Amorfrutin-A, a membrane-targeting phytochemical active against multidrug-resistant Staphylococcus aureus: In vitro and in vivo activity and mechanism of action.

There is an urgent need to develop new antibiotics to combat multidrug-resistant (MDR) Staphylococcus aureus infections. Amorfrutins are plant-derived prenylated benzoic acids whose antimicrobial potential is largely unexplored. This study aimed to investigate the antibacterial properties of amorfrutins. To this end, the antibacterial activity of ten amorfrutins purified from Glycyrrhiza foetida (Fabaceae) was evaluated by determining their minimum inhibitory concentrations (MICs) against reference bacterial strains, as well as their cytotoxicity in human cell lines. The most promising compound was investigated to identify its target, mode of action (MoA), interactions with standard-of-care anti-staphylococcal antibiotics, and inhibition of biofilm and persister cells. The MoA was corroborated by molecular dynamics simulations and in vitro assays using artificial membranes and cellular models. Toxicity and antibacterial efficacy were also assessed in an insect infection model. Among the tested amorfrutins, amorfrutin-A (AMF-A) exhibited the strongest activity against S. aureus (MIC=4-16 mg/L) without causing detectable cytotoxicity. AMF-A rapidly disrupted S. aureus membranes, eradicated biofilm and persister cells, and displayed an undetectable frequency of resistance development. AMF-A did not antagonize standard-of-care anti-staphylococcal antibiotics and showed synergistic effects with daptomycin, piperacillin, and azithromycin. AMF-A showed no in vivo toxicity and protected Galleria mellonella larvae from lethal S. aureus infection. We conclude that AMF-A is a promising membranolytic phytochemical with potential for treatment of MDR S. aureus infections.