LAUSR

The testing association of environmental variables with genetic and epigenetic variation could be crucial to deciphering the effects of environmental factors playing roles as selective drivers in ecological speciation. Although ecological speciation may occur in closely related species, species boundaries may not be established over a short evolutionary timescale. Here, we investigated the genetic and epigenetic variations using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP), respectively, and tested their associations with environmental variables in populations of four closely related species in the R. pseudochrysanthum complex. No distinctive species relationships were found using genetic clustering analyses, neighbor-joining tree, and neighbor-net tree based on the total AFLP variation, which is suggestive of the incomplete lineage sorting of ancestral variation. Nonetheless, strong isolation-by-environment and adaptive divergence were revealed, despite the significant isolation-by-distance. Annual mean temperature, elevation, normalized difference vegetation index, and annual total potential evapotranspiration were found to be the most important environmental variables explaining outlier genetic and epigenetic variations. Our results suggest that the four closely related species of the R. pseudochrysanthum complex share the polymorphism of their ancestor, but reproductive isolation due to ecological speciation can occur if local environmental divergence persists over time.