Exposure of Staphylococcus aureus to Subinhibitory Concentrations of β-Lactam Antibiotics Induces Heterogeneous Vancomycin-Intermediate Staphylococcus aureus

Abstract

h Glycopeptides are known to select for heterogeneous vancomycin-intermediate Staphylococcus aureus (h-VISA) from susceptible strains. In certain clinical situations, h-VISA strains have been isolated from patients without previous exposure to glycopeptides, such as cystic fibrosis patients, who frequently receive repeated treatments with beta-lactam antibiotics. Our objective was to determine whether prolonged exposure to beta-lactam antibiotics can induce h-VISA. We exposed 3 clinical vancomycin-susceptible methicillin-resistant Staphylococcus aureus (MRSA) strains to ceftazidime, ceftriaxone, imipenem, and vancomycin (as a control) at subinhibitory concentrations for 18 days in vitro. Population analyses showed progressive increases in vancomycin resistance; seven of the 12 derived strains obtained after induction were classified as h-VISA according to the following criteria: area under the curve (AUC) on day 18/AUC of Mu3 of >90% and/or growth on brain heart infusion (BHI) agar with 4 mg/liter vancomycin. The derived isolates had thickened cell walls proportional to the level of glycopeptide resistance. Genes known to be associated with glycopeptide resistance (vraSR, yvqF, SA1703, graRS, walKR, and rpoB) were PCR sequenced; no de novo mutations were observed upon beta-lactam exposure. To determine whether trfA, a gene encoding a glycopeptide resistance factor, was essential in the selection of h-VISA upon beta-lactam pressure, a trfA-knockout strain was generated by allelic replacement. Indeed, beta-lactam exposure of this mutated strain showed no capacity to induce vancomycin resistance. In conclusion, these results showed that beta-lactam antibiotics at subinhibitory concentrations can induce intermediate vancomycin resistance in vitro. This induction required an intact trfA locus. Our results suggest that prior use of beta-lactam antibiotics can compromise vancomycin efficacy in the treatment of MRSA infections.

Stepwise Decrease in Daptomycin Susceptibility in Clinical Staphylococcus aureus Isolates Associated with an Initial Mutation in rpoB and a Compensatory Inactivation of the clpX Gene

“…Spx controls gene expression by interacting directly with the ␣ subunit of the RNA polymerase (45). The Spx-controlled trfA gene was recently shown to be required for the ability of subinhibitory concentrations of ␤-lactam antibiotics to induce the hVISA phenotype, raising the intriguing possibility that the elevated Spx level may contribute to the decreased susceptibility to daptomycin, vancomycin, and ␤-lactams (46,47). Unfortunately, attempts to examine this hypothesis by introducing an spx mutation into single mutants were unsuccessful.…”

Multifaceted Interfaces of Bacterial Competition

“…However, many of these metabolically expensive products operate between inactive and inhibitory concentrations and may act as chemical cues for competitors (131). Exposure to subinhibitory antibiotic concentrations can induce resistant states (132)(133)(134), select for resistant competitors (135), stimulate biofilm formation (136)(137)(138), and stimulate motility (139). The activation of a resistant state allows a competitor unrestricted access into a previously protected niche.…”

Regulation of virulence and antibiotic resistance in Gram-positive microbes in response to cell wall-active antibiotics

“…YirB is induced by the CssRS TCS system in response to CWA antibiotics (52), pointing toward interaction of the Spx pathway with TCS sensory systems. Further, TrfA, a factor contributing to glycopeptide and oxacillin resistance, is induced by Spx in response to CWA antibiotics (55)(56)(57). TrfA imparts both specificity and timing to protease-mediated degradation of antitoxins and other factors implicated in the response to antibiotic stress (58).…”