Genetic variability was examined at 16 putative allozymic loci in the narrow endemic fern Polystichum otomasui. Although this species is distributed in only a few valleys within an approximately 10 km 6 km area on the Kyusyu Island of Japan, there are a relatively large number of individuals in this area. The percentage of polymorphic loci (P), the number of alleles per locus (A), the observed heterozygosity (H O ) and the expected heterozygosity (H E ) were 61.9, 1.93, 0.166 and 0.177, respectively, at the population level, and P, A and H E were 81.3, 2.69 and 0.199, respectively, at the species level, indicating that P. otomasui has extremely high allozymic variation for a fern species. The G ST value among the 10 populations examined within the range was 0.108, suggesting that the interpopulation gene flow is high enough to impede genetic structuring. The mating system estimates, Wright's fixation index and the intragametophytic selfing rate, indicate that the species is outcrossing. High genetic variability, in spite of narrowness of the distribution, may have resulted from a recent rapid decline in the population and/or the large effective population size resulting from the extensive gene flow among the populations.