Rafehi Haloom, Fearnley Liam G, Read Justin, Snell Penny, Davies Kayli C, Scott Liam, Gillies Greta, Thompson Genevieve C, Field Tess A, Eldo Aleena, Bodek Simon, Butler Ernest, Chen Luke, Drago John, Goel Himanshu, Hackett Anna, Halmagyi G Michael, Hannaford Andrew, Kotschet Katya, Kumar Kishore R, Kumble Smitha, Lee-Archer Matthew, Malhotra Abhishek, Paine Mark, Poon Michael, Pope Kate, Reardon Katrina, Ring Steven, Ronan Anne, Silsby Matthew, Smyth Renee, Stutterd Chloe, Wallis Mathew, Waterston John, Wellings Thomas, West Kirsty, Wools Christine, Wu Kathy H C, Szmulewicz David J, Delatycki Martin B, Bahlo Melanie, Lockhart Paul J
Population Health and Immunity Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia.
Department of Medical Biology, University of Melbourne, Parkville, Victoria 3052, Australia.
Genome Res. 2025 Apr 14;35(4):769-785. doi: 10.1101/gr.279634.124.
The cerebellar ataxias (CAs) 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 evaluates 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) was performed to identify pathogenic RE. SR-GS provided a genetic diagnosis for 38% of the cohort (40/110) including seven non-RE pathogenic variants. RE causes disease in 33 individuals, with the most common condition being SCA27B ( = 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 enhances diagnostic outcomes. Orthogonal testing for SCA27B revealed a 15% 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 used 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 before clinical implementation.
小脑性共济失调（CA）是一组异质性疾病，其特征为进行性共济失调。17个重复序列扩增（RE）位点已被确定为主要遗传病因，占基因诊断的80%以上。尽管如此，诊断检测有限且效率低下，通常采用单基因检测方法。本研究评估长读长和短读长测序作为CA诊断工具的有效性。我们招募了110名临床诊断为CA的个体（48名女性，62名男性）。进行短读长基因组测序（SR-GS）以鉴定与CA相关的356个基因中的致病性RE以及非RE变异。独立地，采用适应性采样的长读长测序（LR-AS）来鉴定致病性RE。SR-GS为38%的队列（40/110）提供了基因诊断，包括7个非RE致病性变异。RE导致33名个体患病，最常见的情况是SCA27B（=24）。相比之下，LR-AS在29名个体中鉴定出致病性RE。除了4例因读长深度低而未被LR-AS检测到的SCA27B病例外，两种方法对RE的鉴定结果一致。对于这两种技术，人工检查RE比对可提高诊断结果。对SCA27B的正交检测显示，SR-GS和LR-AS的假阳性率分别为15%和0%。总之，这两种技术都是CA的强大筛查工具。SR-GS是诊断机构目前使用的成熟技术，只需在生物信息学工作流程上进行微小更改即可实现CA诊断。LR-AS在RE检测和特征分析方面具有显著优势，但在临床应用前需要优化。
