Convergent Metabolic Specialization through Distinct Evolutionary Paths in Pseudomonas aeruginosa

Evolution by natural selection under complex and dynamic environmental conditions occurs through intricate and often counterintuitive trajectories affecting many genes and metabolic solutions. To study short- and long-term evolution of bacteria in vivo, we used the natural model system of cystic fibrosis (CF) infection. In this work, we investigated how and through which trajectories evolution of Pseudomonas aeruginosa occurs when migrating from the environment to the airways of CF patients, and specifically, we determined reduction of growth rate and metabolic specialization as signatures of adaptive evolution. We show that central metabolic pathways of three distinct Pseudomonas aeruginosa lineages coevolving within the same environment become restructured at the cost of versatility during long-term colonization. Cell physiology changes from naive to adapted phenotypes resulted in (i) alteration of growth potential that particularly converged to a slow-growth phenotype, (ii) alteration of nutritional requirements due to auxotrophy, (iii) tailored preference for carbon source assimilation from CF sputum, (iv) reduced arginine and pyruvate fermentation processes, and (v) increased oxygen requirements. Interestingly, although convergence was evidenced at the phenotypic level of metabolic specialization, comparative genomics disclosed diverse mutational patterns underlying the different evolutionary trajectories. Therefore, distinct combinations of genetic and regulatory changes converge to common metabolic adaptive trajectories leading to within-host metabolic specialization. This study gives new insight into bacterial metabolic evolution during long-term colonization of a new environmental niche.IMPORTANCE Only a few examples of real-time evolutionary investigations in environments outside the laboratory are described in the scientific literature. Remembering that biological evolution, as it has progressed in nature, has not taken place in test tubes, it is not surprising that conclusions from our investigations of bacterial evolution in the CF model system are different from what has been concluded from laboratory experiments. The analysis presented here of the metabolic and regulatory driving forces leading to successful adaptation to a new environment provides an important insight into the role of metabolism and its regulatory mechanisms for successful adaptation of microorganisms in dynamic and complex environments. Understanding the trajectories of adaptation, as well as the mechanisms behind slow growth and rewiring of regulatory and metabolic networks, is a key element to understand the adaptive robustness and evolvability of bacteria in the process of increasing their in vivo fitness when conquering new territories.