Sub-minimum inhibitory concentrations of fosfomycin enhance Staphylococcus aureus virulence through the agr-PSM pathway.

OBJECTIVE: Fosfomycin is increasingly used in combination therapies for multidrug-resistant bacterial infections. This study aimed to investigate the effect of sub-minimum inhibitory concentrations (sub-MICs) of fosfomycin on the virulence of Staphylococcus aureus to optimize therapeutic strategies.

METHODS: Hemolytic activity was assessed using disk diffusion and E-test assays on blood agar plates. The expression of virulence genes was quantified using reverse transcription polymerase chain reaction. Protein profiles were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The involvement of the accessory gene regulator (agr), α-hemolysin (hla), and phenol-soluble modulins (psm) was validated using mutant and complementation strains.

RESULTS: Sub-MICs of fosfomycin significantly enhanced hemolysis in both methicillin-sensitive and methicillin-resistant S. aureus (MSSA and MRSA) strains. MALDI-TOF MS analysis revealed a marked increase in the production of phenol-soluble modulins (PSMs) in S. aureus harvested from fosfomycin sub-inhibitory zones. Genetic analyses confirmed that sub-MICs of fosfomycin induced hemolysis was abolished in the agr or psm mutants and restored upon psm complementation. Moreover, sub-MICs of other agents targeting cell wall synthesis, including β-lactams (ceftazidime) and glycopeptides (vancomycin), significantly enhanced S. aureus hemolysis.

CONCLUSION: Sub-MICs of fosfomycin enhance S. aureus hemolysis by activating the agr-PSM pathway. These findings highlight the potential clinical risks associated with subtherapeutic fosfomycin exposure, which may exacerbate staphylococcal virulence during infection.