A-052. Effect of Goldenseal Leaf Extract on the Multi Drug Resistance Pumps of Staphylococcus aureus
Background: Efflux-related multi drug resistance mechanisms have long been described in S. aureus. The most common of these include those mediated by QacA and NorA, which are MDR transporters, and the more specific MsrA and TetK. Here we report that Goldenseal (Hydrastis canadensis) leaf extract, although limited in its antimicrobial activities, potentiated the antimicrobial activities of different agents on S. aureus by repression of the MDR pump genes. Methods: S. aureus strains expressing different MDR pumps are listed in Table. Minimal inhibitory concentration (MIC) was determined by the broth dilution method. Total RNA was extracted from strains grown in sub lethal doses of leaf extract or reserpine for 18hrs at 37°C. qPCR was performed using SYBR green fluorescence dye and normalized to the 16s RNA housekeeping gene. Total RNA from untreated cells was used as control. Uptake of EtBr was achieved by adding 20µg/ml EtBr and 20µg/ml reserpine to cells in log phase at 37°C for 1 hr. Reserpine facilitates the uptake of EtBr by the cells. In a separate experiment, efflux of EtBr was measured by loading the cells with 20µg/ml EtBr and 20µg/ml reserpine for 30 min at room temp, washed and resusupended in media with or without inhibitors (reserpine or leaf extract). Effect of inhibitors was measured at 530/600 nm (excitation/emission). Results: Decrease in the MIC (Table 1) and repression of respective gene (Image 1) were observed when treated with the Goldenseal Leaf extract. EtBr uptake studies showed that addition of reserpine marked an increase in the uptake. No change was observed on addition of leaf extract. Reserpine decreased the efflux of EtBr while leaf extract did not. Conclusion: qPCR results indicated that Goldenseal leaf extract repressed the overexpression of MDR pump genes while reserpine did not. EtBr uptake/efflux results indicated that Goldenseal leaf extract has little effect on the normal function of the efflux pumps while reserpine did.
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MDR pump inhibitory activity of Leaf extract. MIC of Leaf extract in parenthesis |
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Antimicrobial/Concentration in µg/ml |
SA1199B(1000) |
SA K2068(500) |
RN4220(1000) |
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Ciprofloxacin |
8 |
4 |
0.25 |
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Tetracycline |
0.25 |
1 |
0.25 |
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Erythromycin |
0.25 |
0.5 |
64 |
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List of strains and MDR pumps expressed |
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Strain |
MDR Pump |
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SA 1199B |
NorA MDR pump |
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SA K2068 |
MepA MDR pump |
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RN 4220 |
MsrA macrolide efflux pump |