Abstract
Chronic bacterial infections are difficult to eradicate, though they are caused primarily by drug-susceptible pathogens. Antibiotic-tolerant persisters largely account for this paradox. In spite of their significance in the recalcitrance of chronic infections, the mechanism of persister formation is poorly understood. We previously reported that a decrease in ATP levels leads to drug tolerance in Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus We reasoned that stochastic fluctuation in the expression of tricarboxylic acid (TCA) cycle enzymes can produce cells with low energy levels. S. aureus knockouts in glutamate dehydrogenase, 2-oxoketoglutarate dehydrogenase, succinyl coenzyme A (CoA) synthetase, and fumarase have low ATP levels and exhibit increased tolerance of fluoroquinolone, aminoglycoside, and β-lactam antibiotics. Fluorescence-activated cell sorter (FACS) analysis of TCA genes shows a broad Gaussian distribution in a population, with differences of over 3 orders of magnitude in the levels of expression between individual cells. Sorted cells with low levels of TCA enzyme expression have an increased tolerance of antibiotic treatment. These findings suggest that fluctuations in the levels of expression of energy-generating components serve as a mechanism of persister formation.IMPORTANCE Persister cells are rare phenotypic variants that are able to survive antibiotic treatment. Unlike resistant bacteria, which have specific mechanisms to prevent antibiotics from binding to their targets, persisters evade antibiotic killing by entering a tolerant nongrowing state. Persisters have been implicated in chronic infections in multiple species, and growing evidence suggests that persister cells are responsible for many cases of antibiotic treatment failure. New antibiotic treatment strategies aim to kill tolerant persister cells more effectively, but the mechanism of tolerance has remained unclear until now.
Highlights
Chronic bacterial infections are difficult to eradicate, though they are caused primarily by drug-susceptible pathogens
Proteome analysis confirms that this is the case in a stationary-phase culture, where the level of tricarboxylic acid (TCA) cycle enzymes increases whereas the abundance of glycolytic enzymes decreases
The stationary phase is characterized by nutrient depletion, which accounts for previously observed low levels of ATP and high tolerance of antibiotics [5]
Summary
Chronic bacterial infections are difficult to eradicate, though they are caused primarily by drug-susceptible pathogens. Sorted cells with low levels of TCA enzyme expression have an increased tolerance of antibiotic treatment. These findings suggest that fluctuations in the levels of expression of energy-generating components serve as a mechanism of persister formation. Expression of cap5A and arcA was induced by treatment with arsenate, which depletes ATP through a futile cycle: ADP-As to ADP plus As (spontaneous hydrolysis) This suggests that these markers respond to ATP decreases and that the rare stationary-phase-like cells in a growing population have low September/October 2019 Volume 10 Issue 5 e01930-19. We show that cells in a growing population that have low levels of expression of tricarboxylic acid (TCA) cycle enzymes are tolerant of killing by antibiotics
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