How to Use a Mutant Library to Identify Genes Required for Biofilm Formation in the Pathogenic Fungus Candida albicans.

With over 1 billion infections and the causative agents showing critical diseases such as pancreatic cancer, the study of pathogenic fungi has never been more critical. In 2017, the United States spent $7.2 billion on fungal diseases. $4.5 billion was allocated to 75,055 hospitalizations, while $2.6 billion went to 8,993,230 outpatient visits. For Candida infections specifically, the cost was $1.4 billion. Currently, there are few classes of antifungals available, and resistance is growing. The identification of genes required for biofilm formation is essential for new antifungal development. This review details how to identify, verify, and characterize defective biofilm formation mutants in C. albicans. This includes how to run an in vitro biofilm formation assay, how to create clean deletions using the modified CRISPR-Cas9 system, how to assay to identify the potential causes of the defect, and how to create complementation strains to confirm the mutant defect.