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Adaptation and heterogeneity of Escherichia coli MC1000 growing in complex environments.

ABSTRACT: In a study aiming to assess bacterial evolution in complex growth media, we evaluated the long-term adaptive response of Escherichia coli MC1000 in Luria-Bertani (LB) medium. Seven parallel populations were founded and followed over 150 days in sequential batch cultures under three different oxygen conditions (defined environments), and 19 evolved forms were isolated. The emergence of forms with enhanced fitness was evident in competition experiments of all evolved forms versus the ancestral strain. The evolved forms were then subjected to phenotypic and genomic analyses relative to the ancestor. Profound changes were found in their phenotypes as well as whole-genome sequences. Interestingly, considerable heterogeneity was found at the intrapopulational level. However, consistently occurring parallel adaptive responses were found across all populations. The evolved forms all contained a mutation in galR, a repressor of the galactose operon. Concomitantly, the new forms revealed enhanced growth on galactose as well as galactose-containing disaccharides. This response was likely driven by the LB medium.

SUBMITTER: Puentes-Tellez PE

PROVIDER: S-EPMC3568559 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature

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Adaptation and heterogeneity of Escherichia coli MC1000 growing in complex environments.

Puentes-Téllez Pilar Eliana PE Hansen Martin Asser MA Sørensen Søren Johannes SJ van Elsas Jan Dirk JD

Applied and environmental microbiology 20121130 3

In a study aiming to assess bacterial evolution in complex growth media, we evaluated the long-term adaptive response of Escherichia coli MC1000 in Luria-Bertani (LB) medium. Seven parallel populations were founded and followed over 150 days in sequential batch cultures under three different oxygen conditions (defined environments), and 19 evolved forms were isolated. The emergence of forms with enhanced fitness was evident in competition experiments of all evolved forms versus the ancestral str ...[more]

PMID: 23204426

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Project description:Bacterial metabolism is necessary for adaptation to the host microenvironment. Flexible metabolic pathways allow uropathogenic Escherichia coli (UPEC) to harmlessly reside in the human intestinal tract and cause disease upon extraintestinal colonization. E. coli intestinal colonization requires carbohydrates as a carbon source, while UPEC extraintestinal colonization requires gluconeogenesis and the tricarboxylic acid cycle. UPEC containing disruptions in two irreversible glycolytic steps involving 6-carbon (6-phosphofructokinase; pfkA) and 3-carbon (pyruvate kinase; pykA) substrates have no fitness defect during urinary tract infection (UTI); however, both reactions are catalyzed by isozymes: 6-phosphofructokinases Pfk1 and Pfk2, encoded by pfkA and pfkB, and pyruvate kinases Pyk II and Pyk I, encoded by pykA and pykF UPEC strains lacking one or both phosphofructokinase-encoding genes (pfkB and pfkA pfkB) and strains lacking one or both pyruvate kinase genes (pykF and pykA pykF) were investigated to determine their regulatory roles in carbon flow during glycolysis by examining their fitness during UTI and in vitro growth requirements. Loss of a single phosphofructokinase-encoding gene has no effect on fitness, while the pfkA pfkB double mutant outcompeted the parental strain in the bladder. A defect in bladder and kidney colonization was observed with loss of pykF, while loss of pykA resulted in a fitness advantage. The pykA pykF mutant was indistinguishable from wild-type in vivo, suggesting that the presence of Pyk II rather than the loss of Pyk I itself is responsible for the fitness defect in the pykF mutant. These findings suggest that E. coli suppresses latent enzymes to survive in the host urinary tract.IMPORTANCE Urinary tract infections are the most frequently diagnosed urologic disease, with uropathogenic Escherichia coli (UPEC) infections placing a significant financial burden on the health care system by generating more than two billion dollars in annual costs. This, in combination with steadily increasing antibiotic resistances to present day treatments, necessitates the discovery of new antimicrobial agents to combat these infections. By broadening our scope beyond the study of virulence properties and investigating bacterial physiology and metabolism, we gain a better understanding of how pathogens use nutrients and compete within host microenvironments, enabling us to cultivate new therapeutics to exploit and target pathogen growth requirements in a specific host environment.

Project description:Experimental evolution of bacterial populations in the laboratory has led to identification of several themes, including parallel evolution of populations adapting to carbon starvation, heat stress, and pH stress. However, most of these experiments study growth in defined and/or constant environments. We hypothesized that while there would likely continue to be parallelism in more complex and changing environments, there would also be more variation in what types of mutations would benefit the cells. In order to test our hypothesis, we serially passaged Escherichia coli in a complex medium (Luria-Bertani broth) throughout the five phases of bacterial growth. This passaging scheme allowed cells to experience a wide variety of stresses, including nutrient limitation, oxidative stress, and pH variation, and therefore allowed them to adapt to several conditions. After every ~30 generations of growth, for a total of ~300 generations, we compared both the growth phenotypes and genotypes of aged populations to the parent population. After as few as 30 generations, populations exhibit changes in growth phenotype and accumulate potentially adaptive mutations. There were many genes with mutant alleles in different populations, indicating potential parallel evolution. We examined 8 of these alleles by constructing the point mutations in the parental genetic background and competed those cells with the parent population; five of these alleles were found to be adaptive. The variety and swiftness of adaptive mutations arising in the populations indicate that the cells are adapting to a complex set of stresses, while the parallel nature of several of the mutations indicates that this behavior may be generalized to bacterial evolution. IMPORTANCE With a growing body of work directed toward understanding the mechanisms of evolution using experimental systems, it is crucial to decipher what effects the experimental setup has on the outcome. If the goal of experimental laboratory evolution is to elucidate underlying evolutionary mechanisms and trends, these must be demonstrated in a variety of systems and environments. Here, we perform experimental evolution in a complex medium allowing the cells to transition through all five phases of growth, including death phase and long-term stationary phase. We show that the swiftness of selection and the specific targets of adaptive evolution are different in this system compared to others. We also observe parallel evolution where different mutations in the same genes are under positive natural selection. Together, these data show that while some outcomes of microbial evolution experiments may be generalizable, many outcomes will be environment or system specific.

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Adaptation and heterogeneity of Escherichia coli MC1000 growing in complex environments.

Publications

Adaptation and heterogeneity of Escherichia coli MC1000 growing in complex environments.

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