Synergistic association of clioquinol with antifungal drugs against biofilm forms of clinical Fusarium isolates

Summary Background The influence of biofilm on the complexity of fungal diseases has been reported in recent years, especially in non‐invasive mycoses such as keratitis and onychomycosis. The difficulty in treating cases of fusariosis in the human medical clinic exemplifies this situation, because when Fusarium spp. are present in the form of biofilm, the permeation of antifungal agents is compromised. Objectives This study proposes an association of clioquinol, an inhibitor of fungal cells with antifungal drugs prescribed to combat fusariosis in humans. Methods Susceptibility was assessed by microdilution in broth. Formation of biofilm by staining with violet crystal. Inhibition and removal of biofilm using the MTT colorimetric reagent. Time‐kill combination, hypoallergenicity test, cytotoxicity test and toxicity prediction by computer analysis were also performed. Results Clioquinol associated with voriconazole and ciclopirox inhibited biofilm formation. Possibly, clioquinol acts in the germination and elongation of hyphae, while voriconazole prevents cell adhesion and ciclopirox the formation of the extracellular polymeric matrix. The CLIO‐VRC association reduced the biofilm formation by more than 90%, while the CLIO‐CPX association prevented over 95%. None of the association was irritating, and over 90% of the leucocytes remained viable. Computational analysis does not reveal toxicity relevant to CLIO, whereas VRC and CPX showed some risks for systemic use, but suitable for topical formulations. Conclusions The combination of CLIO‐VRC or CLIO‐CPX proved to be a promising association strategy in the medical clinic, both in combating fungal keratitis and onychomycosis, since they prevent the initial process of establishing an infection, the formation of biofilm.