The taxonomic diversity of actinomycetes from soils in a cool-temperate area (74 samples from Rishiri Island) and a subtropical area (94 samples from Iriomote Island) of Japan was compared. Soil samples were treated with five different pre-treatment methods and subsequently cultured on humic acid-vitamin agar, resulting in the selective isolation of 1,234 actinomycete strains. These isolates were classified using 16S rDNA sequence analysis. 16S rDNA sequence similarity values of 97% and 98:7% were tentatively used to identify isolates at the genus and species ranks, respectively. The actinomycete assemblages isolated comprised diverse species. Specifically, 668 of the Rishiri isolates fell into 17 families, 40 genera, and 178 species, and 566 of the Iriomote isolates belonged to 21 families, 59 genera, and 194 species. Diversity analyses based on the Shannon index revealed that there was no significant difference between the species diversity of the Rishiri isolates and that of the Iriomote isolates. However, there was little taxonomic overlap between the Rishiri isolates and the Iriomote isolates, as only 66 of the species occurred in both groups. Several species that were reportedly isolated from other tropical areas were found among the Iriomote isolates but not among the Rishiri isolates. This result suggests that climate may influence the species types of soil-inhabiting actinomycetes.