Quantitative evaluation of tissue invasion by wild type, hyphal and SAP mutants of Candida albicans, and non‐albicans Candida species in reconstituted human oral epithelium

Abstract

Oral candidiasis is a common problem in compromised patients. Although several non-albicans Candida species have emerged as pathogens the majority of candidal infections are caused by Candida albicans. Morphogenesis from the blastospore to filamentous phase, and production of secretory aspartyl proteinases (SAP) are two major virulence attributes of these opportunistic yeast. Histopathology of oral candidiasis is characterized by fungal invasion of the superficial epithelium although the invasive potentials of different Candida species vary. Computerized image analysis systems (IAS) utilizing immunohistochemistry have been successfully employed for quantification of such histopathological features. The purpose of this study was to evaluate quantitatively the in vitro invasive potential of C. albicans and its hyphal and SAP mutants, and five other non-albicans Candida species using a computerized IAS. In vitro human oral candidiasis was produced using five wild type and one reference C. albicans isolates, hyphal and SAP mutants of C. albicans SC 5314, and one wild type and one reference isolate each of C. tropicalis, C. dubliniensis, C. glabrata, C. parapsilosis and C. krusei in a reconstituted human oral epithelium (RHOE) model. The infected tissues were examined histologically at 12, 24 and 48 h. Invading fungal elements were visualized by periodic acid-Schiff (PAS) staining and quantitatively evaluated as a percentage of total tissue invasive area, using a computerized IAS. All C. albicans isolates including hyphal mutant cph1/cph1 and SAP mutants; sap 1-3, sap 4-6 produced hyphae and differentially (P < 0.05) invaded the tissue over 48 h. The invasive potential of hyphal mutant cph1/cph1 and SAP mutants (sap 1-3, sap 4-6) were similar to the parent wild-type isolate at 12 h although after 24 h their invasion was dissimilar (P < 0.05). Non-albicans Candida species and hyphal mutants; efg1/efg1, efg1/efg1 cph1/cph1 were all non-invasive. RHOE model in combination with computerized image analysis permits for the first time, the assessment of invasive potential of Candida species in a quantitative manner. The differential tissue invasive patterns of various C. albicans isolates, their mutants and other Candida species are also described.