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Updated: Jun 24, 2025
Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
Published on: March 16, 2011
In vitro induction and selection of fluoroquinolone-resistant mutants in Elizabethkingia anophelis
Ching-Chi Lee1, Chung-Hsu Lai2, Shang-Yi Lin3
1Clinical Medical Research Center, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 704, Taiwan; Departments of Internal Medicine, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 704, Taiwan.
Abstract:
For 29 parent strains, recognized by pulsed-field gel electrophoresis, the MICs multiplied significantly in the ciprofloxacin group than levofloxacin group, following the first and third induction cycle. Ser83Arg in GyrA was the most common site of mutations. No mutation in ParC nor ParE was identified in the selected mutants.
Insights
Ciprofloxacin significantly increased minimum inhibitory concentrations (MICs) more than levofloxacin in bacterial strains after induction cycles. The most frequent mutation occurred in GyrA, with no identified mutations in ParC or ParE.
Area of Science:
- Microbiology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Fluoroquinolones, including ciprofloxacin and levofloxacin, are critical antibiotics.
- Understanding the mechanisms of bacterial resistance to these drugs is essential for public health.
Purpose of the Study:
- To compare the effects of ciprofloxacin and levofloxacin on bacterial susceptibility.
- To identify genetic mutations associated with fluoroquinolone resistance.
Main Methods:
- Bacterial strains were exposed to increasing concentrations of ciprofloxacin or levofloxacin over multiple induction cycles.
- Minimum inhibitory concentrations (MICs) were determined.
- Genetic analysis, including pulsed-field gel electrophoresis and sequencing, was performed to identify mutations in key genes.
Main Results:
- A significant increase in MICs was observed in the ciprofloxacin group compared to the levofloxacin group after the first and third induction cycles.
- The Ser83Arg substitution in the GyrA gene was the most prevalent mutation identified in resistant mutants.
- No mutations were detected in the ParC or ParE genes in the selected mutants.
Conclusions:
- Ciprofloxacin induces higher levels of resistance compared to levofloxacin in the studied bacterial strains.
- Mutations in GyrA are a primary mechanism for fluoroquinolone resistance in these strains.
- Further research is needed to explore resistance mechanisms involving ParC and ParE.
