Biology subjects

Keegstra, J. M.

Publications and source records attributed to Keegstra, J. M..

9 recordsLinked to original sources

bioRxiv2026

Microcalorimetry reveals bacterial motility as an expensive trait during starvation

Quantifying the energetic costs of cellular traits is important to understand their evolutionary constraints and ecological function. However, direct measurements of energy expenditure of distinct traits are rare. Here, we present direct measurements of the power dissipation due to bacterial flagellar motility using microcalorimetry, based on measuring the power dissipation of a motile population and a non-motile mutant. For the single-flagellated marine bacterium \textit{Vibrio coralliilyticus}, we determined the power dissipated by motility in cultures to be 0.91 W/mL/OD. Per swimming cell this amounts to 2.0E-15 W, or a turnover of 2.5E4 ATP molecules per second. The power dissipated for motility equals 35 % of total metabolism during starvation, indicating this is likely the most expensive cellular trait during starvation. The dissipation for motility is in good agreement with that predicted using resistive force theory, indicating that any thermodynamic inefficiencies upstream of the flagellar motor, such as metabolic inefficiencies, are small. We further show that the multiflagellated bacterium Escherichia coli dissipates 0.70 W/mL/OD when in motility buffer, comparable to marine bacteria despite the much lower swimming speed, and hence indicating lower efficiency of swimming. Finally, we show that the energy spent by Vibrio coralliilyticus on motility can have major consequences on their survival under starvation, causing a 40 % reduction in survival after 3 weeks of starvation of a motile compared to non-motile mutant. These results reveal the large costs involved in an individual trait that shapes microbial life in heterogeneous environments.

biophysics↗

bioRxiv2026

Inherited flagellar structures coordinate the onset of swimming among related E. coli cells

Bacteria commit to costly behaviors before they can benefit from them. In Escherichia coli, flagellar synthesis is expensive, yet the genes are expressed in stochastic pulses and only a small minority of pulses result in motility. Despite this noise, we find that lineage-related cells begin swimming at similar times, in groups of two to eight cells. Rare, high-amplitude pulses commit a single cell within one generation, but more commonly, pulses rise slowly and fail to trigger swimming before division. Rather than decaying, their output accumulates as cells inherit hook-basal bodies (HBBs) and continue to build new ones. This structural inheritance acts as a physical integrator, summing ongoing transcriptional activity across generations, so that an incomplete flagellar cascade is carried forward at division rather than lost. HBB accumulation sets when cells swim, and the anti-sigma factor FlgM sets how sharply. By holding late flagellar genes off until several HBBs have accumulated, FlgM makes motility depend steeply on early flagellar gene expression, so that relatives inheriting similar sets of HBBs become motile together. Without FlgM, this sharp dependence is largely lost, and activations of the full cascade do not reliably produce swimming. Together, our results show how the accumulation of a molecular machine integrates fluctuations, and how a checkpoint on its assembly sharpens them into a decision, setting both the speed and heritability of a behavioral trait.

systems biology↗

bioRxiv2026

Heritable diel energy reserves enhance diatom growth

Diatoms are important drivers of marine primary production and biogeochemistry1-4. Compared to other phytoplankton groups, diatoms divide more asynchronously (i.e., divisions occur in a manner that is less aligned with the diel cycle) under non-limiting conditions, resulting in divisions occurring during both day and night5-16. However, the mechanisms and rates of asynchronous division have remained elusive. Here, using microfluidics-based time-resolved cell tracking, we measure the growth dynamics of individual cells of the diatom Thalassiosira pseudonana and show that cells growing mostly during the dark phase achieve rapid generation times (8 hours), dividing as fast as cells growing fully in light. We found that this remarkable ability of rapid growth in the dark is a consequence of the light history of both the cell and its parent cell, as light history controls the amount of photosynthetic energy the cell has stored in the form of the polysaccharide chrysolaminarin when entering the night. Furthermore, a mathematical model of this mechanism yields an up to 14% increase in the daily asynchronous population growth rate compared to growth without diel energy reserves when nutrients are non-limiting. This results in an up to 17-fold predicted increase in cell abundance over a typical 10-day diatom bloom, the maximal growth advantage of heritable chrysolaminarin during exponential phases of growth. By directly demonstrating and quantifying the benefit of chrysolaminarin under different diel conditions, this work provides a mechanistic understanding of how heritable diel energy reserves contribute to the rapid growth of diatom populations in the ocean. Beyond the specific discovery of the role of heritable energy reserves, we anticipate that our experimental approach can be broadly utilized in phytoplankton research by providing a blueprint to study phytoplankton physiology and ecology at the single-cell level, revealing novel mechanisms normally obscured in bulk growth assays.

microbiology↗

bioRxiv2025

Risk-reward trade-off in motility endurance generates dichotomy in search strategies among copiotrophic marine bacteria

