The additional diagnostic yield of long-read sequencing in undiagnosed rare diseases

Abstract

Long-read sequencing (LRS) is a promising technology positioned to study the significant proportion of rare diseases (RDs) that remain undiagnosed as it addresses many of the limitations of short-read sequencing, detecting and clarifying additional disease-associated variants that may be missed by the current standard diagnostic workflow for RDs. Some key areas where additional diagnostic yields may be realized include: (1) detection and resolution of structural variants (SVs); (2) detection and characterization of tandem repeat expansions; (3) coverage of regions of high sequence similarity; (4) variant phasing; (5) the use of de novo genome assemblies for reference-based or graph genome variant detection; and (6) epigenetic and transcriptomic evaluations. Examples from over 50 studies support that the main areas of added diagnostic yield currently lie in SV detection and characterization, repeat expansion assessment, and phasing (with or without DNA methylation information). Several emerging studies applying LRS in cohorts of undiagnosed RDs also demonstrate that LRS can boost diagnostic yields following negative standard-of-care clinical testing and provide an added yield of 7%–17% following negative short-read genome sequencing. With this evidence of improved diagnostic yield, we discuss the incorporation of LRS into the diagnostic care pathway for undiagnosed RDs, including current challenges and considerations, with the ultimate goal of ending the diagnostic odyssey for countless individuals with RDs.