Evaluation of using WGS/WES to characterize ACMG actionable genes in genetic testing reports

Abstract

Since the American College of Medical Genetics and Genomics (ACMG) published recommendations for reporting incidental findings in clinical exome and genome sequencing in 2013, the list of suggested actionable genes appear more and more frequently in the reports of genetic testing. However, the existence of low-coverage regions, no matter happening systematically or randomly, in whole genome sequencing (WGS) or whole exome sequencing (WES) raise the chances of false negatives when reporting pathogenicity in variant discovery. To evaluate the effectivity of different sequencing methods, this study developed a workflow to examine low-coverage regions in ACMG actionable genes (version 2 contains 59 genes) on three groups of sequencing data, including 5 samples of 30X WGS, 41 samples of 40X WGS, and 11 samples of WES. By extracting the ACMG regions of the alignment files, we first found that per-base mean coverage in WES is significantly higher in some chromosomes (2, 5, 6, 9, 13, 14, 17, 18) than the others, and the overall average (44.5) is higher than 30X WGS (32.4) and 40X WGS (36.2). However, the total length of the low-coverage regions in ACMG actionable genes is higher in WES samples: 23 times higher than 40X WGS, and 8 times higher than 30X WGS. When looking into ClinVar significance of pathogenic/likely pathogenic (P/LP) variants (the 2020/03/16 release contains 21,388 P/LP variants falling in the 59 actionable genes), 100% (11/11) WES samples and 80% (4/5) 30X WGS samples have more than 20 P/LP variants in the low-coverage regions, while only 2.4% (1/41) 40X WGS samples have such a risk of missing >20 P/LP variants. We conclude that in order to reduce the possibility of missing clinically significant variants, 40X WGS are more recommended than 30X WGS and WES to achieve satisfied quality of characterizing ACMG actionable genes in genetic testing.