Elucidating the mechanisms of colistin resistance and the efficacy of combination therapy in MDR/XDR/PDR Acinetobacter baumannii

Abstract

Background: The rapid dissemination of carbapenem resistance in Gram-negative bacteria, particularly in Acinetobacter baumannii, has gained focus on the “old” drug colistin. Neverthless, the emergence of colistin resistance, last line of defense, causes a serious global threat. Therefore, antimicrobial treatment for these infections remains an important challenge. In this study, we aimed (1) to understand the involvement of the Two-Component system pmrAB and lipid biosynthesis lpx genes in colistin resistance and their corresponding relative expression levels and (2) implement suitable combination therapy options. Methods and materials: Eighty clinical isolates of A. baumannii were identified phenotypically and confirmed by PCR of GyrA gene. Isolates were subjected to antimicrobial susceptibility testing by BMD. The role of pmrAB and lpx genes on colistin resistance was determined by qRT-PCR. Assessment of the effect of combination therapy was done by the checkerboard analysis, Time kill experiments and the neutropenic pneumonia-induced mouse model. The effect of the electrostatic interaction of colistin action was seen by the Scanning Electron Microscopy. Results: Data have shown that 13 of the 80 (16%) isolates were resistant to colistin. The efficacy of colistin in combination with tigecycline, fosfomycin, zerbaxa, tazocin, teicoplanin, amikacin, levofloxacin, rifampicin and meropenem was assessed against colistin resistant A. baumannii isolates. Checkerboard and Time-kill assays demonstrated a synergestic effect between colistin and rifampicin and between colistin and teicoplanin, with bactericidal activity occuring prior to 4 h of incubation. A preliminary neutropenic pneumonia-induced mouse model, demonstrated improved survival and pathogen clearance from the blood and lung tissue cultures in the rifampicin combination. Up-regulation of pmrAB genes was recorded with a down-regulation of the lpx genes in vitro, nonethless this was not the case in in vivo studies. In addition, qRT-PCR did not show any significant differences between resistant and susceptible isolates upon examining the levels of expression of lpx and pmrAB genes. Conclusion: In conclusion, further understanding of the various molecular mechanisms of colistin resistance in A. baumannii is a must. Our current efforts focused on correlating the results of combination therapy in relation to the different resistance mechanisms to colistin, paving the path for potential treatment options against XDR and PDR A. baumannii infections.