Study On The Antibacterial Effect Of Imipenem Combined With Fosfomycin On Multidrug-resistant Acinetobacter Baumannii In Vivo And In Vitro

Objective To explore the in vitro and in vivo antibacterial methods of imipenem-based drugs against clinically isolated multidrug-resistant Acinetobacter baumannii（MDR-AB）.Method The minimal inhibitory concentration（MIC）and combined MIC of12 clinical isolates of MDR-AB against imipenem（IMP）,fosfomycin（FOS）,amikacin（AMK）,tegacycline（TGC）and polymyxin B（PB）were determined by broth dilution method and chessboard method,and the fractional inhibitory concentration indices（FICI）was calculated.Evaluation of germicidal efficacy of imipenem combined with fosfomycin by time-killing curves.Eight drug resistance genes（blaOXA-23,blaOXA-24,blaOXA-51,blaOXA-58,blaIMP,blaVIM,blaADC,blaKPC）and five virulence genes（ompA,csuA,csuE,pga A,abaI）were detected by polymerase chain reaction,and the biofilm of the tested strains was semi-quantitatively stained with crystal violet.The effect of IMP combined with FOS on the biofilm of the strain was observed by laser confocal microscope.The effect of IMP combined with FOS on the expression of bacterial reactive oxygen species ROS was detected by flow cytometry.The infection model of Galleria mellonella was established,and the survival rate of Galleria mellonella was recorded when imipenem and fosfomycin were used alone or in combination.Results IMP combined with FOS had synergistic effect on 8 strains of MDR-AB.Imipenem combined with fosfomycin reduced the colony number of strains AB6624 and AB0153 by more than 2log10 CFU·mL^-1at 8h and 4h respectively,and the colony number of strains AB7182 at 8h was more than3log10 CFU·mL^-1.12 strains carried drug resistance genes blaOXA-23,blaOXA-51 and blaADC and five virulence genes ompA,csuA,csuE,pga A and abaI,and 8 strains carried drug resistance gene blaOXA-58.10 strains carried drug resistance gene blaVIM.2 strains carried drug resistance gene blaKPC.All of the MDR-AB tested has strong biofilm forming ability.IMP combined with FOS treatment inhibited the biofilm formation of strains AB6624 and AB0153 and increased the proportion of dead bacteria in the biofilm.IMP combined with FOS could increase the intracellular ROS level of strain AB0153,the difference was statistically significant（P<0.05）.The LD80 of strain AB6624 was 1.0×10⁵CFU·mL^-1,and the LD80 of strain AB0153 was 1.0×10⁶CFU·mL^-1.The survival rate of Galleria mellonella treated with IMP combined with FOS was improved compared with the control group.Conclusion Imipenem combined with fosfomycin has a synergistic effect on MDR-AB and may play a role by inhibiting the formation of biofilm and increasing the level of intracellular ROS of bacteria.