In Vitro and In Vivo Synergistic Antibacterial Functions of the Recombinant Ib-AMP4 and E50-52 Antimicrobial Peptides against Multidrug-Resistant Acinetobacter baumannii

Background: Most Acinetobacter baumannii (A. baumannii) species have become resistant to all common antibiotics. Many antimicrobial peptides (AMPs) have been identified with efficient functions in infection management. E50-52 (UniProtKB: P85148) and Ib-AMP4 (UniProtKB: O24006) AMPs have shown marked antibacterial functions. Objectives: This investigation was designed to produce E50-52 and Ib-AMP4 AMPs through recombinant protein production. Subsequently, the synergistic antimicrobial functions of these two peptides were assessed under in vitro and in vivo circumstances on multidrug-resistant A. baumannii to investigate the antimicrobial effects of Ib-AMP4 and E50-52 AMPs on drug-resistant Acinetobacter. Methods: The gene sequence of E50-52 and Ib-AMP4 AMPs were codon optimized and separately inserted into the pET-32α vector. The recombinant structures were expressed in host bacteria. The antibacterial functions of the individual and combined application of the purified refolded E50-52 and Ib-AMP4 AMPs against multidrug-resistant A. baumannii were evaluated through the time-kill, minimum inhibitory concentration (MIC), growth kinetic assays, and in vivo (mouse body) systemic infection. Results: The minimum concentrations of the produced refolded E50-52 and Ib-AMP4 AMPs against A. baumannii were 0.325 and 0.0625 mg/mL, respectively. Moreover, the checkerboard procedure confirmed the synergic effects of the produced AMPs. The use of E50-52 and Ib-AMP4 AMPs in combination resulted in an over five times reduction in log10 CFU/mL of alive cells during the first 240-min exposure. The antibacterial efficiency of the produced AMPs was confirmed by growth kinetic assays, scanning electron microscopy (SEM) results, and in vivo evaluation tests. The in vivo assay on rats confirmed the significant antibacterial functions of the produced recombinant proteins on A. baumannii systemic infection. Conclusions: The results proved the considerable synergistic antibacterial functions of the produced recombinant Ib-AMP4 and E50-52 AMPs to treat A. baumannii systemic infection effectively.