Thymol as a Novel Plant-Derived Antibacterial Agent for Suppressing Xanthomonas citri pv. malvacearum in Cotton

Xanthomonas citri pv. malvacearum (Xcm) associated with bacterial blight disease is a significant and widespread pathogen affecting cotton worldwide. The excessive use of harmful chemicals to control plant pathogens has exerted a negative impact on environmental safety. Thymol is a monoterpene phenol present in the essential oils of plants belonging to Lamiaceae family. In this study the antimicrobial activity of thymol was evaluated against Xcm. The minimum inhibitory concentration (MIC) and 99.9% bactericidal concentration (MBC) of thymol against Xcm were 2 and 4 mg/mL, respectively. The effect of MIC and MBC of thymol against Xcm was assessed on the Luria-Bertani medium. The effect of thymol on intercellular ATP levels, membrane potential, and motility in Xcm was assessed using fluorescence spectrometry for membrane potential and firefly luciferase-based assay for ATP levels. Thymol ruptured the cellular membrane of Xcm, resulting in decreased intracellular ATP concentrations, intracellular leakage of genetic material, and changes in membrane potential. Scanning electron microscopy images supported the impact of thymol on the cell membrane of Xcm. Moreover, thymol inhibited the swimming motility and biofilm formation of Xcm at concentrations equal to or above the MIC and MBC. In contrast, sub-MIC concentrations of thymol had little to no impact on the virulence of Xcm. In conclusion, thymol demonstrated the potential as a strong bactericidal compound against Xcm.