Waite, A. J. | bioRxiv Science

bioRxiv2022

The Yeast Lifespan Machine: a microfluidic platform for automated replicative lifespan measurements

The budding yeast, Saccharomyces cerevisiae, has emerged as a model system for studying the aging processes in eukaryotic cells. However, the full complement of tools available in this organism has not been fully applied, in part because of limitations in throughput that restrict the ability to carry out detailed analyses. Recent advances in microfluidics have provided direct longitudinal observation of entire yeast lifespans, but have not yet achieved the normal scale of operation possible in this model system. Here we present a microfluidic platform, called the Yeast Lifespan Machine, where we combine improvements in microfluidics, image acquisition, and image analysis tools to increase robustness and throughput of lifespan measurements in aging yeast cells. We demonstrate the platforms ability to measure the lifespan of large populations of cells and distinguish long- and short-lived mutants, all with minimal involvement of the experimenter. We also show that environmental pH is capable of significantly modulating lifespan depending on the growth media, highlighting how microfluidic technologies reveal determinants of lifespan that are otherwise difficult to ascertain.

cell biology↗

bioRxiv2018

Microscopy quantification of microbial birth and death dynamics

Microbes 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 how Saccharomyces cerevisiae mutants 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.

microbiology↗