Stochastic processes and changes in evolutionary rate are associated with diversification in a lineage of tropical hard pines (Pinus)

The study of the patterns of polymorphism and molecular evolution among closely related species is key to understanding the evolutionary forces involved in the diversification of lineages. This point is a big challenge in species with slow evolutionary rates, long life cycles, and ancient, shared polymorphisms such as conifers. Under the premise of divergence in a stepwise migration process, we expect clinal geographical patterns of purifying selection efficiency, and genetic structure related to latitude or longitude. If migration is accompanied by changes in the environment, we could further expect a role of positive selection in driving species divergence. Here, we infer patterns of polymorphism, efficiency of purifying selection, and molecular evolution using a dataset of 161 nuclear genes (similar to 71 Kb) in a lineage of hard pines from North America, the Caribbean, Mexico, and Central America presumed to have migrated from North America toward lower latitudes with tropical conditions. Under the premise of differences in selective pressures, we also look for possible signals of positive selection. To test our hypothesis, first we estimated different indices to infer patterns of polymorphism and efficiency of purifying selection (K-a, K-s, K-a/K-s, d(N), d(S), d(N)/d(S), and d(xy)) and compared these metrics across five clades. Also, we investigated possible clinal patterns in these indices and morphological traits (needle length and cone length). Then we inferred genetic structure and environmental differences among species to test for possible signals of positive selection using phylogenetic methods in specific clades. We found differences among clades using K-a, K-s, and K-a/K-s with a relaxation of purifying selection, especially in the Elliotti and Patula clades. We also found environmental differences related to geographic distance, and among clades suggesting differences in selective pressures. The indices K-s, d(xy), and needle length had relationships with geography but not ovulate cone length. Finally, we found that most analyzed genes are under purifying selection, but there was an exception of faster evolutionary rate in some pine species, suggesting the possible action of positive selection in divergence. Our study indicated that stochastic processes have played a key role in the diversification of the group, with a possible input of positive selection in pines from Mexico and Central America.