Abstract
Emergence of drug-resistant strains of Candida and inefficiency of conventional antifungal therapy has necessitated the search for alternative and new antifungal agents. Inhibition of virulence and biofilm are the potential drug targets. In this study, the oils of Carum copticum, Thymus vulgaris and their major active compound thymol as revealed by Gas chromatography and gas chromatography-mass spectrometry (GC-GC/MS) analysis were tested for their inhibitory activity against growth to determine sub-MIC values against 27 drug-resistant strains of Candida spp.
Brothmacrodilution method was used for determination of MIC of test oils against Candida strains. The spectrophotometric methods were used for detection and inhibition assays for virulence factors in Candida spp. Light and electron microscopy was performed to observe morphological effects of oils on biofilms. GC-GC/MS were used to evaluate the major active compounds of test oils.
Virulence factors like proteinase and haemolysin were detected in 18 strains, both in solid and liquid media. A 70% of the test strains exhibited hydrophobicity and formed moderate to strong biofilms (OD280 0.5- > 1.0). Test oils exhibited MICs in the range of 45-360 μg.mL(-1) against the majority of test strains. All the oils at 0.25× and 0.5× MICs induced >70% reduction in the cell surface hydrophobicity, proteinase and haemolysin production. At 0.5× MIC, thymol and T. vulgaris were most inhibitory against biofilm formation. At sub-MICs electron microscopic studies revealed the deformity of complex structures of biofilms formed and cell membranes appeared to be the target site of these agents.
Therefore, our findings have highlighted the concentration dependent activity of oils of C. copticum and T. vulgaris against virulence factors and biofilms in proteinase and haemolysin producing drug-resistant strains of Candida spp. The above activities of test oils are supposed to be mainly contributed due to their major active compound thymol. Further mechanism involving anti-proteinase, anti-haemolysin and anti-biofilm activities of these oils and compounds are to be explored for possible exploitation in combating Candida infections.