Simultaneous detection of fetal aneuploidy, de novo FGFR3 mutations and paternally derived β-thalassemia by a novel method of noninvasive prenatal testing.

Objective: The aim is to develop a novel noninvasive prenatal testing (NIPT) method that simultaneously performs fetal aneuploidy screening and the detection of de novo and paternally derived mutations.

Methods: A total of 68 pregnancies, including 26 normal pregnancies, 7 cases with fetal aneuploidies, 7 cases with fetal achondroplasia or thanatophoric dysplasia, 18 cases with fetal skeletal abnormalities, and 10 cases with β-thalassemia high risk were recruited. Plasma cell-free DNA was amplified by Targeted And Genome-wide simultaneous sequencing (TAGs-seq) to generate around 99% of total reads covering the whole-genome region and around 1%  covering the target genes. The reads on the whole-genome region were analyzed for fetal aneuploidy using a binary hypothesis T-score and the reads on target genes were analyzed for point mutations by calculating the minor allelic frequency of loci on FGFR3 and HBB. TAGs-seq results were compared with conventional NIPT and diagnostic results.

Results: In each sample, TAGs-seq generated 44.7-54 million sequencing reads covering the whole-genome region of 0.1-3× and the target genes of >1000×depth. All cases of fetal aneuploidy and de novo mutations of achondroplasia/thanatophoric dysplasia were identified with high sensitivities and specificities except for one false-negative paternal mutation of β-thalassemia.

Conclusions: TAGs-seq is a novel NIPT method that combines the fetal aneuploidy screening and the detection of de novo FGFR3 mutations and paternal HBB mutations.