Whole mitochondrial DNA (mtDNA) sequencing is now systematically used in clinical laboratories to screen patients with a phenotype suggestive of mitochondrial disease. Next Generation Sequencing (NGS) has significantly increased the number of identified pathogenic mtDNA variants. Simultaneously, the number of variants of unknown significance (VUS) has increased even more, thus challenging their interpretation. Correct classification of the variants' pathogenicity is essential for optimal patient management, including treatment and genetic counseling. Here, we used single muscle fiber studies to characterize eight heteroplasmic mtDNA variants, among which were three novel variants. By applying the pathogenicity scoring system, we classified four variants as “definitely pathogenic” (m.590A>G, m.9166T>C, m.12293G>A, and m.15958A>T). Two variants remain “possibly pathogenic” (m.4327T>C and m.5672T>C) but should these be reported in a different family, they would be reclassified as “definitely pathogenic.” We also illustrate the contribution of single‐fiber studies to the diagnostic approach in patients harboring pathogenic variants with low level heteroplasmy.

中文翻译：

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用于分配与线粒体疾病相关的八种线粒体 DNA 变异的致病性的单纤维研究。

全线粒体 DNA (mtDNA) 测序现在被系统地用于临床实验室，以筛选具有提示线粒体疾病表型的患者。下一代测序 (NGS) 显着增加了已鉴定的致病性 mtDNA 变异的数量。同时，未知意义的变异（VUS）的数量增加得更多，从而对他们的解释提出了挑战。正确分类变异的致病性对于优化患者管理（包括治疗和遗传咨询）至关重要。在这里，我们使用单肌纤维研究来表征八种异质性 mtDNA 变体，其中包括三个新变体。通过应用致病性评分系统，我们将四种变异分类为“绝对致病”（m.590A>G、m.9166T>C、m.12293G>A 和 m.15958A>T）。两种变异仍然“可能致病”（m.4327T>C 和 m.5672T>C），但如果它们在不同的家族中报告，它们将被重新分类为“绝对致病”。我们还说明了单纤维研究对携带低水平异质性致病变异的患者的诊断方法的贡献。