Bacterial persistence and proton-motive force

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ABSTRACT Bacterial persistence is a robust mechanism employed by bacteria to survive antibiotic stress, and is likely to be a major cause of chronic infections. Bacterial persisters are genetically indistinct from the rest of the population, and likely emerge due to phenotypic variations that are inherently stochastic. Although all major bacterial pathogens produce persisters, the mechanisms of persistence are poorly understood. Persisters do not exhibit any discernible morphological differences from the rest of the cell population. This adds to the challenges of studying how persisters emerge in natural bacterial populations. Among the known mechanisms involve toxin-antitoxin (TA) modules, which dissipate the proton-motive force (PMF) and promote persister formation. Persisters may also arise due to energetic deficiencies, which induce dormancy in cells. However, persister populations are quite heterogeneous in many regards and identifying the mechanisms of persistence will continue to remain challenging until the sources of heterogeneity are determined. In the proposed work, the objective is to quantify the heterogeneity in the PMF of persisters and study key factors that influence fitness in persister populations. The long-term goal is to develop a mechanistic understanding of the stochastic phenomenon governing persistence. The proposed work is innovative as it combines single-cell biophysical assays including optical trapping with fluorescence techniques and quantitative image analysis – these tools will be used to quantify the heterogeneities in the PMF that might prevail in persister populations of Escherichia coli. The innovative approaches proposed will also help test correlations between the measured heterogeneities and the occurrence of persistence under different antibiotic stresses. In particular, the central hypothesis that low energy levels can predict the probability of persister formation will be tested. Successful execution of this project will lay the foundation for future efforts to determine how bacterial energetics influence persistence mechanisms. As persisters likely play a major role in clinical recurrence of disease, they are a major burden on the public healthcare. This coupled with the emerging threat of antibiotic resistance makes the proposed work timely and critical.