Can Altered Bioenergetics Drive Antibiotic Persistence in Low-Oxygen Conditions?
Abstract
Topic Area: Antimicrobial Resistance Challenge: Antibiotic-resistant bacteria pose a major threat to health care in both military and civilian populations, accounting for at least 2 million infections and 23,000 deaths annually in the U.S. alone. Antibiotic resistance is greatly compounded by antibiotic persistence – a phenomenon where a small fraction of infecting bacteria (known as persisters) can withstand lethal doses of antibiotics despite lacking any genetic mechanisms of resistance. Once treatment ceases, persisters resume growth, thereby posing a unique challenge for infectious disease management. Antibiotic persistence is extremely sensitive to external cues such as environmental oxygen. Notably, oxygen depletion (hypoxia) is associated with heightened persistence in most major pathogens including E. coli, M. tuberculosis, B. pseudomallei, S. aureus, A. baumannii, and P. aeruginosa. Yet, the mechanisms by which hypoxia potentiates bacterial persistence are incompletely understood. As hypoxia is a common feature of biofilms, granulomas, abscesses, and poorly perfused infected tissues, our ability to effectively treat infections and prevent relapse would benefit tremendously from a deeper understanding of how hypoxic microenvironments influence antibiotic persistence. Hypothesis, Innovation, and Proposed Approach: With support from the Peer Reviewed Medical Research Program, we expect to address the above challenge by investigating a potential link between hypoxia, single cell bioenergetics, and antibiotic persistence. Specifically, we hypothesize that anaerobic adaptation in bacteria exposed to hypoxic environments causes a sharp decrease in cellular energy (viz., ATP) in a subset of cells, likely driven by stochastic fluctuations in ATP homeostasis. We speculate that these sub-threshold, energy-deficient cells are the ones that successfully evade antibiotic lethality in an otherwise susceptible population. Bacteria reared in aerobic environments (such as aerated broth cultures) would not experience an ATP deficit to the same extent as anaerobically adapted bacteria owing to the higher ATP output of oxidative respiration. As a result, antibiotic persistence would be preferentially stimulated in hypoxic conditions, consistent with what is observed in most pathogens. Our hypothesis is motivated by recent studies that observed increased levels of persisters following chemically induced depletion of ATP in bacteria. However, these efforts did not investigate whether anaerobic adaptation can sufficiently lower ATP production in individual cells, prompting them to adopt a persistence phenotype. Such studies are challenging due to a lack of methods for long term, single cell tracking of ATP in hypoxic environments. Here, we propose to overcome this technical hurdle by engineering the first oxygen-independent fluorescent reporter for single cell ATP imaging. Next, we will use this ATP sensor to test our hypothesis by tracking how single cell ATP concentrations correlate with a bacterium’s ability to survive antibiotic treatment in hypoxic environments. If our hypothesis is correct, there should be an energetic threshold below which cells are primed to become persisters. By quantifying this ATP threshold, we will establish the first (to our knowledge) quantitative physiological predictor for persistence in single bacterial cells. Applicability and Impact: The core outcome of this research will be a deeper understanding of bacterial bioenergetics and antibiotic persistence in an understudied, yet clinically important context – hypoxia. The methods developed in this work can be readily adapted to study how various other aspects of bacterial physiology and metabolism (e.g., gene expression, cyclic-di-GMP signaling) are tied to a state of persistence at the single cell level in clinically relevant hypoxic conditions. A greater understanding of the mechanisms by which hypoxia potentiates persistence w
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Mar 10, 2021
- Source ID
- W81XWH2010101
Entities
People
- Arnab Mukherjee
Organizations
- United States Army
- University of California, Santa Barbara