Biofilms of Candida spp. from the ocular conjunctiva of horses with reduced azole susceptibility: a complicating factor for the treatment of keratomycosis?

Abstract

Objectives This study aimed to assess the biofilm-forming ability of Candida spp. from the ocular conjunctiva of horses and to investigate the antifungal susceptibility of these biofilms. Procedures Initially, the biofilm-forming ability of 15 strains was assessed by crystal violet staining, which reveals the fungal biomass adhered to the polystyrene plates, and scanning electron microscopy. Then, the minimum inhibitory concentrations (MICs) of amphotericin B, fluconazole, itraconazole, and caspofungin were initially determined against strains in planktonic form. Afterward, antifungal susceptibility of mature biofilms was evaluated by exposing them to 10 9 MIC and 50 9 MIC of the tested drugs, followed by the assessment of their metabolic activity, using the oxidoreduction indicator XTT. Results were analyzed through ANOVA and Tukey’s post-test, and Pvalues below 5% led to significant conclusions. Results Eight strains produced biofilms and were classified as strong (1/15), moderate (3/15) and weak (4/15) producers, according to the amount of crystal violet retained by the adhered fungal biomass. Biofilm metabolic activity of one C. tropicalis did not decrease after exposure to the tested antifungals, while biofilm metabolic activity of five strains was reduced by amphotericin B, but not the other drugs. One C. parapsilosis sensu stricto and one C. glabrata showed significant reduction in biofilm metabolic activity after exposure to fluconazole, itraconazole, and caspofungin, but not amphotericin B. Conclusions The results demonstrate that Candida from the ocular conjunctiva of horses can pose as a risk to animal health as they are capable of forming biofilms, which are commonly involved in fungal keratitis.