Mitochondrial diseases are genetic disorders with multi-system involvement. They can be caused by alterations in mitochondrial genome or nuclear genome. These alterations disrupt the normal functions of mitochondria and subsequently lead to dysfunctions of energy-demanding organs such as skeletal muscles, heart and brain. Before the era of next-generation sequencing (NGS), molecular diagnosis of mitochondrial diseases caused by mitochondrial DNA mutations was usually made by targeted analysis of mutation hotspots using Sanger sequencing. However, there were two major limitations: (1) structural rearrangements or other uncommon mutations could not be identified; (2) mutations at low heteroplasmy levels (i.e. 2% heteroplasmic level can be confidently called. From 2019 to 2022, we have received samples from 71 families for mitochondrial DNA testing. We identified pathogenic variants in 9 (12.7%) families, and variants of uncertain significance (VUS) in 7 (9.8%) other families, for which further investigations would be required. Among the positive results with pathogenic variants detected, 5 out of 9 were not reported in an early version of morbid map of the human mitochondrial genome (i.e. not mutation hotspots). To conclude, the testing modality can expand the genetic analysis of mitochondrial diseases from several mutation hotspots to the whole mitochondrial genome. The application of NGS also enables a promising analytical sensitivity in the detection of low-level heteroplasmic mitochondrial variants.
