Pharmacodynamics Of Linezolid-plus-fosfomycin Against Vancomycin-susceptible And-resistant Enterococci In Vitro And In Vivo Of A Galleria Mellonella Larval Infection Model

Objectives:To explore the activity of linezolid in combination with fosfomycin against vancomycin-susceptible and-resistant enterococci in vitro and the Galleria mellonella infection model,providing a theoretical basis for the treatment of vancomycin resistance Enterococcus.Methods: In this experiment,we used the checkerboard method and time-kill experiment to evaluate the effect of linezolid plus fosfomycin.The transmission electron microscope was used to observe the bacterial cell morphology of vancomycin-susceptible enterococcus faecium No.1 and vancomycin-resistant enterococcus faecium No.2 followed by each drug alone and in combination,which further elucidated the mechanism of action of antibiotic combination therapy.In vivo,the efficacy in vivo of the sensitive antibacterial drug in vitro was determined using Galleria mellonellla larvae,and then we tested the effect of linezolid in combination with fosfomycin in an in vivo survival assay using Galleria mellonellla larvae.Results: In the checkerboard method,linezolid combined with fosfomycin showed synergistic or additive effects in 5 of 6 strains(the FICI of these 5 strains ranged from 0.5 to 1.0),and no antagonistic effect was observed.Two clinical isolates(No.1,No.2)showed synergy.The SBPI > 2 for all strains manifested potential synergism.In the time-kill study,linezolid monotherapy showed bacteriostatic effect against all strains;Fosfomycin monotherapy showed good bacteria-killing activity in the first 8 h or 12 h,but regrowth appeared in all four strains and achieved the level of the control in two strains;While the combination of linezolid and fosfomycin at 1?MIC or 2?MIC led to higher degree of bacterial killing without regrowth for all isolates tested than each monotherapy;Fosfomycin resistance was noted by 24 h when used as monotherapy.MICs for all isolates were > 1024mg/L,indicating the emergence of fosfomycin resistant bacteria.The results of transmission electron microscopy showed that linezolid combined with fosfomycin damaged No.1 and No.2 more obviously than the linezolid or fosfomycin monotherapy.In the Galleria mellonella infeciton model of enterococci,the sensitive antibacterial drugs in vitro showed good antibacterial activity in vivo.A statistically significant larger level of survival rate was observed for No.1 and No.2 in the combination of linezolid and fosfomycin compared with linezolid monotherapy(P<0.05).fosfomycin showed excellent antibacterial efficacy against both No.1 and No.2,which was approximately equivalent to the linezolid plus fosfomycin(P>0.05).Conclusions:The checkerboard method and the time-kill curve experiment showed that the combination of linezolid and fosfomycin showed synergistic or additive effects on vancomycin-sensitive and-resistant enterococci in vitro,and no antagonistic effect was observed.Transmission electron microscopy showed that linezolid combined with fosfomycin damaged bacterial the bacterial cell morphology more obviously,further confirming the synergistic effect of the combination of the two drugs.The establishment of the Galleria mellonella infeciton model indicates that the activity in vivo is consistent with the sensitive activity in vitro.Compared with linezolid monotherapy,linezolid combined with fosfomycin significantly improved the survival rate of the larvae infected by vancomycin-susceptible and-resistant enterococci,which is consistent with the synergy effect observed in vitro.In conclusion,the combination of linezolid and fosfomycin is a potential therapeutic regimen for VRE infection.