An improved viability assay for Acanthamoeba castellanii trophozoites reveals drug-induced pseudocyst formation

Acanthamoeba castellanii is a free-living amoeba (FLA) that causes fatal human infections with few effective treatments. One limitation of new drug development is the lack of accurate, high-throughput, and quantitative viability assays that score both trophozoites and cysts. A colorimetric assay using Sulforhodamine B (SRB), which measures cell adherence, has previously been adapted for A. castellanii trophozoites. In this study, we demonstrated that the SRB assay can be optimized to serve as a robust, high-throughput platform that can measure viable Acanthamoeba trophozoites, pseudocysts and cysts. We used this assay to measure the IC50 of 18 commonly used drugs and disinfectants on A. castellanii trophozoites, demonstrating that several clinically used drugs induce pseudocyst formation rather than amoeba death.