Phenotypic and metabolic characterization of an ADH2 deletion mutant associated with reduced fluconazole susceptibility in Candida albicans.

INTRODUCTION

Alcohol dehydrogenases (ADHs) participate in fungal carbon metabolism and redox homeostasis, but the contribution of ADH2 to virulence-associated phenotypes and fluconazole response in Candida albicans (C. albicans) remains incompletely defined.

METHODS

An ADH2 deletion mutant, adh2Δ/Δ, was generated in a SC5314 genetic background. Growth kinetics, adhesion, filamentous development, biofilm metabolic activity, cell surface hydrophobicity, intracellular ATP, reactive oxygen species, mitochondrial membrane potential, NAD+/NADH ratio, selected glucose-metabolism-related transcript levels, fluconazole response, and in vivo infection burden were evaluated.

RESULTS

The adh2Δ/Δ mutant displayed modestly altered growth kinetics and showed a shorter doubling time than SC5314. However, qualitative assays showed reduced adhesion and filamentous development, accompanied by decreased expression of ALS1, ALS3, and HWP1. The mutant displayed increased biofilm metabolic activity but decreased cell surface hydrophobicity, together with increased CSH1 expression. Metabolic analyses showed reduced ATP levels and basal intracellular ROS-sensitive fluorescence, together with an increased NAD+/NADH ratio and elevated mitochondrial membrane potential. Among selected glucose-metabolism-related genes, HXK1 was significantly upregulated after correction for multiple testing, whereas CDC19, PGI1, and PGK1 were not significantly altered. The adh2Δ/Δ mutant showed a two-fold increase in fluconazole MIC and stronger growth under FLCZ stress, accompanied by increased ERG11 transcript levels, whereas MDR1, CDR1, and CDR2 were not significantly altered after correction for multiple testing; ADH1 was also unchanged. In mice, adh2Δ/Δ infection resulted in higher renal fungal burden and higher residual renal fungal burden after fluconazole treatment.

DISCUSSION: These findings define an ADH2 deletion-associated phenotypic and metabolic/redox state and provide a hypothesis-generating framework for exploring its association with reduced fluconazole susceptibility in C. albicans.