coverages (such 2.2M RPS) has been demonstrated previously [ 14 ]. However, at lower sequencing depths (below 5M RPS), the compared computational tools yielded trisomy risks with differing accuracies, reflected by mostly with a systematically increasing numbers of false-positive and false-negative trisomy cases ( Figs 1 and S1 and S2 ).…”

Systematic evaluation of NIPT aneuploidy detection software tools with clinically validated NIPT samples

“…We also observed that a well-chosen computational NIPT software in combination with appropriately applied Z-score thresholds can be used at lower sequencing coverages, even below 5M RPS. This is in line with the current knowledge, also NIPT potential at lower coverages (such 2.2M RPS) has been shown (18). However, at lower sequencing coverages (below 5M RPS), various computational tools infer trisomy risks with a quite different accuracy, mostly with a systematically increasing number of false-positive and false-negative trisomy cases ( Figure 1, Supplementary Figure 1, Supplementary Figure 2 ).…”

“…However, Koc et al . also observed that NIPT test failures (as no result) are often related to the low FF estimation in the case of lower sequencing coverage (18). Also, depending on the software used for FF calculation, FF estimator (17) accuracy can significantly decrease at low sequencing coverages in the case of truly low FF ( Figure 3, Supplementary Figure 6 ).…”

Progress and Challenges in Laboratory-Based Diagnostic and Screening Approaches for Aneuploidy Detection during Pregnancy

“…69 The MiSeq and NextSeq platforms from Illumina have also been successfully used with cffDNA for aneuploidy detection, with a 10.9% failure rate in 3594 MiSeq cases and 8.7% failure rate in 1000 NextSeq cases. 70 Targeted NGS has also been utilized to analyze selected genomic regions of interest for NIPT. 71 In a report by Koumbaris et al, a blind study showed 100% diagnostic sensitivity while being more cost-effective as an NGS-based method for NIPT.…”