Long-term experimental evolution in Escherichia coli. XIII. Phylogenetic history of a balanced polymorphism

We investigated the phylogenetic history of a balanced polymorphism that evolved in an experimental population of Escherichia coli. Previous work showed that two ecologically and morphologically distinct types, designated L (large) and S (small), arose by generation 6000 and coexisted for more than 12,000 generations thereafter. Here, we performed RFLP analyses using Insertion Sequence elements to resolve the phylogenetic history of L and S. Specifically, we sought to determine whether the derived S morph was monophyletic, indicating a long history of coexistence with L or, alternatively, S was repeatedly regenerated from L, indicating a series of periods with only transiently stable coexistence. Phylogenetic analysis of some 200 clones collected throughout the history of this population demonstrates that S is monophyletic. We then performed competition assays using clones of both morphs from different generations to determine whether either or both lineages continued to undergo genetic adaptation. Indeed, both lineages continued to adapt, and their continued evolution contributed to fluctuations in their relative abundance over evolutionary time. Based on their phylogenetic history and independent evolutionary trajectories, S and L fulfill Cohan's criteria for being different asexual species.