COMPARATIVE STUDY OF BERBERINE AND CAPSAICIN AS EFFLUX PUMP INHIBITORS IN QUINOLONE RESISTANT STAPHYLOCOCCUS AUREUS ISOLATES FROM BOVINE MASTITIS
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Date
2021-08-30
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COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR
Abstract
Antimicrobial resistance which causes failure of antibiotic therapy has
become a global issue nowadays. Bacteria develop resistance towards an antibacterial
agent via many mechanisms, the most important one is the drug efflux mediated
through efflux pumps. Bacterial resistance causes reduction in the usage of several
chemotherapeutic agents in the treatment of bacterial diseases. In this context,
research on natural products from plants which can be used as plausible candidates to
combat antimicrobial resistance will be of utmost importance. Hence, a study was
undertaken to assess the antibacterial activity and resistance modulation through
down regulation of norA, norB, and norC by two plant alkaloids berberine and
capsaicin against quinolone-resistant S. aureus from bovine mastitis.
Disc diffusion assay and broth microdilution assay were performed to assess
the antibacterial activity of berberine and capsaicin alone (at various concentrations
viz 8 g/L to 0.25 g/L) and in combination of antibiotics with enrofloxacin,
norfloxacin and nalidixic acid. Resistance modulating potential through efflux pump
inhibition by berberine and capsaicin in combination with quinolones at sub
inhibitory concentration in down regulating norA, norB, and norC was executed by
reverse transcription and quantitative real-time PCR. Antibiofilm assay was
conducted for various combinations of quinolones with berberine and capsaicin using
congo red agar plate method. In silico studies were carried out using autodock
software to analyse the binding of berberine and capsaicin to NorA protein.
The findings of disc diffusion assay revealed that berberine and capsaicin in
combination with enrofloxacin, norfloxacin and nalidixic acid increased the zone of
inhibition significantly (p-value < 0.05) in a dose-dependent manner. Checkerboard
assay showed that berberine lowered the MIC of antibiotics tested up to eightfold and
capsaicin up to fourfold. The combination of enrofloxacin with berberine exhibited
synergy and with capsaicin exhibited partial synergy. The combination of norfloxacin
with test compounds exhibited partial synergy and the combination of berberine and
capsaicin with nalidixic acid exhibited additivity. Gene expression studies indicated
downregulation in the expression of norA and norC genes, while norB was not
expressed in any of the isolates. Berberine and capsaicin in combination with
quinolones could inhibit the biofilm formation by S. aureus isolates in a dose dependent manner. In silico studies revealed that capsaicin bind to NorA protein.
To consolidate, berberine and capsaicin can be promising drug molecules in
modulating the drug resistance mediated by the Nor efflux pumps in S. aureus from
bovine mastitis. Thus, berberine and capsaicin can be used as an adjunct to antibiotics
for the treatment of bacterial diseases.
Description
Submitted in partial fulfillment of the requirement for the degree of Master of Veterinary Science in Veterinary Pharmacology and Toxicology