Out of 90 Portuguese patients with mitochondrial cytopathy, six harbored the A3243G mutation in the mtDNA tRNA(Leu(UUR)) gene ('MELAS mutation'). They had heterogeneous clinical features, including myopathy with stroke-like episodes, progressive external ophthalmoparesis, diabetes mellitus, and subacute encephalopathy. Histochemical and biochemical analyses of muscle biopsies showed abundant ragged-red fibers reacting positively with the cytochrome oxidase stain, and decreased respiratory chain enzyme activities. On average, the proportion of mutated mtDNA was 67% (20-88%) in tissues from patients and 21% (0-49%) in blood from 20 maternal relatives. The proportion of mutated mitochondrial genomes in muscle did not correlate with clinical presentation or duration of disease. This study, the first in Portuguese patients, confirms the frequent occurrence of the A3243G mutation in patients with mitochondrial diseases, and emphasises the usefulness of genetic testing in reaching a correct diagnosis.