The Role of Antimicrobial Peptide Buforin2 in the Activity of AdeIJK Efflux Pump among Carbapenem-ResistantAcinetobacter baumannii Clinical Isolates

Abstract

Acinetobacterbaumannii is a multidrug-resistant (MDR) nosocomial pathogen that can cause severe infections. The efflux pumps especially the adeIJK system they developed are particularly important with respect. In this paper, we explore the impact of the antibacterial peptide Buforin2 on the activity of adeIJK efflux pump in carbapenem-resistant clinical isolates of A. baumannii. In this study, a total of 150 different clinical specimens (burns, urine, blood and sputum) were collected from two hospitals in Baghdad. Identification of A. baumannii isolates was performed using CHROMagar Acinetobacter medium, biochemical tests, and VITEK 2 system. Antibiotic susceptibility testing was carried out by the disc diffusion method. The role of efflux pumps was assessed by measuringthe minimum inhibitory concentrations (MICs) of antimicrobial agents (Meropenem, Amikacin, Ciprofloxacin, and Tigecycline) and Buforin2 in the presence and absence ofthe efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP).  Quantitative RT-qPCR was usedto evaluate the expression levels of efflux pump genes (adeJ and adeK). Among 150 clinical specimens, 65 (43.3%) were identified as A. baumannii, with the highest prevalence in burn infections (47.7%). Antibiotic susceptibility testing revealed high resistance rates, with 64.6% of isolates classified as MDR.Carbonyl cyanide 3-chlorophenylhydrazone significantly reduced MICs of Meropenem (8–64 fold) and Amikacin (8–32 fold), while it had a minimal effect on Tigecycline (1–2 fold) and ciprofloxacin (4-8 fold). Buforin2 exhibited a (2–4-fold) decrease in MICs with CCCP, suggesting a limited role of efflux pumps in its activity. Gene expression analysis showed significant upregulation of adeJ and adeK in response to Meropenem (4.62 and 3.51 fold, respectively), whereas Buforin2 led to downregulation of adeJ (0.41 fold) and a minimal change in adeK (1.24 fold). The adeIJK efflux pump contributes to antibiotic resistance in A. baumannii, particularly against carbapenems and aminoglycosides. Buforin2 exhibited limited susceptibility to efflux pump inhibition, suggesting a different mechanism of action. These findings highlight the potential of antimicrobial peptides as alternative therapeutics against MDR A. baumannii infections.