Small molecule II-6s synergises with fluconazole against Candida albicans.

• The small molecule II-6s shows fungicidal effects against Candida albicans (C. albicans) and sensitizes C. albicans resistant strain to fluconazole. • II-6s synergises with fluconazole by inhibiting the membrane efflux pump of C. albicans. • II-6s induces mitochondrial damage of C. albicans and affects energy metabolism. • II-6s inhibits MAPK signalling pathway, especially Hog1 of C. albicans. Candida albicans (C. albicans) is the most common opportunistic fungal species in the oral cavity. The emergence of drug resistance of C. albicans has necessitated the development of novel antifungal agents. This study evaluated the antifungal activity of a previously developed antimicrobial small molecule, namely II-6s, and explored its potential synergism with fluconazole against C. albicans and the underlying mechanisms. The antifungal effects of II-6s against C. albicans were evaluated with microdilution and time-killing assays. Synergism of II-6s with fluconazole was determined by calculating the fractional inhibitory concentration index (FICI). The effects of II-6s on efflux pump, mitochondrial function and energy metabolism were examined to investigate the underlying mechanism of synergism. The antifungal mechanism of II-6s against C. albicans was further explored with RNA-seq and validated with specific mutant strains. II-6s exhibited a fungicidal effect against C. albicans with a minimum fungicidal concentration of 31.25 µg/mL. II-6s also inhibited C. albicans biofilm with a sessile minimum inhibitory concentration at 500 µg/mL. More importantly, II-6s showed a synergistic effect with fluconazole against a fluconazole-resistant strain of C. albicans , which expressed elevated levels of CDR1 (FICI < 0.5). II-6s inhibited the efflux pump activity of C. albicans. Consistently, II-6s inhibited energy metabolism of C. albicans by reducing the mitochondrial membrane potential and ATP generation, and inhibited utilisation of the non-fermentable carbon source. II-6s also inhibited the mitogen-activated protein kinase signal pathway, particularly HOG1 , which may explain its antifungal activity against C. albicans. The small molecule II-6s inhibits the growth of C. albicans by targeting HOG1. II-6s also synergises with fluconazole by inhibiting the drug efflux pump, representing a promising antifungal agent for the control of candidiasis. [ABSTRACT FROM AUTHOR]