Genetic basis of persister formation in Escherichia coli

Abstract

Persisters are a small population of dormant bacterial cells that are tolerant to antibiotics. Persister cells have been discovered in many notorious human pathogens and are often responsible for recalcitrance of chronic infections and failure of antibiotic therapy. The goal of my research is to understand the mechanism of persister formation using Escherichia coli as a model. I conducted the first genome-wide screen on persisters using the recently developed transposon-sequencing approach. I constructed a large transposon mutant library and quantified the contribution of every non-essential gene of E. coli to persister formation. The screen allowed me to identify multiple independent pathways that are involved in persister formation. I found that persister levels are significantly increased by activation of amino acid synthesis and motility. We also sought to determine whether various stresses can induce expression of mRNA interferase encoding toxins and persister formation. Among the five stresses we tested, only isoleucine starvation led to toxin-antitoxin dependent persisters, and the effect is limited to fluoroquinolones. The 16S ribosomal RNA promoter rrnB P1 was proposed to be a persister reporter due to activation of toxins. Through fluorescent based cell sorting, we found that the promoter reports persisters independently of toxin-antitoxin systems. Further investigation revealed that the rRNA promoter reports persisters by sensing intracellular ATP levels. A drop of ATP increases persisters by decreasing the activity of antibiotic targets. Noise in the levels of expression of energy generating processes, such as Krebs cycle enzymes, can affect persister formation. In conclusion, energy depletion largely determines persister formation in E. coli, and noise in energy generating components is an important mechanism of persister formation.