The Roles of Life History and Mating System in Speciation: Genomic Evidence From the <i>Incarvillea sinensis</i> Complex

The temporal sequence and mechanistic interplay between life-history and mating-system transitions in plants remain poorly understood. Here, we combine crossing experiments with population genomic analyses to investigate these processes in the Incarvillea sinensis complex (Bignoniaceae), which contains annual selfing and perennial outcrossing populations. Crossing experiments revealed complete post-zygotic isolation between annuals and perennials. Genomic data derived from de novo assemblies of annual and perennial individuals, along with SNP and chloroplast sequencing of 126 individuals from 30 populations, demonstrated strong genetic divergence between life histories, with no evidence of contemporary gene flow between them. Compared to perennials, annuals exhibited significantly reduced genetic diversity, elevated differentiation and a greater number of chromosomal rearrangements-particularly translocations. Divergence time estimates indicated that annual and perennial lineages split during the early Pleistocene (ca. 2.22 Mya), whereas the shift to selfing from outcrossing within the annual lineage was a more recent event, occurring during the late Pleistocene (~0.03 Mya). These results indicate that the mating-system shift was not the initial driver of divergence, and we infer that the life-history transition likely occurred earlier, although its precise timing could not be directly determined. Our findings support a two-stage model of divergence in which adaptation to seasonally arid environments first drove the evolution of annuality, initiating speciation through ecological isolation and genetic divergence. The subsequent evolution of selfing then likely further promoted reproductive isolation via both pre- and post-zygotic mechanisms, potentially accelerating rapid genomic differentiation and effectively completing the speciation process.