Candida albicans is a prevalent opportunistic pathogenic fungus [1]. On October 25, 2022, the World Health Organization issued its first fungal priority pathogens list, which classified C. albicans as a critical-priority pathogen for research, with an emphasis on its increasing risk of azole resistance [2]. Several studies have demonstrated that biofilm formation is crucial for drug resistance in C. albicans [3]. Biofilms are complex three-dimensional structures comprising yeast spores, hyphae and an extracellular matrix. Compared with non-C. albicans, C. albicans biofilms have greater maturity and structural complexity. Genes upregulated during biofilm formation are evolutionarily more recent than those that are downregulated [4]. The prevalence of biofilm-associated Candidal infections ranges from 67.9 to 94.0% [5]. Biofilm-associated fluconazole resistance can reach 70.5% and biofilm formation increases C. albicans azole resistance by >1 000× [5,6]. Therefore, strategies are urgently needed to overcome biofilm-induced resistance. We previously found that disrupting C. albicans zinc homeostasis reverses the high resistance of biofilms to fluconazole, suggesting that zinc could be a key target for reversing biofilm resistance [[6], [7], [8]].

Zinc is the second largest trace metal required by living organisms. It is crucial for the growth and pathogenicity of pathogenic fungi, as it is a key structural or catalytic cofactor of numerous proteins. Approximately 9% of C. albicans proteins depend on bound zinc for their activity, 70% of which are protein structural cofactors, 18% are enzyme catalytic cofactors and 2% are zinc-transporting proteins [9]. Fungi are susceptible to zinc deficiency and zinc chelation in cells has a broad-spectrum antifungal effect [10]. The ZRT/IRT-like protein (ZIP) family, which regulates zinc homeostasis in C. albicans, is highly evolutionarily conserved [11]. Csr1, a ZRT/IRT-like protein (ZIP) family member involved in regulating homeostasis in C. albicans, is a highly conserved transcription factor that positively regulates zinc uptake [11]. Biofilm formation is the main factor contributing to the enhanced pathogenicity and azole resistance of C. albicans; however, its regulatory mechanisms remain unclear [3].

In this study, we used the zinc chelator N,N,N′,N′-Tetrakis(2-pyridylmethyl)-ethylenediamine (TPEN) and constructed a CSR1-knockout strain to induce intracellular zinc deficiency to investigate the effects and mechanisms of zinc deficiency on biofilm formation and azole resistance in C. albicans [11,12]. Our findings provide a novel strategy for the treatment of fungal infections exhibiting biofilm-related drug resistance and recurrence.