Live real-time imaging of life-threatening invasive fungal infections

Candida bloodstream infections remain a significant problem in severely ill patients, particularly those in intensive care units and surgical patients. Mortality rates remain high due to difficulties in diagnosis, delays in initiation of antifungal therapy and limited choices of antifungal drugs. Animal models of Candida bloodstream infection remain a vital element in the development of new antifungal agents and diagnostic tests, as well as increasing our understanding of how infection initiates and progresses. However, in temporal studies there is potential for large numbers of animals to be used. Because of inherent biological variation, groups of 3-10 animals are sampled at the each time point, with analyses based upon the mean or median values for each group. In vivo imaging of fungi in a single group of animals over the same time period would significantly (75-90%) reduce the number of animals required for the same study. However, in vivo imaging of invasive fungal infection requires a reporter which can be detected in the internal organs of mice. To date, none of the reporters developed for C. albicans has been able to accurately reflect disease progression. Publication of new near-infrared reporters for mammalian cells has demonstrated that the near-infrared fluorescence can be detected deep within mice. In addition, only an endogenous compound, biliverdin, is required for fluorescence. The aim of this project is to develop a codon-optimized near-infrared reporter (iRFP) for use in in vivo imaging of C. albicans. A range of constructs will be produced, with the Candida optimized reporter (camiRFP) expression controlled by various Candida gene promoters. Strains expressing the various strains will be extensively analysed in laboratory tests to identify the most suitable reporter for in vivo imaging. Finally, the Candida reporter gene will be validated in small scale experimental infection studies.

• Further Funding: NC3Rs Engagement Award, Opening Doors: the 3Rs in medical sciences, August 2016, £930
• Further Funding: NC3Rs PhD Studentship, Reducing animal usage in antifungal drug development: iRFP reporter strains and imaging infection, October 2016, £90,000 
• Further Funding: NC3Rs PhD Studentship, An in vitro model system to assay kidney-pathogen interactions determining outcome of Candida albicans infection, October 2010, £120,000