The excess female transmission of multiple sclerosis (MS) and the observation of an MS-like illness in patients with Leber's hereditary optic neuropathy who carry a mitochondrial DNA mutation may indicate that mitochondrial genes contribute to the genetic susceptibility to MS. We sequenced the protein- and RNA-coding sequences of 9 patients with MS who had a family history of MS consistent with maternal transmission. Four base-pair (bp) changes of particular interest were identified. Those at bp 4216 and 4917 may play a role in the etiology of Leber's hereditary optic neuropathy. Two others, at bp 11447 and 14766, were found in all MS patients sequenced. Restriction enzyme analysis used to screen 175 unrelated MS patients and 233 healthy control subjects showed that each of these changes was present in MS patients at a similar frequency to control subjects. The 4216 and 4917 changes were at a higher frequency in north European control subjects than previously documented. We conclude that variation in mitochondrial DNA is unlikely to contribute to susceptibility to MS. The etiology of the overlap between Leber's hereditary optic neuropathy and MS remains unexplained.