ABSTRACT

Background: The widespread methicillin-resistant Staphylococcus aureus (MRSA) dissemination poses significant public health challenges, particularly because of limited elimination options for related infections. the present study antimicrobial effects of the natural compound Kaempferol against MRSA, singly and in combination with others (such as metformin), and examined its impact on the expression of key virulence genes associated to MRSA pathogenicity.
Methods: Ten MRSA isolates were collected from clinical specimens of 12–55 years-aged patients. The minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs), of the agents were tested within a range of 0.5–1024 µg/mL. Anti-biofilm effects were assessed using a microplate tissue assay. The expression levels of selected virulence genes (spa, fnbA and pvl) were quantified by real-time PCR following exposure to the agents.
Results: There was a synergistic and additive antibacterial effects between kaempferol and metformin. Also, metformin and kaempferol had anti-biofilm effects. Kaempferol at sub-MIC concentration could decrease the expression of fnbA, pvl, and spa genes by mean ±SD of 1.6 ±0.6 (p<0.05), 1.8 ±0.1 (p<0.05), and 1.2 ±0.4 fold (p<0.05), respectively in comparison with control. The effect of metformin at sub-MIC level was also significant on these genes including 1.9±0.6 (p<0.05), 2.2 ±0.3 (p<0.05), and 1.9 ±0.4 fold (p<0.05), respectively.
Conclusion: Kaempferol and metformin had antibacterial and anti-biofilm activities against MRSA isolates. Further studies with larger numbers of isolates and detailed molecular mechanistic research is warranted to confirm findings.

KEYWORDS

Staphylococcus aureus Methicillin resistance, Virulence genes, Kaempferol, Antibiotic synergy

REFERENCES

1) Nazli, A., et al., Treatment of MRSA infection: where are we? Current medicinal chemistry, 2024. 31(28): p. 4425–4460.
2) Kaushik, A., et al., Biofilm producing methicillin-resistant Staphylococcus aureus (MRSA) infections in humans: Clinical implications and management. Pathogens, 2024. 13(1): p. 76.
3) Silva, V., et al., Biofilm formation of multidrug-resistant MRSA strains isolated from different types of human infections. Pathogens, 2021. 10(8): p. 970.
4) Patel, H. and S. Rawat, A genetic regulatory see-saw of biofilm and virulence in MRSA pathogenesis. Frontiers in microbiology, 2023. 14: p. 1204428.
5) Zhou, H., et al., The antibacterial activity of kaempferol combined with colistin against colistin-resistant gram-negative bacteria. Microbiology Spectrum, 2022. 10(6): p. e02265–22.
6) Periferakis, A., et al., Kaempferol: antimicrobial properties, sources, clinical, and traditional applications. International journal of molecular sciences, 2022. 23(23): p. 15054.
7) Hairil Anuar, A.H., et al., Critical evaluation of green synthesized silver nanoparticles-kaempferol for antibacterial activity against methicillin-resistant Staphylococcus aureus. International Journal of Nanomedicine, 2024: p. 1339–1350.
8) Deng, S.-P., et al., Facile synthesis of long-term stable silver nanoparticles by kaempferol and their enhanced antibacterial activity against Escherichia coli and Staphylococcus aureus. Journal of Inorganic and Organometallic Polymers and Materials, 2021. 31(7): p. 2766–2778. 
9) Majumdar, G. and S. Mandal, Antibacterial activity analysis of kaempferol and its derivatives targeting virulence and quorum sensing associated proteins by in silico methods. The Microbe, 2025. 6: p. 100259.
10) Wang, L., et al., Linalool as a potential agent for inhibiting Escherichia coli biofilm formation and exopolysaccharide production. BMC Veterinary Research, 2025. 21(1): p. 235.
11) Lahiri, D., et al., Antibiofilm and anti‐quorum sensing activities of eugenol and linalool from Ocimum tenuiflorum against Pseudomonas aeruginosa biofilm. Journal of Applied Microbiology, 2021. 131(6): p. 2821–2837.
12) Hsu, C.-C., et al., The inhibitory activity of linalool against the filamentous growth and biofilm formation in Candida albicans. Medical Mycology, 2013. 51(5): p. 473–482.
13) Manoharan, R.K., et al., Inhibitory effects of the essential oils α-longipinene and linalool on biofilm formation and hyphal growth of Candida albicans. Biofouling, 2017. 33(2): p. 143–155.
14) Rocha, F.R., et al., Effect of bioactive compounds on the regulation of quorum sensing network-associated genes and virulence in Streptococcus mutans—A systematic review. Archives of Oral Biology, 2020. 119: p. 104893.
15) Cheruvanachari, P., et al., Deciphering the antibiofilm potential of 2-Phenylethyl methyl ether (PEME), a bioactive compound of Kewda essential oil against Staphylococcus aureus. Microbial Pathogenesis, 2023. 179: p. 106093.
16) Vieira, S.F., et al., Plant-derived bioactive compounds as key players in the modulation of immune-related conditions. Phytochemistry Reviews, 2025. 24(1): p. 343–460.
17) Chadha, J., et al., Anti-virulence prospects of Metformin against Pseudomonas aeruginosa: A new dimension to a multifaceted drug. Microbial Pathogenesis, 2023. 183: p. 106281.
18) Khayat, M.T., et al., Synergistic benefits: exploring the anti-virulence effects of metformin/vildagliptin antidiabetic combination against Pseudomonas aeruginosa via controlling quorum sensing systems. Biomedicines, 2023. 11(5): p. 1442.
19) Barisch, C., J.C. Holthuis, and K. Cosentino, Membrane damage and repair: a thin line between life and death. Biological chemistry, 2023. 404(5): p. 467–490.
20) Murínová, S. and K. Dercová, Response mechanisms of bacterial degraders to environmental contaminants on the level of cell walls and cytoplasmic membrane. International journal of microbiology, 2014. 2014(1): p. 873081.
21) Dalbey, R.E., P. Wang, and J.M. van Dijl, Membrane proteases in the bacterial protein secretion and quality control pathway. Microbiology and molecular biology reviews, 2012. 76(2): p. 311–330.
22) Mitchell, A.M. and T.J. Silhavy, Envelope stress responses: balancing damage repair and toxicity. Nature Reviews Microbiology, 2019. 17(7): p. 417–428.
23) Abbas, H.A., A.M. Elsherbini, and M.A. Shaldam, Repurposing metformin as a quorum sensing inhibitor in Pseudomonas aeruginosa. Afr Health Sci, 2017. 17(3): p. 808–819.
24) Hegazy, W.A.H., et al., Repurposing Anti-diabetic Drugs to Cripple Quorum Sensing in Pseudomonas aeruginosa. Microorganisms, 2020. 8(9).
25) Khayat, M.T., et al., Synergistic Benefits: Exploring the Anti-Virulence Effects of Metformin/Vildagliptin Antidiabetic Combination against Pseudomonas aeruginosa via Controlling Quorum Sensing Systems. Biomedicines, 2023. 11(5).
26) S, M.S., et al., Crippling of Klebsiella pneumoniae virulence by metformin, N-acetylcysteine and secnidazole. BMC Microbiol, 2023. 23(1): p. 229.