Quorum Sensing Inhibitory Activities of Various Folk-Medicinal Plants and the Thyme-tetracycline Effect

Pseudomonas aeruginosa is an opportunistic, nosocomial pathogen for which antibiotic resistance and biofilm development is common. Quorum sensing communication is known to be a major controlling factor in virulence gene expression, biofilm development, antibiotic resistance factors, and specifically MexAB-OprM multi-drug efflux pump expression in P.aeruginosa. MexAB-OprM efflux pumps contribute to antibiotic resistance of tetracycline and other antibiotics in pseudomonads and other organisms. P.aeruginosa infections are problematic in cystic fibrosis and burn patients; it is also the number one causative agent of respiratory infections for intensive care unit patients. Present day antibiotics are losing the battle against these infections. In theory, quorum sensing inhibitors (QSI) reduce pathogencity of the organism; making it less virulent, thus allowing either the host immune system to clear the infection or use of a QSI in combination with an antibiotic to clear more persistent pathogens. For these reasons two alternative modes of treatment were explored in this study: quorum sensing inhibition by folk-medicinal plant extracts and an example of combination drug therapy, the ―thymetetracycline effect‖. Fifty folk-medicinal plant extracts were screened for potential anti-quorum sensing activity using two quorum sensing inhibition (QSI) reporter strains, Pseudomonas aeruginosa QSIS2 and Chromobacterium violaceum 12725. These were used to test specifically for C4-C6 and C12 HSL quorum sensing inhibition. Of the fifty plants tested, thirty plant families were represented. Eleven plant extracts (basil, chaparral, clove, cranberry, oregano, pomegranate, rosemary, sage, sassafras, thyme and witch hazel) showed C4 HSL quorum sensing inhibition as determined by both assays. Interestingly, five of the plants were from the Lamiaceae family. Thymus vulgaris (thyme), also from the Lamiaceae family, was chosen for further assessment. Previous research has shown that thyme extract can synergistically augment tetracycline activity against tetracycline-resistant Pseudomonas aeruginosa, creating the ―thyme-tetracycline effect.‖ Disc diffusion assay, thin layer chromatography (TLC), and TLC bioassay techniques were used to show that thymol is the active component in the thyme extract that augments tetracycline activity against resistant Pseudomonas. This study also showed that thymol is a potent C4 HSL quorum sensing inhibitor. The collective data suggests a potential mode of action for the thyme-tetracycline effect: thymol appears to prevent MexAB-OprM efflux pump gene expression. By blocking MexAB-OprM expression, tetracycline antibiotic accumulation can occur within the cell, thus allowing cellular damage. INDEX WORDS: Thymus vulgaris, Quorum sensing inhibition, Medicinal plants, Pseudomonas aeruginosa QUORUM SENSING INHIBITORY ACTIVITIES OF VARIOUS FOLK MEDICINAL PLANTS AND THE ELUCIDATION OF THE THYME-TETRACYCLINE EFFECT