Identification, antifungal resistance profile, in vitro biofilm formation and ultrastructural characteristics of Candida species isolated from diabetic foot patients in Northern India

摘要

Purpose: Diabetic foot ulcers are a serious cause of diagnostic and therapeutic concern. The following study was undertaken to determine thefungal causes of diabetic foot ulcers, with their phenotypic and genotypic characterisation. Materials and Methods: A total of 155 diabetic footulcers were studied for 1 year. Deep tissue specimen was collected from the wounds, and crushed samples were plated on Sabouraud dextroseagar with chloramphenicol (0.05 g). Identification was done by growth on cornmeal agar, germ tube formation and urease test. For molecularidentification, conserved portion of the 18S rDNA region, the adjacent internal transcribed spacer 1 (ITS1) and a portion of the 28S rDNA regionwere amplified, using the ITS1 and ITS2 primers. Antifungal susceptibility against voriconazole, fluconazole and amphotericin B was determined bystandard broth microdilution method. Biofilm formation was studied in three steps. First, on the surface of wells of microtiter plates followed byquantification of growth by fungal metabolism measurement. Finally, biofilms were analysed by scanning electron microscopy (SEM). Results:Fungal aetiology was found in 75 patients (48.38%). All were identified as Candida species (100%). The prevalence of different species wasCandida tropicalis (34.6%), Candida albicans (29.3%), Candida krusei (16.0%), Candida parapsilosis (10.6%), Candida glabrata(9.33%). All were susceptible to amphotericin B (100%). On microtiter plate, all the isolates were viable within 48 h showing biofilms. Themetabolic activity of cells in the biofilm increased with cellular mass, especially in the first 24 h. On SEM, majority showed budding yeast form.Conclusion: Non-albicans Candida spp. with potential biofilm forming ability are emerging as a predominant cause of diabetic foot ulcers.