Mating System and Fine-Scale Spatial Genetic Structure of the Tropical Epiphytic Orchid Rhynchostylis gigantea

Habitat fragmentation caused by human activities threatens plant genetic diversity, but the mechanisms shaping gene flow and spatial genetic structure in epiphytic orchids remain poorly understood. Here, we investigated the genetic structure and gene flow patterns of the epiphytic orchid Rhynchostylis gigantea in a human-modified landscape on Hainan Island, China, using SNP markers. Based on 2005 high-quality SNPs generated via double-digest restriction site-associated DNA sequencing (ddRADseq) from 275 individuals, we assessed genetic diversity, population differentiation, and fine-scale spatial genetic structure (FSGS) and further explored mating patterns through parentage analysis of 100 F1 offspring. The adult population maintained moderate genetic diversity (H_o = 0.243, F_IS = 0.082), whereas offspring cohorts showed stronger heterozygote deficiency (F_IS = 0.198), suggesting that high contemporary geitonogamous selfing may be counterbalanced by post-zygotic selective mortality during early life stages prior to adult recruitment. The two adult subpopulations separated by agricultural fields exhibited significant genetic differentiation (Φ_PT = 0.131, p = 0.001), indicating reduced gene exchange connectivity caused by landscape fragmentation. Significant FSGS was detected across the population and subpopulations, with three-dimensional spatial analyses revealing the influence of both spatial distance and host-tree distribution. Parentage analysis revealed a high rate of geitonogamy and a pattern in which single pollen donors fertilized multiple flowers on the same recipient plant. This indicates that pollinator movement is highly restricted, leading to localized mating within immediate flower clusters. Our results demonstrate that the genetic structure of R. gigantea is shaped by the combined effects of mixed mating systems, limited seed dispersal, host-tree dependence, and habitat fragmentation. Conservation of epiphytic orchids should therefore integrate the protection of orchid populations, host trees, and landscape connectivity to maintain long-term genetic diversity.