Analysis of mitochondrial DNA heteroplasmy in sporadic ALS suggests technical limitations rather than disease association

Mitochondrial DNA (mtDNA) has received increasing attention in amyotrophic lateral sclerosis (ALS) following the recent report of recurrent low-heteroplasmy mtDNA variants in patients. Here, we performed mtDNA analysis on an independent cohort of 20 sporadic ALS patients using an in-house next-generation sequencing pipeline designed for diagnostics. Using standard filters, none of the previously reported low-heteroplasmy mtDNA variants were detected. These variants only appeared in the low-quality data and were present at similar rates in a large reference population without ALS, localizing to homopolymeric regions that are prone to sequencing errors. Our findings suggest that these low-level mtDNA variants are a result of the technical limitations of short-read next-generation sequencing rather than being associated with the disease. • mtDNA variants previously reported were not detected using standard filters. • Low-level variants were only observed when removing quality filters. • Similar frequencies were found in ALS and non-ALS subjects. • Variants were mapped to homopolymeric regions prone to sequencing errors. • These findings suggest sequencing artifacts over disease association.