Mechanism of Action and Biofilm Inhibitory Activity of Lupinifolin Against Multidrug-Resistant Enterococcal Clinical Isolates.

The treatment of enterococcal infections is becoming more difficult because of multidrug resistance (MDR). Lupinifolin, a prenylated flavonoid isolated from Benth., showed a potent antimicrobial activity against enterococci. The aim of this study was to investigate antibacterial activity and action of lupinifolin against MDR enterococcal clinical isolates. Antibacterial properties of lupinifolin against 21 MDR isolates were assessed using broth microdilution method and time-kill assay. To study mode of action of lupinifolin on the isolates, propidium iodide intensity, salt tolerance assay, and electron microscopic analyses were performed. Antibiofilm formation activity of lupinifolin was conducted using crytal violet assay. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration values of lupinifolin against the isolates ranged between 0.5 and 2.0 μg/mL and between 2 and 16 μg/mL, respectively. Lupinifolin at 2MIC and 4MIC inhibited the bacterial growth >2 log colony-forming units (CFU)/mL at 2 hr incubation by time-kill analysis. The compound increased membrane permeability and caused loss of salt tolerance. SEM and TEM micrographs revealed pronounced morphological and ultrastructural changes in the treated bacteria. Crystal violet staining showed the antibiofilm-producing activity of lupinifolin against four MDR enterococci. This study suggested that lupinifolin is an essential antimicrobial agent that could be useful for the treatment of MDR enterococcal infections.