Persistence and Metabolic Activity in Ofloxacin-treated, Growth-inhibited Populations of Escherichia coli

Abstract

Resistance to antibiotic therapies is an increasing global health problem, and nonheritable bacterial persistence poses an independent set of challenges. Persisters are bacterial cells that can tolerate antibiotic treatment independent of resistance genes and naturally comprise a small fraction of bacterial populations. Persisters are thought to be responsible for relapse infections, with nongrowing populations being particularly difficult to kill. We hypothesized that the cellular metabolic activity in growth-inhibited populations influences the likelihood that a bacterium is a persister during fluoroquinolone treatment. While there have previously been several investigations into persister metabolism focusing on metabolic activity prior to antibiotic treatment, this study examines metabolism during and after antibiotic treatment. We used the fluorescent dye Redox Sensor Green (RSG) to characterize the metabolic activity of stationary-phase Escherichia coli populations during and after ofloxacin treatment, followed by fluorescence-activated cell sorting (FACS) to assess the dependence of persistence on metabolic activity during and after treatment. During treatment, persisters demonstrated a bias for low metabolic activity and were the least likely to be associated with the most metabolically active population; however, culturability was also highest in cells from the population with the lowest metabolic activity, making it difficult to draw strong conclusions. After treatment, cells with above average metabolic activity were most likely to become persisters while those with the highest metabolism were still the least likely to be persisters. This study confirms some association between metabolism and persistence, but future work may include investigating the metabolic response of persisters using alternative techniques such as time-lapse microscopy, exploring different nutrient and environment conditions, or more closely examining the metabolic processes during treatment that enable survival.