Species delimitation using multiple sources of evidence from the Pinus strobiformis-Pinus ayacahuite Species Complex

Background: The Trans-Mexican Volcanic Belt (TMVB) in central Mexico is characterized by peaks of high altitude and geologic instability. In this zone, Pinus strobiformis and Pinus ayacahuite form a contact zone with Pinus veitchii. The taxonomical circumscription of white pines in Central Mexico has been unstable, especially regarding the status of P. veitchii. Questions: What are the species boundaries of the montane Mexican white pines species complex? Is Pinus veitchii a hybrid or an independently evolving lineage? Studied species: Pinus strobiformis, Pinus veitchii and Pinus ayacahuite species complex. Study site and dates: United States of America and Mexico from 2003 to 2022. Methods: We performed multivariate analyses on 10 morphological characters and characterized the climatic niche divergence and the genetic differentiation using SNPs. Results: Our results showed that P. veitchii is morphologically similar to P. strobiformis, but does not have intermediate morphological values with P. ayacahuite. The ecological niche differentiation was not significant.  Genetic analyses showed P. veitchii as an independent lineage with evidence of admixture with P. ayacahuite, suggesting a gene flow but not a hybrid origin. Conclusions: Two of the three lines of evidence support three independent lineages. Environmental information showed niche conservatism, morphology and genetic structure showed differentiation of all three taxa, with a greater morphological similarity between P. strobiformis and P. veitchii, and genetic analyses recovered evidence of introgression, suggesting a complex demographic history in the Trans Mexican Volcanic Belt.