Variant detection from long-read genome sequencing (lrGS) has proven to be more accurate and comprehensive than variant detection from short-read genome sequencing (srGS). However, the rate at which lrGS can increase molecular diagnostic yield for rare disease is not yet precisely characterized. We performed lrGS using Pacific Biosciences “HiFi” technology on 96 short-read-negative probands with rare diseases that were suspected to be genetic. We generated hg38-aligned variants and de novo phased genome assemblies, and subsequently annotated, filtered, and curated variants using clinical standards. New disease-relevant or potentially relevant genetic findings were identified in 16/96 (16.7%) probands, nine of which (8/96, ∼9.4%) harbored pathogenic or likely pathogenic variants. Nine probands (∼9.4%) had variants that were accurately called in both srGS and lrGS and represent changes to clinical interpretation, mostly from recently published gene-disease associations. Seven cases included variants that were only correctly interpreted in lrGS, including copy-number variants (CNVs), an inversion, a mobile element insertion, two low-complexity repeat expansions, and a 1 bp deletion. While evidence for each of these variants is, in retrospect, visible in srGS, they were either not called within srGS data, were represented by calls with incorrect sizes or structures, or failed quality control and filtration. Thus, while reanalysis of older srGS data clearly increases diagnostic yield, we find that lrGS allows for substantial additional yield (7/96, 7.3%) beyond srGS. We anticipate that as lrGS analysis improves, and as lrGS data sets grow allowing for better variant-frequency annotation, the additional lrGS-only rare disease yield will grow over time.

中文翻译：

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长读长基因组测序和变异再分析可提高神经发育障碍的诊断率


长读长基因组测序 （lrGS） 的变异检测已被证明比短读长基因组测序 （srGS） 的变异检测更准确、更全面。然而，lrGS 提高罕见病分子诊断率的速度尚未准确表征。我们使用 Pacific Biosciences“HiFi”技术对 96 名患有疑似遗传性疾病的短读阴性先证者进行了 lrGS。我们生成了 hg38 对齐的变异和从头分阶段基因组组装，随后使用临床标准对变异进行注释、过滤和整理。在 16/96 （16.7%） 的先证者中发现了新的疾病相关或潜在相关的遗传发现，其中 9 例 （8/96， ∼9.4%） 携带致病性或可能的致病性变异。9 个先证者 （∼9.4%） 具有在 srGS 和 lrGS 中被准确调用的变异，代表了临床解释的变化，主要来自最近发表的基因-疾病关联。7 例包括仅在 lrGS 中正确解释的变异，包括拷贝数变异 （CNV） 、倒置、移动元件插入、2 次低复杂度重复扩增和 1 bp 缺失。虽然回想起来，这些变体中的每一个的证据在 srGS 中都是可见的，但它们要么没有在 srGS 数据中被调用，要么由大小或结构不正确的调用表示，要么质量控制和过滤失败。因此，虽然对较旧的 srGS 数据进行重新分析明显提高了诊断率，但我们发现 lrGS 允许在 srGS 之外获得大量额外的产量 （7/96， 7.3%）。我们预计，随着 lrGS 分析的改进，以及随着 lrGS 数据集的增长，允许更好的变异频率注释，额外的仅 lrGS 罕见病产量将随着时间的推移而增长。