Levels and distribution of genetic variation were investigated in the homosporous fern, Polystichum munitum. Homosporous ferns differ from higher vascular plants in that they possess potentially bisexual gametophytes which can produce a completely homozygous sporophyte in a single generation. Because of this, it has long been maintained that ferns possess an inbreeding mating system, resulting in low levels of genetic variation and high levels of homozygosity within populations. The four populations sampled maintain high levels of genetic variation (P̄ = 0.542; H̄ = 0.111; Ā = 2.23), comparable to that maintained by populations of outcrossing seed plants. The mean fixation index, F, for the four populations was 0.052, indicating no significant deviations from Hardy-Weinberg genotypic expectations. Polystichum munitum distributes most of its genetic variation within rather than among populations. Population-genetic structure was assessed by subdividing each of two large populations into 10 x 10-m subpopulations. Comparisons of genetic variation within and among subpopulations indicated little genetic substructure within either of the artificially subdivided populations. Estimates of interpopulational gene flow (Nm) are extremely high, comparable to those reported for gymnosperms. Statistical estimates of intragametophytic selfing are very low, ranging from 0 to 3%. This study suggests that Polystichum munitum is an outcrossing species. Evidence from this and other investigations indicates that fern species do not typically self-fertilize and that mating systems in ferns vary as they do among species of seed plants.