Abstract: The mating system of plants is a pivotal determinant in shaping genetic diversity and fine-scale spatial genetic structure (FSGS), with self-fertilizing species generally demonstrating reduced genetic diversity and more pronounced FSGS compared to outcrossing species. Nevertheless, the extent to which this pattern is modulated by the interplay of population history, microhabitat variability, and seed dispersal mechanisms across different habitats remains insufficiently explored. This study undertakes a comparative analysis of two Roscoea species (Zingiberaceae) with distinct mating systems to investigate the influence of demographic history, microhabitat conditions, and seed dispersal on FSGS. Utilizing thousands of single nucleotide polymorphisms (SNPs) identified in the outcrossing Roscoea cautleoides (oRC) and the selfing Roscoea schneideriana (sRS), we assessed their genetic diversity, FSGS, and historical population dynamics across various habitats within the same geographic region. Consistent with prevailing theories, our findings indicate that oRC exhibits greater genetic diversity and less pronounced FSGS at finer scales compared to sRS. Outlier SNPs revealed a markedly higher genetic diversity within the populations of both species, indicating potential adaptive divergence in response to distinct microhabitats. Both oRC and sRS populations exhibited patterns of expansion and recent bottlenecks which aligns with climatic fluctuations during the Pleistocene and subsequent climate warming, respectively. Field observations on dispersal mechanisms revealed that seeds of sRS were transported by ants significantly farther from the parent plants compared to seeds of oRC. Our results imply the dominant role of mating systems along with the potential impacts of seed dispersal, environmental heterogeneity and historical population dynamics.