Strengths and caveats of parallel sequencing approaches in nemaline myopathy

Abstract

Background: Nemaline myopathy (NM) is a form of congenital myopathy defined histopathologically by the feature of nemaline bodies on skeletal muscle biopsy. The traditional approach to diagnosis has been by skeletal muscle biopsy and candidate gene sequencing. Aim: To determine the diagnostic utility of different massively parallel sequencing (MPS) approaches in NM. Methods: A cohort of 21 individuals with NM were investigated using a range a different MPS techniques (neuromuscular gene panel, whole exome sequencing (WES), whole genome sequencing (WGS), RNA-Sequencing (RNA-Seq). Results: A genetic diagnosis was obtained in 20/21 NM families investigated using MPS approaches (95% diagnosis). Pathogenic splicing variants were identified for 6/20 families; extended splice site or intronic variants relied heavily upon mRNA studies frommuscle biopsies to support pathogenicity. This study redefines the relative incidence ofNMgenotypes in our Australian cohort of probands from 52 families with a histopathological diagnosis of NM and a molecular genetic diagnosis. Discussion/Conclusion: this study confirms the utility of massively parallel sequencing as a first-tier investigation for patients with congenital myopathy, while emphasising the continuing importance of muscle biopsy in a subset of patients. We highlight challenges interpreting variants simultaneously identified in multiple plausible candidate genes, provide specific recommendations regarding analysis and interpretation of splicing variants, and propose a diagnostic algorithm for genetic diagnosis of congenital myopathy.L6298899942019-11-22 <br />