Selected Essential Oils Modulate Ethidium Bromide Accumulation and Exhibit Antibacterial Activity Against Multidrug-Resistant Escherichia coli and Klebsiella pneumoniae

The increasing prevalence of multidrug-resistant (MDR) Gram-negative bacteria has stimulated the search for plant-derived antimicrobials with alternative modes of action. This study investigated whether chemically distinct essential oils (EOs) differ in their antibacterial effects and their ability to modulate bacterial efflux-associated processes in clinically relevant MDR pathogens. The chemical compositions of tea tree (Melaleuca alternifolia), lemon (Citrus limon), Scots pine (Pinus sylvestris), and Eucalyptus radiata EOs, as well as their blend, were characterized by gas chromatography–mass spectrometry (GC-MS). Terpinen-4-ol, limonene, α-pinene, and eucalyptol were identified as major constituents. Antibacterial activity against Escherichia coli and Klebsiella pneumoniae strains with different resistance profiles was evaluated by broth microdilution, while effects on efflux-associated processes were assessed using an ethidium bromide accumulation assay. The EOs showed strain-dependent antibacterial effects, with E. coli being more susceptible than K. pneumoniae. E. radiata showed the lowest MIC against E. coli 130063 (0.312 mg/mL), whereas no EO inhibited the K. pneumoniae strains at concentrations below 5 mg/mL. The highest RFI values were observed for lemon oil against K. pneumoniae 33443 (1.41) and the EO blend against E. coli 128673 (1.27). These findings support further investigation of selected EOs as complementary antimicrobial agents.