Antifungal Activity of Carvacrol on Ergosterol Synthesis in Multidrug Resistant Candida albicans

Abstract

Introduction: Greatly increased use of antifungal therapies has resulted in the development of multidrug resistant. The phenolic compound carvacrol have been reported to have anti-Candida activity. This work is an attempt to examine effect of carvacrol on ergosterol synthesis against multidrug resistant Candida albicans. Methods: This cross-sectional study has been conducted on 30 immune-compromised patients from vagina, mouth and skin surfaces during 2016-2017. Colonizing clinical isolates of C. albicans were identified and drug resistant isolates detected using WHONET software. The susceptibility tests for carvacrol were carried out in terms of disk diffusion, broth microdilution and time kill assays against multidrug resistant C. albicans. The ability to change from yeast to hyphal morphology exploited using a light microscopy. Ergosterol quantification has been investigated by spectrophotometric analysis. The expression profile of ERG11 gene was studied using quantitative real time RT-PCR to admit the possibility of further associated ergosterol pathway. Statistical analysis was performed with SPSS 21.0. Comparisons were performed using one-way ANOVA, with Tukey's tests. Results: Ten colonizing clinical isolates of C. albicans were identified. Multidrug resistant isolates of C. albicans were detected. Carvacrol was found to have MIC90 of 100-200 µg/ml for multidrug drug resistant isolates of C. albicans. The time kill curve results show that carvacrol could significantly inhibit the growth of C. albicans (P≤0.05). In multidrug resistant C. albicans treated with carvacrol there was a marked reduction of the transition of yeast cells to hyphal cells. Significant decrease of ergosterol content was noted in multidrug resistant C. albicans treated with carvacrol. Furthermore, significant downregulation was observed on ERG11 gene in multidrug resistant C. albicans treated with carvacrol (P≤0.05). Conclusion: Carvacrol show strong antifungal activity against multidrug resistant C. albicans. These results provide proof of concept for the implementation of carvacrol that may have potential applications in the treatment of drug resistance C. albicans infections.

Inhibitory effect of menthol on expression of aspartyl proteinase 1 in fluconazole-resistant Candida albicans

“…Our data support the effects of menthol in fluconazole-resistant C. albicans vaginal isolates. Several lines of evidence demonstrate that natural pure compounds derived from plants are the most potent products to kill antifungal resistant C. albicans (23,30).…”

“…The disk diffusion (CLSI M44-A2) and broth microdilution (CLSI M27-A3 and CLSI M27-S4) methods were used to test antifungal susceptibility of fluconazole against C. albicans. Results of susceptibility of isolates to fluconazole were analyzed using WHONET software (23).…”

“…The lowest concentration in which ≥50% or 90% reduction of growth was evident considered to be a relative MIC. Fluconazole was used as a control (22,23).…”

“…The expression assay of SAP1 gene was conducted using relative quantitative Real time PCR (qRT-PCR) that was adapted from a previously described method (23). C. albicans ATCC MYA-573 treated with menthol and fluconazole was induced to form hypha formation.…”

“…The oligonucleotide primers used for qRT-PCR were listed in Table 1. The expression levels of SAP1 gene were calculated by comparative Ct method (2 −∆Ct formula) after normalization with beta actin gene (23,25).…”