Molecular Mechanism of Action of HOCl from Neutral-pH Electrolysed Oxidising Water Against Candida albicans

Chemical disinfection of removable acrylic dental prostheses minimizes the risk of denture stomatitis caused by the opportunistic fungal pathogen <i>Candida albicans</i>. We previously reported that neutral-pH electrolysed oxidising water (EOW), a hypochlorous acid (HOCl)-based biocide, is effective at inhibiting <i>C. albicans</i> biofilm formation on denture resins. Knowledge about the mechanism of action of EOW on <i>C. albicans</i> is lacking. This study investigated the molecular mechanism of action of neutral-pH EOW against <i>C. albicans</i> cells that were incubated with sub-inhibitory concentrations of EOW-HOCl (treatment with 0.125× MIC₉₀ EOW-HOCl (15 µM; T0.125) or treatment with 0.5× MIC₉₀ EOW-HOCl (59 µM; T0.5)). RNA-sequencing (RNA-seq) was used to identify differentially expressed genes (DEGs) which were validated by qRT-PCR. Ninety-five DEGs were identified between the treated and untreated cells after a 60 min exposure. A moderate sub-inhibitory EOW-HOCl concentration (T0.125) caused significant upregulation (log₂ fold change > +2) of genes responsive to oxidative stress (<i>EBP1</i>, <i>GAP6</i>, <i>PRN1</i>, <i>HSP21</i>), weak organic acid stress (<i>PRN1</i>), and heat-shock (<i>HSP21</i>). A higher sub-inhibitory concentration (T0.5) caused a significant downregulation of most DEGs (notably, -1.9 to -3 log₂ fold reduction in <i>SUT1</i>, <i>HNM3</i>, <i>STP4</i> expression), cessation of growth, and an upregulation of genes involved in ammonia transport, carbohydrate metabolism, and the unfolded protein and apoptotic response pathways (<i>ATO2</i>, <i>IRE1</i>). Our findings reveal <i>HSP21</i> and <i>PRN1</i> to be possible key players in protecting <i>C. albicans</i> cells against HOCl, a natural biocide of the innate immune system.