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High-throughput quantification of microbial birth and death dynamics using fluorescence microscopy

  • Samuel F. M. Hart
  • David Skelding
  • Adam J. Waite
  • Justin C. Burton
  • Wenying ShouEmail author
Research Article

Abstract

Background

Microbes live in dynamic environments where nutrient concentrations fluctuate. Quantifying fitness in terms of birth rate and death rate in a wide range of environments is critical for understanding microbial evolution and ecology.

Methods

Here, using high-throughput time-lapse microscopy, we have quantified how Saccharomyces cerevisiae mutants incapable of synthesizing an essential metabolite (auxotrophs) 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.

Results

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 compared to no lysine. 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 can vary with metabolite concentration and time.

Conclusions

Our work reveals how time-lapse microscopy can be used to discover non-intuitive microbial birth and death dynamics and to quantify growth rates in many environments.

Keywords

Saccharomyces cerevisiae fluorescence microscopy microbial growth birth rate death rate 

Notes

Acknowledgements

We thank Jose Pineda for performing the experiment in Supplementary Figure S3 and Li Xie for consultation regarding models.

Supplementary material

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Copyright information

© Higher Education Press and Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Samuel F. M. Hart
    • 1
  • David Skelding
    • 1
  • Adam J. Waite
    • 1
  • Justin C. Burton
    • 1
  • Wenying Shou
    • 1
    Email author
  1. 1.Fred Hutchinson Cancer Research CenterSeattleUSA

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