Abstract
Circulating fetal cell-free DNA (cfDNA) is generally shorter than maternal cfDNA. Size selection of shorter cfDNA in total cfDNA could significantly increase the fetal fraction, but there are few reports of using this method to decrease the false negative rate for NIPT. In this study, nine false negative cases were retrospectively analyzed by NIPT retesting and E-gel based size-selection NIPT and the fetal cfDNA fraction in maternal total cfDNA was evaluated by calculating the proportion of reads from chromosome Y. Fetal placenta karyotypes were confirmed by CNVplex assays to analysis the reasons for false negative cases. Of the 81,601 pregnancies who underwent NIPT, nine false negative cases (0.01%) were found. Of eight retested cases, two (25%) had positive NIPT retest results, and five (62.5%) had positive size-selection NIPT results. For fetal cfDNA fraction, 100% cases had improvement after size-selection NIPT compared with the initial NIPT and retest results, and the fetal cfDNA fraction growth ratio ranged from 99 to 359%. For one twin pregnancy with one T18 fetus, size selection improved the fetal cfDNA fraction to 23.10%, and successfully detected the T18 fetus in NIPT. Placental tissue analysis results for two cases indicated both had confined placental mosaicism (CPM), which was confirmed with size-selection NIPT. In conclusion, size selection can significantly enrich the fetal cfDNA fraction and decrease the false negative rate of NIPT, especially for CPM and twin pregnancies.
Highlights
Non-invasive prenatal testing (NIPT) is widely used for detecting fetal chromosome trisomies 13, 18, and 21 (T13, T18, and T21) and sex chromosome aneuploidies with high sensitivity and specificity (Porreco et al, 2014; Yu et al, 2017; Zhang et al, 2017)
Abnormal ultrasound results were found in 8 false negative cases during the second trimester
A total of 81,601 pregnancies were analyzed using next-generation sequencing-based NIPT, and only 0.01% had false negative results, which is in accordance with a previous study (Suzumori et al, 2019)
Summary
Non-invasive prenatal testing (NIPT) is widely used for detecting fetal chromosome trisomies 13, 18, and 21 (T13, T18, and T21) and sex chromosome aneuploidies with high sensitivity and specificity (Porreco et al, 2014; Yu et al, 2017; Zhang et al, 2017) This method does have limitations such as maternal malignancies and confined placental mosaicism (CPM) that lead to discordance between the fetal karyotype and NIPT results (Wang et al, 2014; Bianchi et al, 2015), Size-Selection NIPT With High Success Rate not all possible genetic disorders are detectable, some false positive or negative cases, as well as 3–5% non-reportable results due to low fetal fraction circulating free DNA (cfDNA) (Liehr et al, 2017; Suzumori et al, 2019). Several studies reported using size selection to improve NIPT specificity (Minarik et al, 2015; He et al, 2018), but the application of this method to reanalyze NIPT false negative cases has not been well described
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