Frog Skin Peptides Hylin-a1, AR-23, and RV-23: Promising Tools Against Carbapenem-Resistant Escherichia coli and Klebsiella pneumoniae Infections

Background/Objectives. One of the pressing challenges in global public health is the rise in infections caused by carbapenem-resistant Enterobacteriaceae. Growing bacterial drug resistance, coupled with the slow development of new antibiotics, highlights the critical need to explore and develop new broad-spectrum antimicrobial agents able to inhibit bacterial growth efficiently. In recent years, antimicrobial peptides (AMPs) have gained significant attention as a promising alternative to conventional drugs, owing to their antimicrobial potency, low toxicity, and reduced propensity for fostering resistance. Our research aims to investigate the antibacterial ability of three amphibian AMPs, namely Hylin-a1, AR-23, and RV-23, against both antibiotic-sensitive and carbapenem-resistant strains of Escherichia coli and Klebsiella pneumoniae. Methods. A 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT) was performed to identify non-cytotoxic concentrations of peptides. A microdilution assay evaluated the antibacterial effect, determining the peptides’ minimum inhibitory concentration (MIC). In addition, the checkerboard test analyzed the compounds’ synergistic effect with meropenem. Results. We demonstrated that peptides with low toxicity profile and resistance to proteolytic activity exhibited strong antibacterial activity, with MIC ranging from 6.25 to 25 μM. The antibiofilm mechanism of action of peptides was also investigated, suggesting that they had a crucial role during the biofilm formation step by inhibiting it. Finally, we highlighted the synergistic effects of peptides with meropenem. Conclusions. Our study identifies Hylin-a1, AR-23, and RV-23 as promising candidates against Gram-negative bacterial infections with a favorable therapeutic profile. This effect could be related to their great flexibility, as evidenced by circular dichroism data, confirming that the peptides could assume an α-helical conformation interacting with bacterial membranes.