Effect of the antidepressant drug paroxetine in downregulating the biofilm-adhering genes in Staphylococcus aureus: In vitro and in silico studies.

Osteomyelitis is a bacterial infection of the bone that affects millions globally. Due to problems in drug delivery, bacterial resistance through biofilm formation, adverse effects of the medications in use, etc, the scientists are searching for novel antimicrobial agents. As drug repurposing is an excellent method to develop new antimicrobials, this study evaluates the antibacterial and antibiofilm effects of the antidepressant paroxetine, combined with hydroxyapatite (HA), against drug-resistant, biofilm-forming Staphylococcus aureus. The antibacterial activity of paroxetine was assessed using the agar diffusion assay, and the minimum inhibitory concentration (MIC) was determined by the microdilution method. The antibiofilm potential of paroxetine was quantified through the crystal violet assay and further examined using scanning electron microscopy and confocal laser scanning microscopy. The bacterial load on drug-loaded hydroxyapatite was determined using the viable colony count method. The expression of bacterial adhesion genes following paroxetine treatment was analyzed using real-time polymerase chain reaction. Molecular docking studies were performed to evaluate the binding affinity of paroxetine to bacterial adhesion proteins and penicillin-binding proteins. The study demonstrated promising antibacterial properties of the drug and the drug-HA combination against S aureus with a MIC of 18.75 µg/mL. Paroxetine prevented the biofilms formation by S aureus, and could eradicate mature biofilms, with 83%, 86%, and 89% efficacy after 1X MIC, and 2X treatment. The antibiofilm effect was further confirmed by in silico, in vitro methods, wherein a strong affinity was noted for biofilm adhesion protein and paroxetine. Paroxetine treatment revealed downregulation of biofilm-adhering genes, like icaA, clfA, cna, fnbpA, and fib, using RT-PCR. When combined with HA, paroxetine displayed synergistic activity, and this was visualized using confocal laser scanning microscopy, which showed 81% and 19% dead/live cells after treatment, respectively. Furthermore, the scanning electron microscopy analysis displayed the impact of the drug paroxetine on S aureus cell morphology, which showed remarkable damage to the bacterial cells. In silico docking revealed that paroxetine's mode of action was mediated through binding with proteins and penicillin-binding protein, thereby inducing cell death. These results suggest that the paroxetine-HA combination may serve as a promising adjunctive strategy for treating biofilm-associated infections caused by S aureus.