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Published on: February 23, 2021
Role of oxidative stress in persister tolerance
Yanxia Wu1, Marin Vulić, Iris Keren
1Antimicrobial Discovery Center, Department of Biology, Northeastern University, Boston, Massachusetts, USA.
Abstract:
Persisters are dormant phenotypic variants of regular cells that are tolerant to antibiotics and play an important role in recalcitrance of chronic infections to therapy. Persisters can be produced stochastically in a population untreated with antibiotics. At the same time, a deterministic component of persister formation has also been documented in a population of cells with DNA damaged by fluoroquinolone treatment. Expression of the SOS response under these conditions induces formation of persisters by increasing expression of the TisB toxin. This suggests that other stress responses may also contribute to persister formation. Of particular interest is oxidative stress that pathogens encounter during infection. Activated macrophages produce reactive oxygen and nitrogen species which induce the SoxRS and OxyR regulons. Genes controlled by these regulons deactivate the oxidants and promote repair. We examined the ability of oxidative stress induced by paraquat (PQ) to affect persister formation. Preincubation of cells with PQ produced a dramatic increase in the number of persisters surviving challenge with fluoroquinolone antibiotics. PQ did not affect killing by kanamycin or ampicillin. Persisters in a culture treated with PQ that survived a challenge with a fluoroquinolone were also highly tolerant to other antibiotics. PQ induces SoxRS, which in turn induces expression of the AcrAB-TolC multidrug-resistant (MDR) pump. Fluoroquinolones are extruded by this MDR pump, and the effect of PQ on antibiotic tolerance was largely abolished in a mutant that was defective in the pump. It appears that PQ, acting through AcrAB-TolC, reduces the concentration of fluoroquinolones in the cells. This allows a larger fraction of cells to become persisters in the presence of a fluoroquinolone. Analysis of a lexA3 mutant indeed showed a dependence of persister induction under these conditions on SOS. These findings show that induction of a classical resistance mechanism, MDR efflux, by oxidative stress leads to an increase in multidrug-tolerant persister cells.
Insights
Oxidative stress, induced by paraquat (PQ), significantly increases antibiotic-tolerant persister cells by activating the multidrug-resistant (MDR) efflux pump. This mechanism enhances survival against fluoroquinolone antibiotics in chronic infections.
Area of Science:
- Microbiology
- Molecular Biology
- Drug Resistance
Background:
- Persister cells are dormant variants contributing to chronic infection treatment failure.
- Antibiotic treatment can induce persister formation, particularly with DNA-damaging agents like fluoroquinolones via the SOS response.
- Oxidative stress, encountered during infection, activates bacterial stress response pathways.
Purpose of the Study:
- To investigate the role of oxidative stress in persister cell formation.
- To determine if oxidative stress enhances tolerance to antibiotics.
- To elucidate the molecular mechanisms underlying oxidative stress-induced persister formation.
Main Methods:
- Exposure of bacterial cultures to paraquat (PQ) to induce oxidative stress.
- Challenging PQ-treated cultures with various antibiotics (fluoroquinolones, kanamycin, ampicillin).
- Assessing persister cell survival and cross-tolerance to antibiotics.
- Investigating the role of the SoxRS regulon and the AcrAB-TolC multidrug-resistant (MDR) pump using mutant strains.
- Analyzing the involvement of the SOS response in persister induction.
Main Results:
- Preincubation with PQ dramatically increased persister survival against fluoroquinolones.
- PQ-induced tolerance was specific to fluoroquinolones and not observed with kanamycin or ampicillin.
- Persisters formed under PQ treatment exhibited cross-tolerance to other antibiotics.
- PQ induced the SoxRS regulon, leading to AcrAB-TolC MDR pump expression, which was crucial for PQ's effect on fluoroquinolone tolerance.
- The effect of PQ was largely abolished in an AcrAB-TolC deficient mutant, indicating reduced intracellular fluoroquinolone concentration.
- Persister induction under these conditions was dependent on the SOS response.
Conclusions:
- Oxidative stress can significantly enhance persister cell formation and multidrug tolerance.
- The AcrAB-TolC MDR efflux pump, induced by oxidative stress via SoxRS, plays a key role in mediating this increased tolerance by reducing intracellular antibiotic levels.
- These findings highlight a novel mechanism linking bacterial stress responses to antibiotic tolerance and persister cell generation, relevant for recalcitrant infections.
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