Long read sequencing characterises a novel structural variant opaque to short reads, with likely functional impact in a case of unexplained severe fatigue

Abstract Despite the recent advances in genomic analysis, causative variants cannot be found for a sizeable proportion of patients with suspected genetic disorders. Many of these disorders involve genes in difficult-to-align genomic regions which are recalcitrant to short read approaches. Structural variants in these regions can be particularly hard to detect or define with short reads, yet may account for a significant number of cases. Long read sequencing can overcome these difficulties and is providing new hope for diagnosis and patient care. Here, we present a case of unusually complex, severe fatigue where a potentially relevant structural variant was indicated but could not be resolved by short-read sequencing. We use nanopore sequencing to identify and fully characterise a large inversion in a highly homologous region spanning the AKR1C gene locus, along with serum steroid analysis to investigate the functional consequences. The DNA inversion appears to increase the expression of AKR1C2 while limiting AKR1C1 activity, resulting in a relative increase of inhibitory neurosteroids and impaired progesterone metabolism. This study provides an example of where long read sequencing may supplement the use of more traditional sequencing methods in clinical care to increase diagnostic yield for rare disease, and highlights some of the challenges that arise in sequencing complex regions containing tandem arrays of genes. It also proposes a novel gene associated with a specific disease aetiology that may be an underlying cause of unexplained severe fatigue.