Sigma S-Dependent Antioxidant Defense Protects Stationary-Phase Escherichia coli against the Bactericidal Antibiotic Gentamicin

Abstract

Stationary-phase bacteria are important in disease. The s -regulated general stress response helps them become resistant to disinfectants, but the role of s in bacterial antibiotic resistance has not been elucidated. Loss of s rendered stationary-phase Escherichia coli more sensitive to the bactericidal antibiotic gentamicin (Gm), and proteomic analysis suggested involvement of a weakened antioxidant defense. Use of the psfiA genetic reporter, 3=-(p-hydroxyphenyl) fluorescein (HPF) dye, and Amplex Red showed that Gm generated more reactive oxygen species (ROS) in the mutant. HPF measurements can be distorted by cell elongation, but Gm did not affect stationary-phase cell dimensions. Coadministration of the antioxidant N-acetyl cysteine (NAC) decreased drug lethality particularly in the mutant, as did Gm treatment under anaerobic conditions that prevent ROS formation. Greater oxidative stress, due to insufficient quenching of endogenous ROS and/or respiration-linked electron leakage, therefore contributed to the greater sensitivity of the mutant; infection by a uropathogenic strain in mice showed this to be the case also in vivo. Disruption of antioxidant defense by eliminating the quencher proteins, SodA/SodB and KatE/SodA, or the pentose phosphate pathway proteins, Zwf/Gnd and TalA, which provide NADPH for ROS decomposition, also generated greater oxidative stress and killing by Gm. Thus, besides its established mode of action, Gm also kills stationary-phase bacteria by generating oxidative stress, and targeting the antioxidant defense of E. coli can enhance its efficacy. Relevant aspects of the current controversy on the role of ROS in killing by bactericidal drugs of exponential-phase bacteria, which represent a different physiological state, are discussed.

RpoN Promotes Pseudomonas aeruginosa Survival in the Presence of Tobramycin

“…In contrast with our observations, the importance of RpoS in the response to aminoglycoside-induced stress has been demonstrated in E. coli (Wang et al, 2014). Furthermore, RpoS protects E. coli and V .…”

Ranking of persister genes in the same Escherichia coli genetic background demonstrates varying importance of individual persister genes in tolerance to different antibiotics

“…In contrast to other knockout mutants tested, the deletion mutant of rpoS displayed a defect in persistence to gentamicin, but a higher persistence phenotype than wild-type strain in exposure to ampicillin or norfloxacin. Our data are compatible with the previous observation ( Hong et al, 2012 ; Wang et al, 2014 ) that loss of RpoS renders stationary-phase E. coli more sensitive to gentamicin by generating more ROS to enhance oxidative stress, whereas compensatory mutations may have occurred in the RpoS mutant induced by ampicillin and norfloxacin, which led to a higher persistence phenotype. Future studies are needed to address this possibility.…”

Antibiotic efficacy is linked to bacterial cellular respiration

“…It has been hypothesized that bactericidal antibiotics lead to metabolic instability and the formation of toxic ROS as part of their lethality (28,29,35,36). Acceleration of cellular respiration by bactericidal antibiotics may be a potential source of ROS (45). Our work supports this model by showing that tuning rates of basal cellular respiration can significantly impact bactericidal efficacy.…”