1032-1037The current treatment options for Candida
albicans biofilm-device related infections are very scarce due to their
intrinsic increased tolerance to antimycotics. The aim of this work was to
study synergistic action of terpenes (eugenol, menthol and thymol) with fluconazole (FLA)
on C. albicans biofilm
inhibition. The minimum inhibitory concentration (MIC) assayed
using CLSI M27-A3 broth micro-dilution method showed
antifungal activity against C. albicans MTCC 227 at a concentration of 0.12 % (v/v) for both thymol and eugenol as
compared to 0.25 % (v/v) for menthol. FLA was taken as
positive control. The effect of
these terpenes on metabolic activity of preformed C. albicans biofilm cells was evaluated using 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide
(XTT) reduction assay in 96-well
polystyrene microtiter plate. Thymol and eugenol were more effective at lower
concentrations of ≥ 1.0 % (v/v) than menthol. Synergistic
studies using checkerboard micro-dilution assay showed
fractional inhibitory concentration index
(Σ FIC=0.31) between thymol/FLA followed by eugenol/FLA (Σ FIC=0.37) and
menthol/FLA (Σ FIC<0.5) against
pre-formed C. albicans biofilms. Thymol with fluconazole showed highest
synergy in reduction of biofilm formation than eugenol and menthol which was
not observed when their activities were observed independently. Adherence assay showed 30% viability of C.
albicans cells after 2 h of treatment with 0.05 %
(v/v) thymol/FLA. Effect of thymol/FLA on C.
albicans adhesion visualized by SEM micrographs showed disruption in number
of candidal cells and alteration in structural design of C. albicans. Thus, the study demonstrated synergistic effect of terpenes
with fluconazole on C. albicans biofilm,
which could be future medications for biofilm infections.
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