CONCENTRATION DEPENDENT ANTIBIOFILM ACTIVITY OF TEA TREE OIL AGAINST CLINICAL ENTEROBACTER CLOACAE ISOLATED FROM URINARY TRACT INFECTIONS

Abstract

Background: Biofilm-associated urinary tract infections (UTIs) caused by Enterobacter cloacae represents a significant clinical challenge due to antimicrobial resistance. Natural plant-derived products such as Tea Tree Oil (TTO) gained attention as an antibiofilm agent. Methods: A total of 23 clinical E. cloacae isolates sampled from Iraqi patients with UTIs, identified using morphological characteristics, biochemical tests and Vitek-2 compact system. Biofilms forming ability was determined using 96-wells polystyrene flat plate assay. Minimum bactericidal concentrations (MBCs), minimum inhibitory concentrations (MICs) and sub-MICs (SMICs) of TTO determined using resazurin-based assay. Antibiofilm activity was determined via crystal violet assay, at different TTO concentrations by measuring optical densities. The presence of csgA detected via PCR technique and expression levels of csgA quantified via real-time PCR, normalized to the housekeeping gene and analyzed using 2^-ΔΔCt. Results: All E. cloacae isolates were capable of forming biofilms with optical densities ranged from 0.150 to 0.700. Tea tree oil exhibited antibiofilm activity in a concentration-dependent manner. MBCs level significantly reduced biofilm (p < 0.0001), with reductions exceeding 50% in several strong and moderate biofilm-formers. MIC levels showed moderate inhibition, whereas SMICs demonstrated limited but detectable inhibition. Molecular analysis revealed that, exposure to TTO resulted in significant downregulation of the csgA compared to control. Conclusion: TTO at the level of MBCs, MICs and SMICs demonstrates potential anti-biofilm activity against E. cloacae and may be considered as potential natural In vitro anti-biofilm agent, however, further In vivo studies are required to evaluate its possibility to control biofilm associated-UTIs and molecular levels highlighted these findings.