Inexpensive whole genome sequencing now permits us to look at genome-wide patterns of adaptation within populations, but this approach is only starting to be used to examine the extensive population structure that arises under long-term evolution studies. To determine the effects that effective population size and variable mutation rates have on genome evolution, we conducted a long-term evolution experiment using Escherichia coli populations with six different genetic backgrounds, carried through 3 years of continuous culture. After whole-population sequencing of these lines, we identified recurring mutations across populations—specifically genes associated with biofilm formation, global regulation, and peptidoglycan recycling.
To further investigate this phenomenon, we isolated eight random clones from twelve long-term evolution populations polymorphic for mutations in fimE (biofilm formation), hns (global regulation), and nlpD/mppA (peptidoglycan recycling). These twelve populations also represent three of the genetic backgrounds used: wild-type (WT), ΔmutL, and a mutL+ line preloaded with mutations for ~4000 generations of mutation accumulation. Genomic sequence of the clones, finds genetic evidence for subpopulation structure in the long-term evolution populations. Further, we find phenotypic differences within populations for lag-time, maximum growth rate, diauxic growth, auxotrophy, and biofilm formation.
For our wild-type focus population—where mutation rate is low—we identified three major haplotypes among the eight clones. Associated with these haplotypes, we identified genetic and phenotypic evidence of auxotrophy for thiamine in five clones. Additionally, these five clones displayed slower maximum growth rates, but shorter lag times and diauxic growth curves (indicating resource prioritization), presenting one possible adaption to coexistence in a fluctuating environment.
In our ΔmutL focus population, with mutation rates ~150 times greater than wild-type, we identified six major haplotypes. Furthermore, because of the increased mutation rate, we were able to conduct pathway analyses shedding light on the specialization of each clone. Interestingly, because of the potential for non-competing niches, independent evolutionary changes within identical pathways occurred in clones belonging to different haplotypes. We conclude that our methods gives us the mutational resolution to describe evolutionary changes within hetrogenous, intraspecies communities, an important step toward understanding intraspecies communities in both nature and disease.