Genomic and phenotypic evolution ofEscherichia coliin a novel citrate-only resource environment

Evolutionary innovations allow populations to colonize new, previously inaccessible ecological niches. We previously reported that aerobic growth on citrate (Cit+) evolved in a population ofEscherichia coliduring adaptation to a minimal glucose medium containing citrate (DM25). Cit+can grow in citrate-only medium (DM0), which is a novel environment forE. coli. To study adaptation to this new niche, we evolved one set of Cit+populations for 2,500 generations in DM0 and a control set in DM25. We identified numerous parallel mutations, many mediated by transposable elements. Several lineages evolved multi-copy amplifications containing themaeAgene, constituting up to ∼15% of the genome. We also found substantial cell death in ancestral and evolved clones. Our results demonstrate the importance of copy-number variation and transposable elements in the refinement of the Cit+trait. However, the observed mortality suggests a persistent evolutionary mismatch betweenE. coliphysiology and a citrate-only environment.