The cerebellar ataxias (CA) are a heterogeneous group of disorders characterized by progressive incoordination. Seventeen repeat expansion (RE) loci have been identified as the primary genetic cause and account for >80% of genetic diagnoses. Despite this, diagnostic testing is limited and inefficient, often utilizing single gene assays. This study evaluated the effectiveness of long- and short-read sequencing as diagnostic tools for CA. We recruited 110 individuals (48 females, 62 males) with a clinical diagnosis of CA. Short-read genome sequencing (SR-GS) was performed to identify pathogenic RE and also non-RE variants in 356 genes associated with CA. Independently, long-read sequencing with adaptive sampling (LR-AS) and performed to identify pathogenic RE. SR-GS identified pathogenic variants in 38% of the cohort (40/110). RE caused disease in 33 individuals, with the most common condition being SCA27B (n=24). In comparison, LR-AS identified pathogenic RE in 29 individuals. RE identification for the two methods was concordant apart from four SCA27B cases not detected by LR-AS due to low read depth. For both technologies manual review of the RE alignment enhanced diagnostic outcomes. Orthogonal testing for SCA27B revealed a 16% and 0% false positive rate for SR-GS and LR-AS respectively. In conclusion, both technologies are powerful screening tools for CA. SR-GS is a mature technology currently utilized by diagnostic providers, requiring only minor changes in bioinformatic workflows to enable CA diagnostics. LR-AS offers considerable advantages in the context of RE detection and characterization but requires optimization prior to clinical implementation.

The authors have declared no competing interest.

This work was supported by the Australian Government National Health and Medical Research Council grants (GNT2001513 and MRFF2007677) to PJL, MBD, HR, MB and DS. HR was supported by a NHMRC Emerging Leadership 1 grant (1194364) and MB was supported by an NHMRC Investigator grant (1195236). Additional funding was provided by the Independent Research Institute Infrastructure Support Scheme, the Victorian State Government Operational Infrastructure Program and the Murdoch Childrens Research Institute. KRK reports financial support was provided by the Ainsworth 4 Foundation unrelated to the current study.

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