Targeting Candida albicans in dual-species biofilms with antifungal treatment reduces Staphylococcus aureus and MRSA in vitro

Polymicrobial biofilms consisting of fungi and bacteria are frequently formed on endotracheal tubes and may contribute to development of ventilator associated pneumonia (VAP) in critically ill patients. This study aimed to determine the role of early Candida albicans biofilms in supporting dual-species (dual-kingdom) biofilm formation with respiratory pathogens in vitro, and investigated the effect of targeted antifungal treatment on bacterial cells within the biofilms. Dual-species biofilm formation between C. albicans and three respiratory pathogens commonly associated with VAP (Pseudomonas aeruginosa, Escherichia coli and Staphylococcus aureus) was studied using quantitative PCR. It was shown that early C. albicans biofilms enhanced the numbers of E. coli and S. aureus (including methicillin resistant S. aureus; MRSA) but not P. aeruginosa within dual-species biofilms. Transwell assays demonstrated that contact with C. albicans was required for the increased bacterial cell numbers observed. Total Internal Reflection Fluorescence microscopy showed that both wild type and hyphal-deficient C. albicans provided a scaffold for initial bacterial adhesion in dual species biofilms. qPCR results suggested that further maturation of the dual-species biofilm significantly increased bacterial cell numbers, except in the case of E.coli with hyphal-deficient C. albicans (Ca_gcn5Δ/Δ). A targeted preventative approach with liposomal amphotericin (AmBisome(®)) resulted in significantly decreased numbers of S. aureus in dual-species biofilms, as determined by propidium monoazide-modified qPCR. Similar results were observed when dual-species biofilms consisting of clinical isolates of C. albicans and MRSA were treated with liposomal amphotericin. However, reductions in E. coli numbers were not observed following liposomal amphotericin treatment. We conclude that early C. albicans biofilms have a key supporting role in dual-species biofilms by enhancing bacterial cell numbers during biofilm maturation. In the setting of increasing antibiotic resistance, an important and unexpected consequence of antifungal treatment of dual-species biofilms, is the additional benefit of decreased growth of multi-drug resistant bacteria such as MRSA, which could represent a novel future preventive strategy.