Analyses of the Antibiofilm Activity of o-Phenanthroline Monohydrate against Enterococcus faecalis and Staphylococcus aureus and the Mechanisms Underlying These Effects

Enterococcus faecalis and Staphylococcus aureus exhibit robust biofilm formation capabilities, the formation of which is closely linked to pathogenicity and drug resistance, thereby resulting in host infection and treatment failure. o-Phenanthroline monohydrate (o-Phen) and its derivatives demonstrate a wide range of antibacterial and antifungal activities. In this study, we aimed to explore the antibiofilm activity of o-Phen to E. faecalis and S. aureus and provide insights into the molecular mechanisms for combating biofilm resistance. We demonstrated that o-Phen possesses significant antibacterial and antibiofilm properties against E. faecalis and S. aureus, inducing alterations in bacterial morphology, compromising cell membrane integrity, and exhibiting synergistic effects with β-lactam antibiotics at sub-MIC concentrations. The adhesion ability and automatic condensation capacity of, and synthesis of, extracellular polymers by E. faecalis cells were reduced by o-Phen, resulting in the inhibition of biofilm formation. Importantly, transcriptome analysis revealed 354 upregulated and 456 downregulated genes in o-Phen-treated E. faecalis. Differentially expressed genes were enriched in 11 metabolism-related pathways, including amino acid metabolism, pyrimidine metabolism, and glycolysis/gluconeogenesis. Moreover, the oppA, CeuA, and ZnuB genes involved in the ABC transport system, and the PBP1A penicillin-binding protein-coding genes sarA and mrcA were significantly downregulated. The multidrug efflux pump system and membrane permeability genes mdtG and hlyD, and bacterial adhesion-related genes, including adcA and fss2 were also downregulated, while mraZ and ASP23 were upregulated. Thus, o-Phen is anticipated to be an effective alternative drug for the treatment of E. faecalis and S. aureus biofilm-associated infections.