Comparing the Influence of Habitat Configuration on Population Connectivity and Genetic Structure Using Congeneric Species Across Multiple Taxa

Abstract Habitat configuration influences population connectivity and, consequently, genetic structure. River networks provide heterogeneous, hierarchically arranged environments that shift drastically from upstream to downstream. We compared congeneric species across Ephemeroptera, Plecoptera, and Trichoptera, emphasizing longitudinal replacement (upstream to downstream) and the rarely studied wet rock (hygropetric) habitats. We surveyed six rivers on the Muroto Peninsula, Japan, qualitatively sampling aquatic insects at 63 sites. Cluster analysis based on the environmental data classified surveyed sites into four clusters. We analyzed a total of 10 species from four genera that exhibit longitudinal replacement patterns within genera. Genetic analyses based on the mitochondrial cytochrome c oxidase subunit I region revealed that upstream species showed higher genetic diversity than downstream species. In contrast, species adapted to hygropetric exhibited the lowest genetic differentiation among all habitat types. A novel contribution of this study is the inclusion of hygropetric species. The surprisingly low differentiation in hygropetric species suggests high connectivity, similar to lentic species. By comparing congeneric taxa across orders within environmentally similar rivers, we reduce phylogenetic and environmental confounds, strengthening inference that habitat configuration and dispersal traits jointly shape genetic structure. These findings provide a new perspective on riverine spatial ecology and underscore the importance of microhabitat-aware comparisons for evolutionary inference.