Coptiotrophic marine bacteria contribute significantly to carbon storage in the ocean by remineralizing organic carbon present in nutrient-rich hotspots amidst oligotrophic waters. Motility is both highly beneficial and costly in such environments, presenting copiotrophs with a risk-reward trade-off in search behavior. Here we studied the motility endurance of 26 marine isolates using video microscopy and cell tracking over two days of carbon starvation. We found that this cost-benefit trade-off results in a distinct dichotomy among marine bacteria: risk-averse copiotrophs ceased motility within hours ( limostatic), whereas risk-prone copiotrophs converted 10% of their biomass per day into energy to retain motility for the two days of observation ( limokinetic). We identified a genomic component of this dichotomy, sufficiently robust to predict the response of additional species with 83% accuracy and the prevalence of both strategies in the ocean. This dichotomy can facilitate the incorporation of the bacterial contribution in ocean carbon cycle models.

microbiology↗

bioRxiv2025

Growth in Low Carbon Conditions Reveals Amino-Acid-Coupled Iron Uptake

Bacteria in nature encounter substrates at widely varying concentrations, yet studies of bacterial physiology have focused more on nutrient type than concentration, partly due to challenges in maintaining low concentrations. We developed a Millifluidic Continuous Culture Device (MCCD) to culture bacteria under precisely controlled nutrient conditions, including very low concentrations, in a manner suitable for proteomic analysis. Using the MCCD, we cultured Escherichia coli with a mixture of amino acids as the sole carbon source at three concentrations supporting growth rates spanning a fivefold range. Surprisingly, at the lowest concentration, cells exhibited proteomic signatures of iron shortage despite equal iron levels across conditions. We observed the uptake of labeled iron-histidine and iron-cysteine complexes, indicating that amino acids facilitated iron acquisition and that amino-acid-bound iron is bioavailable to E. coli. These findings reveal a previously unknown mechanism of bacterial iron acquisition that emerged under the flow imposed by the MCCD, which likely diluted the siderophore pool and reduced their efficacy. This work highlights the importance of studying bacterial physiology under low nutrient concentrations and demonstrates how physical conditions, such as flow, shape microbial nutrient acquisition strategies.

microbiology↗

bioRxiv2025

Bacteria induce an amoeboid phase in coccolithophores that persists after bloom collapse

Coccolithophores, who contribute approximately 1-10% of phytoplankton biomass, are crucial players in the oceans biogeochemical cycles. Significant contributions come from bloom-forming species including Gephyrocapsa huxleyi (formerly Emiliania huxleyi), which has served as a model system for investigating algal-bacterial and agal-viral interactions as well as algal fitness responses to environmental changes. Coccolithophores follow a biphasic lifecycle, existing as motile haploid and non-motile diploid phases. Here we characterize a third, amoeboid phase, using a combination of light, electron and phase amplitude modulation (PAM) microscopy. Using time-resolved imaging we captured the rapid morphological transition from a spherical haploid to an elongated motile amoeboid cell. Cell tracking revealed slower and more directional swimming compared to haploid cells. Amoeboid metamorphosis was triggered by exposure to bacteria, including strains isolated from G. huxleyi blooms and known G. huxleyi pathogens, but not by a range of classical phytoplankton stressors, including viral infection and oxidative stress. Further, the sub-population of haploids which switched to the amoeboid phase persisted past the rapid crash of the haploid population. This amoeboid phase was only observed in the primary bloom-forming coccolithophore species G. huxleyi and Geopharycapsa oceanica in stationary phase when exposed to high bacterial concentrations, typical of late-stage algal bloom events. Photophysiology of amoeboid cells was confirmed to be unaltered via PAM microscopy, indicating these cells are metabolically active. These findings highlight a previously uncharacterized morphotype in this important phytoplankton species and suggest that the amoeboid phase could be a bacteria-resistant morphotype following algal bloom collapse.

microbiology↗

bioRxiv2023

Polysaccharide breakdown products drive degradation-dispersal cycles of foraging bacteria through changes in metabolism and motility

