Thyme essential oil potentials as a bactericidal and biofilm-preventive agent against prevalent bacterial pathogens

Abstract

Antimicrobial resistance represents a significant global issue that requires the investigation of innovative approaches for infection management. In pursuit of alternative natural antimicrobials, nine plant essential oils were evaluated for their antibacterial properties against nine common bacterial pathogens. Among the tested essential oils, thyme essential oil demonstrated the highest antibacterial activity against all tested bacterial species, Thyme essential oil exhibited inhibition zones ranging from 17.3 to 51 mm with relative minimum inhibitory concentrations ranging from 99.2 to 450 µg/ml, implying the bactericidal effect. The ultrastructural changes in bacterial cells treated with thyme essential oil were visualized using transmission electron microscope. Thyme essential oil exhibited a potent inhibitory effect toward the biofilm formations for all the tested pathogenic strains. GC/MS analysis was used to determine the thyme essential oil composition. The major components of thyme essential oil were thymol (28.29%), o-cymene (18.31%), ç-terpinene (8.51%), eucalyptol (5%), linalool (2.86%), borneol (2.17%), á-myrcene (1.55%), à-pinene (1.52%) and camphene (1%). Molecular docking analysis demonstrated that the constituents present in the thyme essential oil had high binding affinity for ECF, FimH, LasR, PrfA and RhlA proteins, which were found to be associated with improved anti-biofilm efficacy. Furthermore, treatment with thyme essential oil led to the downregulation of essential genes associated with virulence and biofilm formation in the tested pathogens. These findings suggest that thyme essential oil has promising potential as an antibacterial and a biofilm inhibitory agent to combat bacterial infections in food and pharmaceutical industries.

Keywords: Antibacterial activity; Biofilm; Chemical composition; Gene expression; Molecular docking; Pathogenic bacteria; Thyme essential oil.

Fig. 1

TEM micrographs of control and essential oil treated Gram-positive bacteria (B. cereus, L. monocytogenes, S. aureus and S. aureus MRSA). (A) Untreated B. cereus (Direct Mag.: 30,000×). (B) Treated B. cereus (Direct Mag.: 40,000×). (C) Untreated L. monocytogenes (Direct Mag.: 30,000×). (D) Treated L. monocytogenes (Direct Mag.: 15,000 and 40,000×). (E) Untreated S. aureus (Direct Mag.: 30,000×). (F) Treated S. aureus (Direct Mag.: 25,000 and 40,000×). (G) Untreated S. aureus MRSA (Direct Mag.: 15,000 and 30,000×). (H) Treated S. aureus MRSA (Direct Mag.: 25,000 and 30,000×).

Fig. 2

TEM micrographs of control and essential oil treated Gram-negative bacteria (E. coli (STEC), E. coli O157:H7, S. typhimurium and P. aeruginosa). (A) Untreated E. coli (STEC) (Direct Mag.: 10,000 and 40,000×). (B) Treated E. coli (STEC) (Direct Mag.: 30,000 and 40,000×). (C) Untreated E. coli O157:H7 (Direct Mag.: 12,000 and 15,000×). (D) Treated E. coli O157:H7 (Direct Mag.: 20,000×). (E) Untreated S. typhimurium (Direct Mag.: 20,000 and 60,000×). (F) Treated S. typhimurium (Direct Mag.: 15,000 and 25,000×). (G) Untreated P. aeruginosa (Direct Mag.: 15,000 and 30,000×). (H) Treated P. aeruginosa (Direct Mag.: 20,000 and 25,000×).

Fig. 3

The impact of different concentrations of thyme essential oil on the inhibition of biofilm formation by pathogenic bacteria. (a) S. aureus, (b) S. aureus MRSA, (c) B. cereus, (d) E. coli (STEC), (e) L. monocytogenes, (f) S. typhimurium, (g) E. coli O157:H7, (h) P. aeruginosa. Data represents the mean of six replicates. Error bars represent standard deviation.

Fig. 4

Chromatographic profiles of the thyme essential oil.

Fig. 5

Molecular docking analysis. 1a. Interaction of thymol against RhlA active site in P. aeruginosa. 1b. Interaction of o-Cymene against RhlA active site in P. aeruginosa. 1c. Interaction of thymol against LasR active site in P. aeruginosa. 2a. Interaction of thymol against FimH active site in E. coli. 2b. Interaction of o-Cymene against FimH active site in E. coli. 3a. Interaction of thymol against ECF active site in S. aureus. 3b. Interaction of o-Cymene against ECF active site in S. aureus. 4a. Interaction of thymol against PrfA active site in L. monocytogenes. 4b. Interaction of o-Cymene against PrfA active site in L. monocytogenes.

Fig. 6

Expression analysis of the virulence and biofilm related genes in E. coli O157:H7 and S. aureus under control and thyme essential oil treatment conditions. Data represents the mean of three independent replicates. Error bars represent standard deviation.