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A Fluorescence-based Method to Study Bacterial Gene Regulation in Infected Tissues
Published on: February 19, 2019
Heterologous expression of the enterococcal vanA operon in methicillin-resistant Staphylococcus aureus
Bruno Périchon1, Patrice Courvalin
1Unité des Agents Antibactériens, Institut Pasteur, 25, rue du Docteur Roux, 75724 Paris Cedex 15, France.
Abstract:
Two methicillin- and vancomycin-resistant Staphylococcus aureus strains, MI-VRSA and PA-VRSA, and Enterococcus faecalis DMC83006B, considered to be the potential donor of glycopeptide resistance to MI-VRSA, were studied. MI-VRSA is highly resistant to both glycopeptides, whereas PA-VRSA displays low-level resistance to vancomycin and reduced susceptibility to teicoplanin. We have analyzed the expression of the vanA operon in the three clinical isolates. Determination of the relative amounts of late peptidoglycan precursors and quantification of the d,d-peptidase activities, in the absence or after induction by glycopeptides, revealed that the resistance genes were expressed at similarly high levels in the three strains. Glycopeptide resistance stability in the three strains was studied by replica plating. Resistance was lost at high frequency, ca. 50%, after overnight growth of PA-VRSA in the absence of antibiotics, whereas it was fully stable in MI-VRSA and E. faecalis DMC83006B. Induction of resistance by vancomycin was significantly delayed in PA-VRSA relative to MI-VRSA. Low-level glycopeptide resistance of S. aureus PA-VRSA is thus likely due to instability of the genetic element, plasmid or transposon, carrying the vanA operon associated with a longer lag phase before growth resumes after induction by vancomycin.
Insights
Investigating vancomycin-resistant Staphylococcus aureus (VRSA) strains revealed that low-level resistance in PA-VRSA is linked to unstable vanA operon expression and delayed induction. High-level resistance in MI-VRSA and Enterococcus faecalis was stable.
Area of Science:
- Microbiology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Methicillin- and vancomycin-resistant Staphylococcus aureus (VRSA) pose significant clinical challenges.
- Enterococcus faecalis is a known reservoir for glycopeptide resistance genes.
- Understanding the mechanisms of vanA operon expression and resistance stability is crucial.
Purpose of the Study:
- To analyze vanA operon expression in two VRSA strains (MI-VRSA and PA-VRSA) and an Enterococcus faecalis strain.
- To investigate the stability of glycopeptide resistance in these clinical isolates.
- To elucidate the factors contributing to low-level vancomycin resistance in S. aureus PA-VRSA.
Main Methods:
- Analysis of vanA operon expression in clinical isolates.
- Quantification of peptidoglycan precursors and d,d-peptidase activity.
- Assessment of glycopeptide resistance stability using replica plating.
- Evaluation of vancomycin induction lag phase.
Main Results:
- VanA operon genes were expressed at high levels in all three strains.
- Glycopeptide resistance was unstable in PA-VRSA (approx. 50% loss) but stable in MI-VRSA and E. faecalis.
- Vancomycin induction of resistance was significantly delayed in PA-VRSA compared to MI-VRSA.
Conclusions:
- Low-level vancomycin resistance in S. aureus PA-VRSA is attributed to the instability of the vanA operon-carrying genetic element.
- A longer lag phase before growth resumption after vancomycin induction contributes to reduced resistance in PA-VRSA.
- Resistance stability and induction kinetics are key factors in glycopeptide resistance expression in staphylococci and enterococci.
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