Berberine disrupts staphylococcal proton motive force to cause potent anti-staphylococcal effects in vitro

Neil Zhao; Selin Isguven; Rachel Evans; Thomas P Schaer; Noreen J Hickok

doi:10.1016/j.bioflm.2023.100117

Berberine disrupts staphylococcal proton motive force to cause potent anti-staphylococcal effects in vitro

Biofilm. 2023 Apr 1:5:100117. doi: 10.1016/j.bioflm.2023.100117. eCollection 2023 Dec.

Authors

Neil Zhao¹, Selin Isguven^{1

2}, Rachel Evans¹, Thomas P Schaer³, Noreen J Hickok¹

Affiliations

¹ Department of Orthopaedic Surgery, Sidney Kimmel Medical College of Thomas Jefferson University, Philadelphia, PA, USA.
² Department of Radiology, Sidney Kimmel Medical College of Thomas Jefferson University, Philadelphia, PA, USA.
³ Department of Clinical Studies, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Kennett Square, PA, USA.

Abstract

The presence of antibiotic resistance has increased the urgency for more effective treatments of bacterial infections. Biofilm formation has complicated this issue as biofilm bacteria become tolerant to antibiotics due to environmental factors such as nutrient deprivation and adhesion. In septic arthritis, a disease with an 11% mortality rate, bacteria in synovial fluid organize into floating, protein-rich, bacterial aggregates (mm-cm) that display depressed metabolism and antibiotic tolerance. In this study, Staphylococcus aureus (S. aureus), which is the most common pathogen in septic arthritis, was tested against different inhibitors that modulate bacterial surface protein availability and that should decrease bacterial aggregation. One of these, berberine, a quaternary ammonium compound, was found to reduce bacterial counts by 3-7 logs in human synovial fluid (aggregating medium) with no effect in tryptic soy broth (TSB, non-aggregating). Unlike traditional antibiotics, the bactericidal activity of berberine appeared to be independent of bacterial metabolism. To elucidate the mechanism, we used synovial fluid fractionation, targeted MRSA transposon insertion mutants, dyes to assess changes in membrane potential (DiSC₃(5)) and membrane permeability (propidium iodide (PI)), colony counting, and fluorescence spectroscopy. We showed that berberine's activity was dependent on an alkaline pH and berberine killed both methicillin-sensitive S. aureus and MRSA in alkaline media (pH 8.5-9.0; p < 0.0001 vs. same pH controls). Under these alkaline conditions, berberine localized to S. aureus where berberine was isolated in cytoplasmic (∼95%) and DNA (∼5%) fractions. Importantly, berberine increased bacterial cell membrane permeability, and disrupted the proton motive force, suggesting a mechanism whereby it may be able to synergize with other antibacterial compounds under less harsh conditions. We suggest that berberine, which is cheap and readily available, can be made into an effective treatment.

Keywords: Berberine; Septic arthritis; Staphylococcus aureus; Synovial fluid.

Abstract

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