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Population and Single-Cell Analysis of Antibiotic Persistence in Escherichia coli
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Birth and Resuscitation of (p)ppGpp Induced Antibiotic Tolerant Persister Cells

Mikkel Skjoldan Svenningsen1, Alexandra Veress2, Alexander Harms2

  • 1Niels Bohr Institute, University of Copenhagen, Blegdamsvej 17, 2100 København Ø, København, Denmark.

Scientific Reports
|April 17, 2019
PubMed

Insights

High levels of the second messenger (p)ppGpp are crucial for bacterial persister cell formation. However, persister cell antibiotic tolerance is stochastic and not directly linked to (p)ppGpp levels, suggesting a transcriptional response drives formation.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Bacterial Physiology

Background:

  • Transient antibiotic treatment eliminates most bacteria, leaving behind a small subpopulation of dormant, tolerant cells known as persisters.
  • The guanosine tetraphosphate (p)ppGpp molecule is a key second messenger implicated in persister cell formation in Escherichia coli, but its precise role remains unclear.
  • Existing hypotheses suggest persisters have low ATP levels or are regulated by toxin-antitoxin (TA) modules, but these have not been definitively proven.

Purpose of the Study:

  • To investigate the mechanisms underlying persister cell formation in Escherichia coli.
  • To directly observe the stochastic nature of persister cell appearance, antibiotic tolerance, and resuscitation.
  • To elucidate the role of the second messenger (p)ppGpp in persister cell development.

Main Methods:

  • Induced (p)ppGpp synthesis by modulating tRNA charging in Escherichia coli.
  • Utilized live microscopy to monitor persister cell formation, antibiotic tolerance, and resuscitation.
  • Employed fluorescent reporters to continuously track physiological parameters of persister cells and their non-persister counterparts.

Main Results:

  • Confirmed that elevated (p)ppGpp levels are essential for persister formation.
  • Observed that persister formation is a stochastic process, with no direct correlation between single-cell (p)ppGpp levels and antibiotic tolerance.
  • Did not find evidence supporting low intracellular ATP concentrations or TA module activation as primary drivers of persister phenotypes.

Conclusions:

  • Persister cell formation under normal conditions is primarily driven by the transcriptional response to increased (p)ppGpp levels.
  • The stochastic nature of persister formation suggests complex regulatory networks beyond simple (p)ppGpp concentration.
  • Further research is needed to fully understand the multifaceted mechanisms governing bacterial persistence and antibiotic tolerance.

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