The Atlantic salmon, Salmo salar, shows geographically structured differentiation at various classes of molecular genetic variation, among and within river stocks. Nuclear microsatellite locus variation at multiple loci has been exploited for more than a decade as a marker for the continental origin of fish caught at sea in distant-water fisheries. However, a simpler, more cost-effective, but still accurate, assignment can be obtained using a single microsatellite locus in combination with a mitochondrial DNA (mtDNA) single-nucleotide polymorphism (SNP) detected by restriction enzyme digestion. Following on from this, a preliminary study was made of the potential for using mtDNA SNP variation to enhance the resolving power and cost-effectiveness of within-continent assignment of European salmon as determined using microsatellites. Variation in 20 mtDNA regions, encompassing ∼43% of this genome, in 330 salmon from 29 rivers across Europe, was analyzed. High levels of inter-individual and inter-river variation were found, as well as evidence of regional differentiation paralleling observed microsatellite differentiation. The observations indicate scope for using mtDNA SNPs along with microsatellites for genetically based assignment of European salmon to region and river of natal origin, but further study is needed.