Long read sequencing enhances pathogenic and novel variation discovery in patients with rare diseases

Abstract With ongoing improvements in accuracy and capacity to detect complex genomic and epigenomic variations, long-read sequencing (LRS) technologies could serve as a unified platform for clinical genetic testing, particularly in rare disease settings, where nearly half of patients remain undiagnosed using existing technologies. Here, we report a simplified funnel-down filtration strategy aimed at identifying large deleterious variants and abnormal episignature disease profiles from whole-genome LRS data. This approach substantially reduced structural and copy number variants by 98.5–99.9%, respectively, while detecting all pathogenic changes in a positive control set (N = 10). When applied to patients who previously had negative short-read testing (N = 39), additional diagnoses were uncovered in 13% of cases, including a novel methylation profile specific to spinal muscular atrophy, thus opening new avenues for diagnosing and treating this life-threatening condition. Our study illustrates the utility of LRS in clinical genetic testing and in the discovery of novel disease variations.