Are essential oils a solution to combat antimicrobial resistance?

Abstract

Essential oils (EOs), derived from aromatic plants, have been widely studied and used as antimicrobials due to their broad-spectrum activity. This offers a potential resolution to the worldwide concern of antimicrobial resistance. However, little is known about the impact EOs will have on resistance emergence due to exposure in relation to resistance mechanisms. This study aimed to investigate resistance mechanisms and the potential induction of resistance against a selection of EOs. The research objectives encompassed four methods comprising of an initial antimicrobial screening of 23 EOs, followed by anti-quorum sensing (anti-QS) activity, biofilm inhibition, and resistance induction assays against five oils. The broth microdilution assay was carried out on 23 EOs, to screen the minimum inhibitory concentrations (MICs) against the ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter aerogenes) pathogens. This included reference, susceptible and resistant clinical strains. Based on the minimum inhibitory activity results, five EOs, Thymus vulgaris Willk. (0.92 mg/mL), Origanum vulgare L. (0.72 mg/mL), Carum carvi L. (1.62 mg/mL), Commiphora myrrha Engl. (3.04 mg/mL), and Matricaria recutita L. (3.36 mg/mL), representing a variety of inhibitory activity (noteworthy - poor) were selected for further investigation into resistance mechanisms. The anti-QS activity was evaluated, using Chromobacterium violaceum (ATCC 12472) as the biomonitor strain. The results revealed that all five EOs exhibited noteworthy (≥ 70.00%) quorum sensing (QS) inhibition at concentrations ranging from 0.02 - 1.00 mg/mL. Essential oils with noteworthy and moderate inhibitory activities had comparable minimum quorum sensing inhibitory concentrations (MQSICs). For instance, C. carvi and O. vulgare both exhibited a MQSIC ranging from 0.03 to 0.06 mg/mL; though, O. vulgare displayed a 3.01% higher inhibition of QS compared to C. carvi. For the biofilm inhibition assay, the crystal violet method was used to quantify the biofilm inhibition percentage. The five EOs were tested against S. aureus ATCC 6538. At a concentration of 0.05 mg/mL, O. vulgare and T. vulgaris inhibited biofilm formation most notably (≥ 70.00%). Matricaria recutita and C. myrrha inhibited more than 70.00% biofilm v formation starting at 2.00 mg/mL, while C. carvi only demonstrated notable inhibition (≥ 70.00%) at the highest concentration (4.00 mg/mL) tested. The resistance induction assay involved exposure of S. aureus ATCC 6538, to the five EOs, over a total of 20 passages. Origanum vulgare and T. vulgaris maintained their noteworthy minimum inhibitory activity over the 20 passages with minor inconsistencies in the MIC (SD ± 0.22 and ± 0.32 respectively). Carum carvi had the most variation in the MIC reported, however, no occurrences of resistance were detected. Commiphora myrrha and M. recutita both had a sudden declivity in their MIC at passage 16, with the MIC difference being more than two serial dilutions lower. There were changes in the growth kinetics of S. aureus as a result of exposure, which was observed in passages 16, 17, and 18. The exponential phase, when resistance is most likely to occur, shifted from 3 to 11 hrs, between passages 16 and 17. Staphylococcus aureus ATCC 6538 displayed heteroresistance patterns to the antibiotics after the exposure of the culture to EOs at certain passages. The exposure of S. aureus to O. vulgare resulted in the most variation in the MIC of the antibiotics, with T. vulgaris being the second. This study revealed that O. vulgare and T. vulgaris exhibited noteworthy activity in the minimum inhibition, anti-QS, and biofilm inhibition assays, within a concentration range of 0.03 - 1.00 mg/mL and 0.02 - 1.00 mg/mL respectively, highlighting these oils as promising antimicrobials to combat resistance. The induction of resistance assay gave key insight into the relationship between minimum inhibitory activity and the ability to detect resistance through repeated exposure. In its entirety, the study affirms the dissertation title by demonstrating that EOs can effectively contribute to combating AMR, contingent upon meticulous selection and a thorough understanding of the role each EO plays in various resistance mechanisms.