Antibiofilm Activity of Three Essential Oils Against ESBL-Producing Klebsiella pneumoniae: An In Vitro and In Silico Investigation of Putative Molecular Targets

Biofilm formation is a major contributor to antibiotic resistance in Klebsiella pneumoniae, posing a serious challenge to current therapeutic strategies. Thus, this study aims to evaluate the antibiofilm activity of three essential oils Thymus hirtus Willd. Ssp. algeriensis Boiss, Syzygiuma romaticum, and Eucalyptus globulus against four clinical isolates of ESBL-producing K. pneumoniae, along with the reference strain K. pneumoniae ATCC 700603. The antibiofilm activity of essential oils was assessed with crystal violet assay using MICs ranging from 3.38 ± 0.2 to 27.1 ± 0.56 mg/mL, 2 ± 0.19 to 32 ± 0.55 mg/mL, and 13.78 ± 0.62 to 110.25 ± 3.37 mg/mL, for TEO, SEO and EEO, respectively. In vitro tests showed that S. aromaticum EO and T. algeriensis EO exhibited the best anti-adhesive activity with a percentage of up to 75.39%, while no difference was observed between the EO in their eradication activity. Microscopic observations confirmed the disorganization of the biofilm after treatment with T. algeriensis. The molecular docking analysis of the three EOs main compounds with MrkH, SdiA and MrkD revealed that SdiA was the most favorable target, with p-cymene (−7.7 kcal/mol), α-pinene (−7.5 kcal/mol), and eucalyptol (−7.1 kcal/mol) showing the strongest binding affinities. Thymol and p-cymene showed also a favorable affinity with MrkD. Overall, p-cymene and α-pinene demonstrated the most favorable binding profiles, whereas linalool exhibited the weakest predicted interactions. These results highlight the promising potential of these EOs, as multi-target antibiofilm agents against MDR- K. pneumoniae biofilms.