Fig. 3. NAMs are potent agents against the formation of antibiotic tolerant, persisters (AT/P) cells and biofilm.

a AT/P cells formation in PA14 and isogenic ∆mvfR in the presence or absence of the indicated compounds. Cells were grown in the presence of 10 μg mL⁻¹ meropenem with or without 10 µM of the compounds for 24 h. Values were normalized to the cells grown for 4 h in the absence of antibiotics and compound. The PA14 cells grown with antibiotics and vehicle (DMSO) were considered control, and the percentage values were calculated compared to control. b Initiation of biofilm formation of PA14 and ∆mvfR cells with or without compound. Biofilm was grown in the 96 well microtiter plate at 37 °C for 24 h containing M63 minimal media in the presence of 10 µM of the compounds or vehicle. After 24 h, the wells were washed to remove planktonic cells, and the biofilm was stained with 0.1% crystal violet. The stained biofilm was washed and solubilized in ethanol: acetone (80:20). OD was measured at 570 nm. The biofilm grown with the vehicle was considered as a control. The percentage value was calculated in comparison to the PA14 control. a, b Data represent at least n = 3 biological replicates, each dot on the graph represents one replicate, and the number of biological replicates for each compound is depicted in the graph. The error bars denote ±SEM. One-way ANOVA followed by Tukey post-test was applied. ∗, ∗∗ and ∗∗∗ indicate significant differences from the control at P < 0.05, P < 0.01, and P < 0.001, respectively.