Agamospermy as a mechanism to maintain species boundaries in sympatric Goodyera species

Abstract Premise Diverse mechanisms of reproductive isolation can limit gene flow between species. The Orchidaceae, one of the most species‐rich and morphologically diverse plant families, often exhibits strong pollinator‐mediated isolation due to highly specialized pollination systems. Autonomous self‐pollination is also common in orchids and contributes to both speciation and the maintenance of reproductive barriers among closely related taxa. Although agamospermy (clonal seed production without fertilization) occurs in several orchid species, its role in reproductive isolation remains unclear. Methods We tested whether agamospermy serves as a reproductive barrier between the agamospermous Goodyera crassifolia and the pollinator‐dependent G. schlechtendaliana by assessing pollen germination, fruit, and seed production after interspecific and intraspecific pollination in conjunction with high‐throughput genomic analysis. Results Goodyera crassifolia pollinia failed to germinate on conspecific and heterospecific stigmas. In contrast, G. schlechtendaliana pollinia germinated readily on conspecific stigmas, with pollen tubes reaching the column base, but did not germinate on G. crassifolia stigmas. No fruit set occurred after G. crassifolia pollinia were placed on G. schlechtendaliana stigmas, whereas G. schlechtendaliana pollinia on G. crassifolia stigmas resulted in seeds that lacked paternal genetic contribution. No evidence of hybridization between G. crassifolia and G. schlechtendaliana was detected in natural populations. Conclusions The agamospermous reproductive mode of G. crassifolia likely serves as an effective barrier to interspecific gene flow from sympatric G. schlechtendaliana at the study site. To our knowledge, this is the first detailed investigation of how agamospermy functions as a mechanism of reproductive isolation in orchids.