Roller, B. R. K. | Search bioRxiv

bioRxiv2023

Direct quantification of sinking velocities across unicellular algae

Eukaryotic phytoplankton, also known as algae, form the basis of marine food webs and drive marine carbon sequestration when their biomass sinks to the ocean floor. Algae must regulate their vertical movement, as determined by motility and gravitational sinking, to balance access to light at the surface and nutrients in deeper layers. However, the regulation of gravitational sinking velocities remains largely unknown, especially in motile species. Here, we directly quantify single-cell masses and volumes to calculate sinking velocities according to Stokes law in diverse clades of unicellular marine microalgae. Our results reveal the cell size, light, and nutrient-dependency of sinking velocities. We identify motile dinoflagellate and green algal species that increase their sinking velocity in response to starvation. Mechanistically, this increased cell sinking is achieved by photosynthesis-driven accumulation of carbohydrates, which increases cell mass and density. Moreover, cell sinking velocities correlate inversely with proliferation rates, and the mechanism regulating cell sinking velocities integrates signals from multiple nutrients. Our findings suggest that the regulation of cell composition according to environmental conditions contributes to the vertical movement of motile cells in the oceans. More broadly, our approach for sinking velocity measurements expands the study of gravitational sinking to motile cells and supports the modeling of marine carbon pump and nutrient cycles.

ecology↗

bioRxiv2023

Single-cell mass distributions reveal simple rules for achieving steady-state growth

Optical density is a common method for measuring exponential growth in bacterial batch cultures. However, there is a misconception that such exponential growth is equivalent to steady-state growth, which is a distinct physiological state that improves experimental reproducibility. Determining precisely when steady-state growth occurs is technically challenging and is aided by paired single-cell and population-level measurements. Using microfluidic mass sensors and optical density, we explore when in typical laboratory batch cultures steady-state growth occurs. We show that cell mass increases by an order of magnitude within a few hours of dilution into fresh medium and that steady-state growth is only achieved when cultures are inoculated with high dilutions from overnight stationary phase cultures. At high dilutions, Escherichia coli and Vibrio cyclitrophicus grown in different rich media achieve steady-state growth approximately 4 total biomass doublings after inoculation. We can decompose these dynamics into 3 doublings of average cell mass and 1 doubling of cell number for both species. We also show that batch cultures in rich media depart steady-state growth early in their growth curves at low cell and biomass concentrations. Achieving and maintaining steady-state growth in batch culture is a delicate balancing act, and we provide general guidance for commonly used rich media. Quantifying single-cell mass outside of steady-state growth is an important first step towards understanding how microbes grow in their natural context, where fluctuations pervade at the scale of individual cells. ImportanceMicrobiologists have watched clear liquid turn cloudy for over 100 years. While the cloudiness of a culture is proportional to its total biomass, growth rates using such optical density measurements are challenging to interpret when cells change size. Many bacteria adjust their size at different steady-state growth rates, but also when shifting between starvation and growth. Optical density cannot disentangle how mass is distributed among cells of different sizes, and directly measuring how mass is distributed among cells has been a major challenge. Here we use single-cell mass measurements to demonstrate that a population of cells in batch culture achieves a stable mass distribution for only a short period of time. Achieving steady-state growth in rich medium requires low initial biomass concentrations and enough time for the coordination of individual cell and population growth. Steady-state growth is important for reliable cell mass distributions in a culture and we discuss how mass variation outside of steady-state can impact physiology, ecology, and evolution experiments.

microbiology↗