Development and validation of a fetal genotyping assay with potential for noninvasive prenatal diagnosis of hereditary hearing loss

Objective Inherited non-syndromic hearing loss (NSHL) is a common sensory disorder that afflicts otherwise healthy individuals. The aim of the study was to evaluate the performance of circulating single molecule amplification and re-sequencing technology (cSMART) for non-invasive prenatal testing (NIPT) of NSHL. Method Neonatal inheritance of NSHL mutations was determined from bloodspots using SNaPshot genotyping. NIPT of cell-free DNA for fetal NSHL mutations in the GJB2, GJB3 and SLC26A4 genes was performed by a multiplex cSMART assay. The percentage of mutant alleles was used to deduce fetal DNA fractions and assign fetal genotypes. Results A total of 25 plasma samples selected with different fetal NSHL genotypes were coded and retrospectively analyzed by NIPT. Three normal fetuses, 18 carrier fetuses comprising seven GJB2 109G>A, four GBJ2 235delC, three GJB2 299-300delAT and four SLC26A4 IVS7-2A>G heterozygotes and four affected fetuses comprising two GJB2 109G>A homozygotes, one GBJ2 235delC homozygote and one compound GJB2 235delC/299-300delAT heterozygote were identified. All 25 fetal genotypes determined by the cSMART assay were concordant with neonatal genotypes. Conclusion The cSMART assay applied to cell-free DNA isolated from maternal plasma of pregnant women is highly accurate for calling correct fetal NSHL genotypes. © 2016 John Wiley & Sons, Ltd.