AbstractMicrobes live in dynamic environments where nutrient concentrations fluctuate. Quantifying fitness (birth and death) in a wide range of environments is critical for understanding microbial evolution as well as ecological interactions where one species alters the fitness of another. Here, using high-throughput time-lapse microscopy, we have quantified howSaccharomyces cerevisiaemutants incapable of synthesizing an essential metabolite grow or die in various concentrations of the required metabolite. We establish that cells normally expressing fluorescent proteins lose fluorescence upon death and that the total fluorescence in an imaging frame is proportional to the number of live cells even when cells form multiple layers. We validate our microscopy approach of measuring birth and death rates using flow cytometry, cell counting, and chemostat culturing. For lysine-requiring cells, very low concentrations of lysine are not detectably consumed and do not support cell birth, but delay the onset of death phase and reduce the death rate. In contrast, in low hypoxanthine, hypoxanthine-requiring cells can produce new cells, yet also die faster than in the absence of hypoxanthine. For both strains, birth rates under various metabolite concentrations are better described by the sigmoidal-shaped Moser model than the well-known Monod model, while death rates depend on the metabolite concentration and can vary with time. Our work reveals how time-lapse microscopy can be used to discover non-intuitive microbial dynamics and to quantify growth rates in many environments.
Isolating live cells after high-throughput, long-term, time-lapse microscopy
