Mitochondrial DNA (mtDNA) variation was examined in 209 Atlantic salmon from two river systems, the R. Itchen in Hampshire (Southern England) and the R. Conwy (North Wales). Within each system, five spawning sites were sampled. Four enzymes (AVA II, HAE III, HINF I and MBO I) revealed restriction fragment polymorphisms that were informative. Ten clonal lines were observed. These clones were differentially distributed between the two river systems and single clonal types were found to predominate at several spawning sites. MBO I variants were found in salmon from the R. Itchen but not the R. Conwy. A significant heterogeneity in frequency distribution of clonal lines between parr and smolt assemblages of the same year class within the R. Itchen was detected. This heterogeneity suggests that differences in survival or migratory behaviour may be identified by changes in mtDNA clonal frequencies.