Adaptive radiation of the Callicarpa genus in the Bonin Islands revealed through double-digest restriction site–associated DNA sequencing analysis

The Bonin Islands, comprising of the Mukojima, Chichijima, and Hahajima Islands, are known for their isolated and distinctive habitats, hosting a diverse array of endemic flora and fauna. In these islands, adaptive radiation has played a remarkable role in speciation, particularly evident in the Callicarpa genus that is represented by three species: Callicarpa parvifolia and Callicarpa glabra exclusive to the Chichijima Islands, and Callicarpa subpubescens , distributed across the entire Bonin Islands. Notably, C. subpubescens exhibits multiple ecotypes, differing in leaf hair density, flowering time, and tree size. In this study, we used double-digest restriction site–associated DNA sequencing to analyze species, ecotypes and geographical variations within Callicarpa in the Bonin Islands. We aimed to determine detailed phylogenetic relationships, investigate species and ecotype diversification patterns, estimate divergence times, and explore cryptic species using genetic and phenotypic data. Genetic analysis revealed that C. parvifolia and C. glabra formed a single, distinct genetic groups. Conversely, C. subpubescens showed seven genetic groups corresponding to different ecotypes and regions, with one ecotype derived from the hybridization of two others. Phylogenetic and population demography analyses, focusing on six Chichijima and Hahajima Islands–based species/ecotypes, indicated the divergence of an ecotype adapted to tall mesic forests approximately 170 kya, whereas the other five species/ecotypes diverged nearly simultaneously around 73–77 kya. Environmental changes during the glacial period likely contributed to this process of adaptive radiation. Moreover, leaf morphology, flowering time, and genetic analyses suggested the presence of two cryptic species within C. subpubescens .