Euglena gracilis as a high-throughput screening platform for antibacterial activity, cytotoxicity and membrane permeability in a one-step and cost-effective assay.

There is an urgent need for the development of new antibacterial drugs, caused by the increasing number of resistances. The first step for the development of new antibacterial compounds is usually a high-throughput screen of naturally occurring or synthetic compounds. Here, we propose to screen compound libraries on the Euglenoid Euglena gracilis. This protozoan has obtained a chloroplast via secondary endosymbiosis of an alga, but still maintains its ability to metabolise organic carbon sources. Importantly, the chloroplast has preserved some bacterial features and can be targeted by a range of antibiotics, resulting in organelle loss and Euglena bleaching, while the mixotrophic metabolism ensures growth of Euglena. Therefore, Euglena allows simultaneous screening of a compound library for (i) antibacterial activity (Euglena bleaches), (ii) absence of toxicity (Euglena grows) and (iii) membrane permeability across at least three membranes of both eukaryotic and prokaryotic origin. We have established and optimised all the parameters needed for such an Euglena-based antibacterial drug screen. We define absorbance parameters to distinguish the three possible outcomes (bleaching, growing and dying) in an automatised way. We have successfully tested the screen on both a commercially available compound library from MedChemExpress and on a self-assembled library of rare natural compounds from myxobacteria and fungi. Our Euglena-based screening platform provides a novel high-throughput method to screen for new compounds with antibiotic properties, in a cost-effective way and with any library. While some classes of antibiotics will be missed, the screen is unbiased and has the potential to discover novel antibiotic targets.