Whole Genome Sequencing in the Evaluation of Fetal Structural Anomalies: A Parallel Test with Chromosomal Microarray Plus Whole Exome Sequencing

Abstract

Whole genome sequencing (WGS) is a powerful tool for postnatal genetic diagnosis, but relevant clinical studies in the field of prenatal diagnosis are limited. The present study aimed to prospectively evaluate the utility of WGS compared with chromosomal microarray (CMA) and whole exome sequencing (WES) in the prenatal diagnosis of fetal structural anomalies. We performed trio WGS (≈40-fold) in parallel with CMA in 111 fetuses with structural or growth anomalies, and sequentially performed WES when CMA was negative (CMA plus WES). In comparison, WGS not only detected all pathogenic genetic variants in 22 diagnosed cases identified by CMA plus WES, yielding a diagnostic rate of 19.8% (22/110), but also provided additional and clinically significant information, including a case of balanced translocations and a case of intrauterine infection, which might not be detectable by CMA or WES. WGS also required less DNA (100 ng) as input and could provide a rapid turnaround time (TAT, 18 ± 6 days) compared with that (31 ± 8 days) of the CMA plus WES. Our results showed that WGS provided more comprehensive and precise genetic information with a rapid TAT and less DNA required than CMA plus WES, which enables it as an alternative prenatal diagnosis test for fetal structural anomalies.

Keywords: chromosomal microarray; fetal structural anomalies; prenatal diagnosis; whole exome sequencing; whole genome sequencing.

Figure 1

Flowchart of the study. A total of 111 fetuses with structural or growth anomalies were subjected to two test strategies: trio genome sequencing (whole genome sequencing (WGS)) and chromosomal microarray (CMA) plus exome sequencing (whole exome sequencing (WES)). WES was sequentially performed in only 102 fetuses with negative CMA results. The positive/negative rates are provided in each box.

Figure 2

Architecture of a mixed cohort referred for prenatal diagnosis. Each slice of the pie chart represents one individual in the prospective cases analyzed by genome sequencing (WGS) and chromosomal microarray (CMA) plus exome sequencing (WES) where clinically relevant findings were identified. Types of variant are indicated by colors (aneuploidy, light blue; duplication and heterozygous (het) single nucleotide variant (SNV), blue; unbalanced translocation, red; intrauterine infection, yellow; compound heterozygous SNV, light gray; heterozygous SNV, dark gray). Additional genetic findings by WGS are indicated by * and additional nongenetic findings by WGS are indicated by #.

Figure 3

Whole genome sequencing (WGS) identified pathogenic copy number variants (CNVs) in twin fetuses arising from their father’s balanced translocation. (A) Distributions of copy number across chromosome 13 and chromosome 15 are shown at the top and bottom, respectively. Dots in red indicate a heterozygous deletion of 19.7 Mb on chromosome 13, and dots in blue indicate a duplication of 1.6 Mb on chromosome 15 in both case 102-1 and case 102-2. The deletion affected three genes that are associated with autosomal dominant developmental disorder. (B) The father’s karyotype of chromosome 13 and chromosome 15 are shown at the top and bottom. The distribution of base pair sequencing depth and Sanger sequencing results across the breakpoints on chromosome 13 and chromosome 15 are displayed, respectively, in the middle. There is a templated 13-bp duplication and a 34-bp deletion in the breakpoint junction on chromosomes 13 and 15. (C) A schematic drawing of the formation of balanced translocations between chromosomes 13 and 15 in the father.

Figure 4

RT-PCR validation of the viral DNA in the amniotic fluid. The load of cytomegalovirus (CMV) was 5.73 × 10⁵ copies/mL, which indicated intrauterine CMV infection in case 5.