Gramicidin D enhances the antibacterial activity of fluoride

Abstract

Fluoride is a toxic anion found in many natural environments. One of the major bacterial defenses against fluoride is the cell envelope, which limits passage of the membrane-impermeant fluoride anion. Accordingly, compounds that enhance the permeability of bacterial membranes to fluoride should also enhance fluoride toxicity. In this study, we demonstrate that the pore-forming antibiotic gramicidin D increases fluoride uptake in Bacillus subtilis and that the antibacterial activity of this compound is potentiated by fluoride. Polymyxin B, another membrane-targeting antibiotic with a different mechanism of action, shows no such improvement. These results, along with previous findings, indicate that certain compounds that destabilize bacterial cell envelopes can enhance the toxicity of fluoride.

Keywords: Antibiotic; Bacillus subtilis; Fluoride reporter; Polymyxin B; Riboswitch.

Figure 1

Synergistic antibacterial function of fluoride and gramicidin D. (A) Gramicidin D increases intracellular fluoride levels in B. subtilis. Reporter gene expression values were normalized to account for culture growth (see Supplemental Material). Error bars are the standard deviation of three replicates. (B) Gramicidin D enhances the antibacterial activity of fluoride. Data points represent the growth of B. subtilis following 16 hours of incubation in the condition indicated for each experiment. Error bars represent the standard deviation of three measurements and the data is fit to a sigmoidal curve. (C) Fluoride enhances the antibacterial activity of gramicidin D. Culture growth is depicted as trend lines, which represent the average optical density of three replicates that were measured every fifteen minutes.

Figure 2

No enhancement of fluoride uptake is observed upon addition of polymyxin B to E. coli. Data points are the average of three replicates and error bars represent the standard deviation, which is smaller than the data point symbols. No growth was observed under the experimental conditions with 2.5 μg mL⁻¹ polymyxin B (data not shown).

Figure 3

Synergistic effect of fluoride and gramicidin D in B. halodurans. Data presented are the average of three replicates and error bars represent standard deviation.