In vivo imaging of disseminated murine Candida albicans infection reveals unexpected host sites of fungal persistence during antifungal therapy

Objectives: Candida albicans is an important fungal pathogen that can cause life-threatening disseminated infections. To determine the efficacy of therapy in murine models, a determination of renal fungal burden as cfu is commonly used. However, this approach provides only a snapshot of the current situation in an individual animal and cryptic sites of infection may easily be missed. Thus, we aimed to develop real-time non-invasive imaging to monitor infection in vivo. Methods: Bioluminescent C. albicans reporter strains were developed based on a bioinformatical approach for codon optimization. The reporter strains were analysed in vitro and in vivo in the murine model of systemic candidiasis. Results: Reporter strains allowed the in vivo monitoring of infection and a determination of fungal burden, with a high correlation between bioluminescence and cfu count. We confirmed the kidney as the main target organ but additionally observed the translocation of C. albicans to the urinary bladder. The treatment of infected mice with caspofungin and fluconazole significantly improved the clinical outcome and clearance of C. albicans from the kidneys; however, unexpectedly, viable fungal cells persisted in the gall bladder. Fungi were secreted with bile and detected in the faeces, implicating the gall bladder as a reservoir for colonization by C. albicans after antifungal therapy. Bile extracts significantly decreased the susceptibility of C. albicans to various antifungals in vitro, thereby probably contributing to its persistence. Conclusions: Using in vivo imaging, we identified cryptic sites of infection and persistence of C. albicans in the gall bladder during otherwise effective antifungal treatment. Bile appears to directly interfere with antifungal activity.