Drug-resistant clinical Candida responded to 5-aminolevulinic acid photodynamic therapy (ALA-PDT) in vitro via enhanced ROS release.

Photodynamic therapy (PDT) has emerged as a potential strategy to combat drug-resistant fungal infections. This study aimed to evaluate the therapeutic efficacy of 5-aminolevulinic acid PDT (ALA-PDT) against drug-resistant Candida strains isolated from patients with candidiasis and to elucidate its underlying mechanisms. Drug-resistant strains were identified by their reduced antifungal susceptibility to azoles, micafungin, amphotericin B, and terbinafine. RNA sequencing (RNA-seq) was used to assess gene expression related to mycelial formation and fungal growth. Scanning electron microscopy (SEM) was employed to observe cell-wall morphology after ALA-PDT treatment. Biofilm formation and intracellular reactive oxygen species (ROS) levels were also measured to investigate the mechanism of ALA-PDT-mediated fungal killing. Candida albicans and Candida tropicalis accounted for over half, predominantly from male ICU patients aged over 40 years. Candida spp. strains exhibited resistance rates of approximately 69.38% to terbinafine, 40.8% to azole antifungals, and 16.3% to micafungin among 49 drug-resistant isolates. Treatment with 20% ALA-PDT effectively eradicated those azole-resistant C. albicans in vitro, increased fungal killing, and significantly increased intracellular ROS levels. SEM analysis revealed varying degrees of cell-wall disruption, and RNA-seq demonstrated downregulation of mycelia-related genes following ALA-PDT. ALA-PDT effectively inhibits drug-resistant Candida spp. growth by inducing ROS-mediated cell damage and suppressing mycelial gene expression. These findings highlight ALA-PDT as a promising therapeutic approach for refractory candidiasis and provide mechanistic insights into its antifungal activity.