Figure 1. Experimental evolution of E. coli to increasing EtOH concentrations. (a) Setup of the evolution experiment with increasing percentage of EtOH. Initially, ancestral cells were grown in the presence of 5% EtOH, the condition that mimics near-lethal stress. Populations that grew until exponential phase were transferred to fresh medium while simultaneously increasing EtOH concentrations with 0.5%. (b) Evolutionary outcome of 20 independent parallel lines. Eight parallel lines evolved to high EtOH tolerance (shown in red). The other 12 lines were only able to acquire low EtOH-tolerance levels (shown in blue). For each line, the relative time (in generations) it spent growing on a certain percentage of EtOH is shown. For details, see Swings et al., 2017.
Mutation rate dynamics in evolutionary tolerance engineering
Changes in cellular mutation rate have mostly been studied under conditions that are mildly stressful to the cells.
Under these conditions, an increased mutation rate can provide rapid adaptation, but the cost of accumulating random and mostly harmful mutations limits the frequency of hypermutation. Biofuels, and more specifically ethanol, usually cause complex and near-lethal stress in the producing organism.
Our experiments show that hypermutation occurs rapidly under extreme stress and is essential for bacteria to adapt to the level of alcohol and to avoid extinction. Populations of bacteria in which hypermutation did not occur were unable to develop tolerance to the alcohol and went to extinction (Figure 1). Further experiments show that an individual population of bacteria can alter the mutation rate repeatedly as a result of changing stress levels. We identified cellular mortality as a critical cost of hypermutation and a major modulator of the cellular mutation rate.
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