Stimulating Transcription in Antibiotic-Tolerant Escherichia coli Sensitizes It to Fluoroquinolone and Nonfluoroquinolone Topoisomerase Inhibitors

LaGree, Travis J.; Byrd, Brandon A.; Quelle, Ryan M; Schofield, Stephanie L; Mok, Wendy W. K.

doi:10.1128/aac.01639-22

Cited by 1 publication

(2 citation statements)

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“…In this study, we delved into the responses of stationary-phase P. aeruginosa to Levo treatment with the two most common reference strains: PAO1 and PA14 ( 35 ). In contrast to previous findings in E. coli fluoroquinolone persisters, we discovered that P. aeruginosa cells retain redox and transcriptional activities in the stationary phase and succumb to Levo during treatment ( 25 , 48 , 56 ). While the nature of clinical infections is far more complex than laboratory monocultures, our findings are nonetheless relevant to understanding how genetically susceptible P. aeruginosa cells survive antibiotic treatment.…”

Section: Discussioncontrasting

confidence: 99%

“…6A ). As expected, stationary-phase E. coli MG1655 has minimal radiolabeled uridine in its extracted nucleic acids, but stationary-phase P. aeruginosa PAO1 and PA14 both have appreciable nucleic acid synthesis activity ( 56 ). Degradation of alkaline-labile RNA in each sample with 3 M KOH before scintillation counting reduced the counts of tritiated uridine.…”

Section: Resultsmentioning

confidence: 54%

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Metabolic and transcriptional activities underlie stationary-phase Pseudomonas aeruginosa sensitivity to Levofloxacin

Hare,

Gonzalez,

Quelle

et al. 2024

Microbiol Spectr

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The ubiquitous opportunistic pathogen Pseudomonas aeruginosa is highly adaptive and refractory to several different classes of antibiotics. However, we found in this study that stationary-phase P. aeruginosa cultures exhibit greater sensitivity to the fluoroquinolone Levofloxacin (Levo) than other bactericidal antibiotics, including an aminoglycoside (Tobramycin) and β-lactam (Aztreonam). To understand the basis of this sensitivity, we conducted time-lapse fluorescence microscopy experiments of cells during Levo treatment. We discovered that stationary-phase P. aeruginosa cells die rapidly during treatment and undergo heterogeneous morphological changes, including explosive lysis, filamentation, and gradual loss of membrane integrity as evidenced by propidium iodide uptake. These morphologies are reminiscent of how the model organism Escherichia coli appears when recovering from fluoroquinolone treatment, a period when activation of the DNA damage-induced SOS response is crucial. Accordingly, we monitored the morphologies and survival of P. aeruginosa Δ recA mutants and found that the SOS response is not involved in P. aeruginosa Levo sensitivity like it is for E. coli . We hypothesized that Levo sensitivity may be due to P. aeruginosa maintaining active metabolism in stationary phase. We determined that stationary-phase P. aeruginosa cells transcribe, maintain reductase activity, and accumulate reactive metabolic species which contribute to Levo-mediated death. By elucidating how P. aeruginosa cells sustain metabolic activity during the stationary phase, we can design strategies to sensitize these persistent subpopulations to Levo and maintain the efficacy of this clinically important fluoroquinolone antibiotic. IMPORTANCE The bacterial pathogen Pseudomonas aeruginosa is responsible for a variety of chronic human infections. Even in the absence of identifiable resistance mutations, this pathogen can tolerate lethal antibiotic doses through phenotypic strategies like biofilm formation and metabolic quiescence. In this study, we determined that P. aeruginosa maintains greater metabolic activity in the stationary phase compared to the model organism, Escherichia coli , which has traditionally been used to study fluoroquinolone antibiotic tolerance. We demonstrate that hallmarks of E. coli fluoroquinolone tolerance are not conserved in P. aeruginosa , including the timing of cell death and necessity of the SOS DNA damage response for survival. The heightened sensitivity of stationary-phase P. aeruginosa to fluoroquinolones is attributed to maintained transcriptional and reductase activity. Our data suggest that perturbations that suppress transcription and respiration in P. aeruginosa may actually protect the pathogen against this important class of antibiotics.

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