Assessing the Emergence of Resistance in vitro and Invivo: Linezolid Combined with Fosfomycin Against Fosfomycin-Sensitive and Resistant Enterococcus

PURPOSE: We aimed to evaluate the synergistic effect of linezolid and fosfomycin on fosfomycin-sensitive and -resistant Enterococcus clinical isolates in vitro and in vivo and whether the emergence of fosfomycin resistance in Enterococcus is associated with changes in strain virulence, from the perspective of fitness cost. METHODS: The synergistic effect of linezolid and fosfomycin was studied via in vitro checkerboard and static time-kill assays, as well as based on the in vivo survival rate and hemolymph load of a Galleria mellonella infection model. Fosfomycin resistance was induced via a stepwise increase in concentration. Changes in the virulence of the strains after drug resistance were investigated using the G. mellonella infection model and reverse transcription quantitative polymerase chain reaction (RT-qPCR). In vitro and in vivo growth curves and competitive experiments were used to study the fitness cost of the strain. Finally, a static time-kill assay was performed to explore the effect of the combined medication. RESULTS: In vitro and in vivo data showed that linezolid combined with fosfomycin had a good synergistic effect on Enterococcus treatment. The G. mellonella infection model and RT-qPCR data showed that the virulence of the resistant strains was weakened to varying degrees. A survival curve and competition experimental data showed that this was related to the fitness cost of strains while acquiring resistance and negatively impacted linezolid treatment; however, the combination still showed a good synergistic effect in drug-resistant strains. CONCLUSION: Linezolid combined with fosfomycin had a synergistic effect on both fosfomycin-sensitive and -resistant Enterococcus strains. Strains incur fitness costs as they develop drug resistance, which leads to a decrease in virulence. There is an interaction between fitness cost, virulence, and drug resistance, which indirectly affects drug treatment.