2023 AIChE Annual Meeting
Proton Motive Force in E. coli Persisters
Persisters are bacterial cells that can survive lethal doses of antibiotics despite lacking genes responsible for antibiotic resistance. Persisters are assumed to be dormant with a low metabolic activity that protects against antibiotics. Once the antibiotic stress decreases, the persister cells resurrect and repopulate, contributing to chronic infections and antibiotic resistance. Understanding of persister physiology, though, is currently limited. We focused on the proton motive force (PMF) in âinducedâ persisters as the PMF fuels several cellular processes, including the uptake of antibiotics. The previous notion in the field was that a low PMF promotes persistence by inducing dormancy. However, our recent findings suggest that the PMF varies considerably in populations of Escherichia coli persisters. Those conclusions were drawn from motility studies, which indicated that a part of the persister population swims robustly. As PMF powers motility in E. coli, this suggests the existence of two sub-populations within the persisters: persisters with low PMF and persisters with high PMF. We quantified the heterogeneity in PMF at a single-cell level using dyes that label cells as a function of their membrane potential, which is the major contributor to the PMF. Briefly, we treated cells with an antibiotic, rifampicin, which induces persistence by inhibiting transcription. We subsequently exposed the cells to lethal concentrations of ampicillin to concentrate persisters and eliminate the remaining population. We then labeled the cells with Thioflavin T (ThT) dye and used photon-sensitive cameras to record fluorescent signals from single cells. We observed a bimodal distribution in the intensity of fluorescence signals in persister populations, indicating that some persisters retained high energy levels, whereas the rest were metabolically dormant. Future experiments are anticipated to help determine if the so-called persisters employ PMF-dependent efflux pumps or decrease their membrane permeability to tolerate antibiotic stress. Insights from this research are likely to help develop therapeutics against persisters.