The genetic diversity of Stenopsyche marmorata was investigated in three spatial hierarchies, among catchments, among reaches within catchments, and among individuals within reaches, by AFLP analysis of 306 individuals in 12 reaches in 3 catchments in Eastern Japan. The analysis of molecular variance revealed that the diversity within reaches accounted for the highest percentage (86%) of the total genetic variation in the three hierarchies. The genetic diversity between reaches increased with geographical distance between reaches on a small spatial scale (=2-74km); however, it did not increase on a large spatial scale (=74-450km). The level of gene flow among adjacent catchments (=55-74km) was lower than that between reaches within catchments (=2-19km). The genetic diversity within reaches (as the percentage of polymorphic loci and expected heterozygosity) positively correlated with altitude, suggesting that altitude is an environmental factor in determining the local population size of S. marmorata in each reach.