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A role for the stringent response in ciprofloxacin resistance in Pseudomonas aeruginosa
Libertad García-Villada1, Natalya P Degtyareva1, Ashley M Brooks2
1Genomic Integrity and Structural Biology Laboratory, NIEHS, Durham, NC, USA.
Abstract:
Pseudomonas aeruginosa is a major cause of nosocomial infections and the leading cause of chronic lung infections in cystic fibrosis and chronic obstructive pulmonary disease patients. Antibiotic treatment remains challenging because P. aeruginosa is resistant to high concentrations of antibiotics and has a remarkable ability to acquire mutations conferring resistance to multiple groups of antimicrobial agents. Here we report that when P. aeruginosa is plated on ciprofloxacin (cipro) plates, the majority of cipro-resistant (ciproR) colonies observed at and after 48 h of incubation carry mutations in genes related to the Stringent Response (SR). Mutations in one of the major SR components, spoT, were present in approximately 40% of the ciproR isolates. Compared to the wild-type strain, most of these isolates had decreased growth rate, longer lag phase and altered intracellular ppGpp content. Also, 75% of all sequenced mutations were insertions and deletions, with short deletions being the most frequently occurring mutation type. We present evidence that most of the observed mutations are induced on the selective plates in a subpopulation of cells that are not instantly killed by cipro. Our results suggests that the SR may be an important contributor to antibiotic resistance acquisition in P. aeruginosa.
Insights
Pseudomonas aeruginosa develops ciprofloxacin resistance through mutations in Stringent Response (SR) genes. These genetic changes, often short deletions, impact bacterial growth and highlight SR
Area of Science:
- Microbiology
- Molecular Biology
- Antimicrobial Resistance Research
Background:
- Pseudomonas aeruginosa is a significant cause of hospital-acquired infections and chronic lung infections in patients with cystic fibrosis and COPD.
- P. aeruginosa exhibits high-level antibiotic resistance and rapidly acquires mutations for multi-drug resistance, complicating treatment.
- Ciprofloxacin is a critical antibiotic for treating P. aeruginosa infections.
Purpose of the Study:
- To investigate the genetic basis of ciprofloxacin resistance in P. aeruginosa.
- To identify specific bacterial pathways involved in the acquisition of antibiotic resistance.
- To understand the role of the Stringent Response (SR) in P. aeruginosa antibiotic resistance.
Main Methods:
- P. aeruginosa strains were cultured on ciprofloxacin-containing agar plates.
- Resistant colonies appearing after 48 hours of incubation were isolated and analyzed.
- Genetic sequencing was performed to identify mutations in SR-related genes, particularly spoT.
- Phenotypic characterization of resistant isolates included growth rate and intracellular ppGpp levels.
Main Results:
- The majority of ciprofloxacin-resistant (ciproR) P. aeruginosa colonies carried mutations in Stringent Response (SR) genes.
- Mutations in spoT, a key SR component, were found in approximately 40% of ciproR isolates.
- Resistant isolates exhibited reduced growth rates, extended lag phases, and altered intracellular ppGpp levels compared to wild-type.
- Insertions and deletions, predominantly short deletions, constituted 75% of the identified mutations.
- Evidence suggests these mutations are induced on selective plates in a resistant subpopulation.
Conclusions:
- The Stringent Response (SR) pathway is a significant contributor to the acquisition of ciprofloxacin resistance in P. aeruginosa.
- Mutations in SR genes, like spoT, confer resistance and alter bacterial physiology.
- Understanding SR's role in resistance can inform future therapeutic strategies against P. aeruginosa infections.
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