Development of a Fluorescent Multiwell Assay for Evaluating the Capacity of the Ciliated Protozoan Tetrahymena for Bacterivory in Water Samples

Abstract Bacterivory by ciliates in various water ecosystems, both natural and artificial, plays a significant role on the microbial population composition and consequently affects water quality. A convenient, rapid and inexpensive methodology to evaluate the capacity of the ciliate protozoan Tetrahymena thermophila for bacterivory was developed utilizing fluorescent protein expressing bacteria (FPEB) in a microtitre plate fluorimeter. Bacterivory was correlated with a loss in fluorescence measured in the fluorimeter and confirmed by fluorescence microscopy showing that the FPEB were engulfed during the assay and subsequently lost their fluorescence, whereas Cytochalasin B, a known inhibitor of phagocytosis, prevented a decrease in relative fluorescence units (RFUs). The ciliate bacterivory (CB) assay has a great dynamic range allowing the assay to be performed with a variety of predator:prey concentrations. A model toxicant, CuCl2, known to have a toxicological impact on protozoa and often present in different types of wastewater, resulted in measurable decreases in bacterivory. As well, starvation of Tetrahymena for 24 h resulted in reduced bacterivory. In the future, the CB assay could be developed for water monitoring purposes to rapidly assess water samples for the capacity to support bacterivory as an indicator of ecosystem health.