The Use of Chemostats in Microbial Systems Biology

Abstract

Cells regulate their rate of growth in response to signals from the external world. As the cell grows, diverse cellular processes must be coordinated including macromolecular synthesis, metabolism and ultimately, commitment to the cell division cycle. The chemostat, a method of experimentally controlling cell growth rate, provides a powerful means of systematically studying how growth rate impacts cellular processes -including gene expression and metabolism -and the regulatory networks that control the rate of cell growth. When maintained for hundreds of generations chemostats can be used to study adaptive evolution of microbes in environmental conditions that limit cell growth. We describe the principle of chemostat cultures, demonstrate their operation and provide examples of their various applications. Following a period of disuse after their introduction in the middle of the twentieth century, the convergence of genome-scale methodologies with a renewed interest in the regulation of cell growth and the molecular basis of adaptive evolution is stimulating a renaissance in the use of chemostats in biological research. Video LinkThe video component of this article can be found at

Steady-State Growth under Inorganic Carbon Limitation Conditions Increases Energy Consumption for Maintenance and Enhances Nitrous Oxide Production in Nitrosomonas europaea

“…We hypothesized that IC limitation coupled with energy wasted on N 2 O production would increase cellular maintenance energy and decrease the optimal growth rate, leading to a washout at a slower doubling time (T d ) than previously reported (31,32). Continuous-culture theory predicts that the dilution rate is equal to at steady state and that cells will begin to wash out when the culture approaches the maximum specific growth rate ( max ) (21,33 (Fig. S1B) (34).…”

“…At this point, medium feed commenced and continuous culturing began. Continuous cultures were allowed at least three population doublings to reach steady state, which was defined as one population doubling with a Յ5% change in cell density (21). Approximately 200-l aliquots of chemostat cultures were plated periodically on Luria-Bertani agar plates to confirm axenic culture conditions.…”

The enduring utility of continuous culturing in experimental evolution

“…Through the process of continuous dilution a growing population of cells can be maintained in a chemostat indefinitely. An essential requirement of the chemostat is the use of a defined medium in which a single nutrient is present at a growth limiting concentration [21]. A nutrient is said to be limiting in the chemostat when its concentration dictates the steady-state cell density, such that increasing the concentration of the limiting nutrient results in a proportional increase in the steady-state cell density.…”

The functional basis of adaptive evolution in chemostats

“…More than 60 years later, the advent of genome-scale experimental methods and a reinvigoration of a quantitative approach to biology (i.e. systems biology) has stimulated renewed interest in the use of chemostats (Hoskisson & Hobbs 2005; Bull 2010; Ziv, Brandt, et al 2013a). The primary research applications of chemostats remain the study of cell growth and adaptive evolution, which owing to their central importance in cell and evolutionary biology, remain vital areas of investigation.…”