An in vitro study of sequential fluconazole / caspofungin treatment against Candida albicans biofilms
Candida albicans is an opportunistic pathogenic fungus causing life threatening diseases in immunocompromised patients. Typically, these infections are associated with the formation of biofilms on both host tissues and implanted biomaterials. Biofilms are defined as complex microbial communities attached to a surface and encased in an exopolymeric matrix. The ability to form biofilms is important from a clinical perspective in that the biofilms show an increased resistance to antifungal therapy and the cells within the biofilms are able to withstand host immune defenses. C. albicans biofilms display intrinsic high levels of resistance to most major classes of antifungal drugs, including the azoles and polyenes. However, our group (and subsequently others) has demonstrated excellent activity of caspofungin, an antifungal drug belonging to the class echinocandins targeting cell wall biosynthesis, against C. albicans biofilms. Our present study indicates that, in a sequential antifungal drug therapy regimen, treatment of fluconazole first followed by caspofungin leads to a significant decrease of the in vitro efficacy of this echinocandin. Multiple clinical isolates of C. albicans demonstrated this antagonism to caspofungin after fluconazole pretreatment, and this effect was also specific only to C. albicans and not manifested by non-albicans Candida spp. Our studies pointed to the role of cell membrane stress response elements, such as calcineurin, in mediating this resistance process. Global gene expression analyses studies on drug treated biofilms revealed increasing levels of gene expression in genes associated with ergosterol and chitin biosynthetic pathways after fluconazole treatment. Expression levels of these genes were further elevated after subsequent caspofungin treatment. Other cell wall associated genes, such as several encoding GPI- anchored proteins were also found significantly regulated during the sequential drug treatment regimen. We hypothesize that treatment of C. albicans biofilms with fluconazole triggers a cell membrane stress response and also results in upregulation of the cell wall salvage pathway, which together render cells in biofilms resistant to caspofungin.