Killing by Bactericidal Antibiotics Does Not Depend on Reactive Oxygen Species
Antimicrobial Discovery Center, Department of Biology, Northeastern University, Boston, MA 021156, USA.Science (Impact Factor: 33.61). 03/2013; 339(6124):1213-6. DOI: 10.1126/science.1232688
Bactericidal antibiotics kill by modulating their respective targets. This traditional view has been challenged by studies that propose an alternative, unified mechanism of killing, whereby toxic reactive oxygen species (ROS) are produced in the presence of antibiotics. We found no correlation between an individual cell's probability of survival in the presence of antibiotic and its level of ROS. An ROS quencher, thiourea, protected cells from antibiotics present at low concentrations, but the effect was observed under anaerobic conditions as well. There was essentially no difference in survival of bacteria treated with various antibiotics under aerobic or anaerobic conditions. This suggests that ROS do not play a role in killing of bacterial pathogens by antibiotics.
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- "A clear example of this is illustrated by the recent debate over oxidative stress as a general antimicrobial mechanism caused by antibiotics. These discussions are fueled by conflicting data over the potential involvement of ROS in bacterial killing by antibiotics      . New analytical tools for the measurement of oxidative stress in bacteria are desperately needed to resolve some of these uncertainties. "
ABSTRACT: Aerobic bacteria are continuously fighting potential oxidative stress due to endogenous and exogenous reactive oxygen species (ROS). To achieve this goal, bacteria possess a wide array of defenses and stress responses including detoxifying enzymes like catalases and peroxidases; however until now, the dynamics of the intra-bacterial redox balance remained poorly understood. Herein, we used redox-sensitive GFP (roGFP2) inside a variety of Gram-negative bacteria to study real-time redox dynamics immediately after a challenge with hydrogen peroxide. Using this biosensor, we determined the individual contributions of catalases and peroxidases and found that each enzyme contributes more to rapid detoxification or to prolonged catalytic activity. We also found that the total catalytic power is affected by environmental conditions. Additionally, using a Salmonella strain that is devoid of detoxifying enzymes, we examined endogenous ROS production. By measuring endogenous ROS production, we assessed the role of oxidative stress in toxicity of heavy metals and antibiotics. We found that exposure to nickel induced significant oxidative stress whereas cobalt (which was previously implicated to induce oxidative stress) did not induce ROS formation. Since a turbulent debate evolves around oxidative stress as a general killing mechanism by antibiotics (aminoglycosides, fluoroquinolones and β-lactams), we measured oxidative stress in bacteria that were challenged with these antibiotics. Our results revealed that antibiotics do not induce ROS formation in bacteria thereby disputing a role for oxidative stress as a general killing mechanism. Together, our results expose how the intra-bacterial redox balance in individual microorganisms is affected by environmental conditions and encounters with stress-inducing compounds. These findings demonstrate the significant potential of roGFP2 as a redox biosensor in Gram-negative bacteria to investigate redox dynamics under a variety of circumstances.
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- "In mammalian cells, bactericidal antibiotics induce mitochondrial dysfunction by disrupting the electron transport chain, which triggers dose-and time-dependent overproduction of reactive oxygen species (ROS), thereby leading to oxidative stress damages (Kalghatgi et al. 2013). However, several reports demonstrated that ROS is not involved in bacterial cell death (Ezraty et al. 2013; Liu et al. 2013; Keren et al. 2013). Aberrant ROS production has always been associated with cellular oxidative damage and death. "
ABSTRACT: The aminoglycoside antibiotic hygromycin B (Hyg) inhibits prokaryotic, chloroplast and mitochondrial protein synthesis. Because of the toxic effect of Hyg on plant cells, the HPT gene, encoding hygromycin phosphotransferase, has become one of the most widely used selectable markers in plant transformation. Yet the mechanism behind Hyg-induced cell lethality in plants is not clearly understood. In this study, we aimed to decipher this mechanism. With Hyg treatment, rice calli exhibited cell death, and rice seedlings showed severe growth defects, leaf chlorosis and leaf shrinkage. Rice seedlings also exhibited severe lipid peroxidation and protein carbonylation, for oxidative stress damage at the cellular level. The production of reactive oxygen species such as O 2·−, H2O2 and OH· was greatly induced in rice seedlings under Hyg stress, and pre-treatment with ascorbate increased resistance to Hyg-induced toxicity indicating the existence of oxidative stress. Overexpression of mitochondrial Alternative oxidase1a gene without HPT selection marker in rice enhanced tolerance to Hyg and attenuated the degradation of protein content, whereas the rice plastidial glutathione reductase 3 mutant showed increased sensitivity to Hyg. These results demonstrate that Hyg-induced cell lethality in rice is not only due to the inhibition of protein synthesis but also mediated by oxidative stress.
- "Recent studies, however, point to the existence of antibiotic killing mechanisms that are not dependent on OH • induction in bacteria (Brochmann et al., 2013; Keren et al., 2013; Liu & Imlay, 2013). For ciprofloxacin, this fits into a scheme containing two lethal pathways of quinolones: one is blocked by inhibitors of protein synthesis and by anaerobic conditions (the chloramphenicol-sensitive, ROS-dependent) and another is active even in the presence "
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