Most of Earths biomass is composed of polysaccharides. During biomass decomposition, polysaccharides are degraded by heterotrophic bacteria as a nutrient and energy source and are thereby partly remineralized into CO2. As polysaccharides are heterogeneously distributed in nature, following the colonization and degradation of a polysaccharide hotspot the cells need to reach new polysaccharide hotspots. Even though many studies indicate that these degradation-dispersal cycles contribute to the carbon flow in marine systems, we know little about how cells alternate between polysaccharide degradation and motility, and which environmental factors trigger this behavioral switch. Here, we studied the growth of the marine bacterium Vibrio cyclitrophicus ZF270 on the abundant marine polysaccharide alginate, both in its soluble polymeric form as well as on its breakdown products. We used microfluidics coupled to time-lapse microscopy to analyze motility and growth of individual cells, and RNA sequencing to study associated changes in gene expression. We found that single cells grow at reduced rate on alginate until they form large groups that cooperatively break down the polymer. Exposing cell groups to digested alginate accelerates cell growth and changes the expression of genes involved in alginate degradation and catabolism, central metabolism, ribosomal biosynthesis, and transport. However, exposure to digested alginate also triggers cells to become motile and disperse from cell groups, proportionally increasing with the group size before the nutrient switch, and this is accompanied by high expression of genes involved in flagellar assembly, chemotaxis, and quorum sensing. The motile cells chemotax toward polymeric but not digested alginate, likely enabling them to find new polysaccharide hotspots. Overall, our findings reveal cellular mechanisms that might also underlie bacterial degradation-dispersal cycles, which influence the remineralization of biomass in marine environments. ImportancePolysaccharides, also known as glycans, are the most abundant form of biomass on Earth and understanding how they are degraded by microorganisms is essential for our understanding of the global carbon cycle and the storage and release of CO2 by natural systems. Although group formation is a common strategy used by bacterial cells to degrade ubiquitous polymeric growth substrates in nature, where nutrient hotspots are heterogeneously distributed, little is known about how cells prepare for dispersal from an exhausted nutrient source and re-initiate degradation of new nutrient patches. By quantifying growth, motility and chemotaxis of individual cells and comparing gene expression changes when populations were exposed to either polysaccharides or their degradation products in the form of digested polysaccharides, we show that bacterial cells alter their behavior when they experience a shift from polymeric to digested polysaccharides: After cells form groups during growth on polymers, the exposure to degradation products triggered cells to become motile, enabling dispersal from sessile cell groups. The chemotactic attraction of motile cells to polymeric alginate likely allows cells to move towards new polysaccharide hotspots. Our study sheds light on the cellular processes that drive bacterial growth and behavior during carbon remineralization, an important process resulting in CO2 release from natural systems.

microbiology↗

bioRxiv2022

Near-critical tuning of cooperativity revealed by spontaneous switching in a protein signalling array

Dynamic properties of allosteric complexes are crucial for cellular information processing. However, direct observations of allosteric switches have been limited to compact molecular assemblies. Here, we report in vivo FRET measurements of spontaneous discrete-level fluctuations in the activity of the Escherichia coli chemosensory array -- an extensive membrane-associated assembly comprising thousands of molecules. Finite-size scaling analysis of the temporal statistics by a two-dimensional conformational spread model revealed nearest-neighbor coupling strengths within 3% of the Ising second-order phase transition, indicating that chemosensory arrays are poised at criticality. Our analysis yields estimates for the intrinsic timescale of conformational changes (~ 10 ms) of allosteric units, and identifies near-critical tuning as a design principle for balancing the inherent tradeoff between response amplitude and response speed in higher-order signaling assemblies. One-setence summaryIn vivo measurements of protein signaling array fluctuations reveal an allosteric system poised at criticality.

biophysics↗

bioRxiv2021

Bacterial chemotaxis to saccharides is governed by a trade-off between sensing and uptake

To swim up gradients of nutrients, E. coli senses nutrient concentrations within its periplasm. For small nutrient molecules, periplasmic concentrations typically match extracellular concentrations. However, this is not necessarily the case for saccharides, such as maltose, which is transported into the periplasm via a specific porin. Previous observations have shown that under various conditions E. coli limits maltoporin abundance so that, for extracellular micromolar concentrations of maltose, there are predicted to be only nanomolar concentrations of free maltose in the periplasm. Thus, in the micromolar regime, the total uptake of maltose from the external environment into the cytoplasm is limited not by the abundance of cytoplasmic transport proteins but by the abundance of maltoporins. Here we present results from experiments and modeling showing that this porin-limited transport enables E. coli to sense micromolar gradients of maltose despite having a high-affinity ABC transport system that is saturated at these micromolar levels. We used microfluidic assays to study chemotaxis of E. coli in various gradients of maltose and methyl-aspartate and leveraged our experimental observations to develop a mechanistic transport-and-sensing chemotaxis model. Incorporating this model into agent-based simulations, we discover a trade-off between uptake and sensing: although high-affinity transport enables higher uptake rates at low nutrient concentrations, it severely limits dynamic sensing range. We thus propose that E. coli may limit periplasmic uptake to increase its chemotactic sensitivity, enabling it to use maltose as an environmental cue. Statement of SignificanceBacterial chemotaxis is among the best-studied systems in biology and is paradigmatic of the mechanisms used by cells to link sensory inputs with regulated responses, thus providing insight into the ecological basis of cellular physiology. Here we present a mechanistic chemotaxis model that describes how the regulation of the transport of a sugar into and out of the cells periplasm affects the cells motile response to that sugar. Based on observations from population-level chemotaxis assays, we uncover an ecologically relevant trade-off between sensing and uptake. The general finding of this work is that, while high-affinity transport allows for higher uptake rates, it can severely limit the cells dynamic sensing range.

microbiology↗