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Adaptation of Escherichia coli to Long-Term Serial Passage in Complex Medium: Evidence of Parallel Evolution.

ABSTRACT: 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.

SUBMITTER: Kram KE

PROVIDER: S-EPMC5340864 | biostudies-literature | 2017 Mar-Apr

REPOSITORIES: biostudies-literature

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Adaptation of <i>Escherichia coli</i> to Long-Term Serial Passage in Complex Medium: Evidence of Parallel Evolution.

Kram Karin E KE Geiger Christopher C Ismail Wazim Mohammed WM Lee Heewook H Tang Haixu H Foster Patricia L PL Finkel Steven E SE

mSystems 20170307 2

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 c ...[more]

PMID: 28289732

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Project description:BackgroundMicrobial metabolism is highly dependent on the environmental conditions. Especially, the substrate concentration, as well as oxygen availability, determine the metabolic rates. In large-scale bioreactors, microorganisms encounter dynamic conditions in substrate and oxygen availability (mixing limitations), which influence their metabolism and subsequently their physiology. Earlier, single substrate pulse experiments were not able to explain the observed physiological changes generated under large-scale industrial fermentation conditions.ResultsIn this study we applied a repetitive feast-famine regime in an aerobic Escherichia coli culture in a time-scale of seconds. The regime was applied for several generations, allowing cells to adapt to the (repetitive) dynamic environment. The observed response was highly reproducible over the cycles, indicating that cells were indeed fully adapted to the regime. We observed an increase of the specific substrate and oxygen consumption (average) rates during the feast-famine regime, compared to a steady-state (chemostat) reference environment. The increased rates at same (average) growth rate led to a reduced biomass yield (30% lower). Interestingly, this drop was not followed by increased by-product formation, pointing to the existence of energy-spilling reactions. During the feast-famine cycle, the cells rapidly increased their uptake rate. Within 10 s after the beginning of the feeding, the substrate uptake rate was higher (4.68 ?mol/gCDW/s) than reported during batch growth (3.3 ?mol/gCDW/s). The high uptake led to an accumulation of several intracellular metabolites, during the feast phase, accounting for up to 34% of the carbon supplied. Although the metabolite concentrations changed rapidly, the cellular energy charge remained unaffected, suggesting well-controlled balance between ATP producing and ATP consuming reactions.ConclusionsThe adaptation of the physiology and metabolism of E. coli under substrate dynamics, representative for large-scale fermenters, revealed the existence of several cellular mechanisms coping with stress. Changes in the substrate uptake system, storage potential and energy-spilling processes resulted to be of great importance. These metabolic strategies consist a meaningful step to further tackle reduced microbial performance, observed under large-scale cultivations.

Dataset Information

Adaptation of Escherichia coli to Long-Term Serial Passage in Complex Medium: Evidence of Parallel Evolution.

Publications

Adaptation of <i>Escherichia coli</i> to Long-Term Serial Passage in Complex Medium: Evidence of Parallel Evolution.

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