Bacteria in biofilms secrete potassium ions to attract free swimming cells. However, the basis of chemotaxis to potassium remains poorly understood. Here, using a microfluidic device, we found that Escherichia coli can rapidly accumulate in regions of high potassium concentration on the order of millimoles. Using a bead assay, we measured the dynamic response of individual flagellar motors to stepwise changes in potassium concentration, finding that the response resulted from the chemotaxis signaling pathway. To characterize the chemotactic response to potassium, we measured the dose-response curve and adaptation kinetics via an Förster resonance energy transfer (FRET) assay, finding that the chemotaxis pathway exhibited a sensitive response and fast adaptation to potassium. We further found that the two major chemoreceptors Tar and Tsr respond differently to potassium. Tar receptors exhibit a biphasic response, whereas Tsr receptors respond to potassium as an attractant. These different responses were consistent with the responses of the two receptors to intracellular pH changes. The sensitive response and fast adaptation allow bacteria to sense and localize small changes in potassium concentration. The differential responses of Tar and Tsr receptors to potassium suggest that cells at different growth stages respond differently to potassium and may have different requirements for potassium.

Solid surfaces submerged in liquid in natural environments alter bacterial swimming behavior and serve as platforms for bacteria to form biofilms. In the initial stage of biofilm formation, bacteria detect surfaces and increase the intracellular level of the second messenger c-di-GMP, leading to a reduction in swimming speed. The impact of this speed reduction on bacterial surface swimming remains unclear. In this study, we utilized advanced microscopy techniques to examine the effect of swimming speed on bacterial surface swimming behavior. We found that a decrease in swimming speed reduces the cell-surface distance and prolongs the surface trapping time. Both these effects would enhance bacterial surface sensing and increase the likelihood of cells adhering to the surface, thereby promoting biofilm formation. We also examined the surface-escaping behavior of wild-type Escherichia coli and Pseudomonas aeruginosa, noting distinct surface-escaping mechanisms between the two bacterial species.
OBJECTIVE: In the early phase of biofilm formation, bacteria identify surfaces and increase the intracellular level of the second messenger c-di-GMP, resulting in a decrease in swimming speed. Here, we utilized advanced microscopy techniques to investigate the impact of swimming speed on bacterial surface swimming, focusing on Escherichia coli and Pseudomonas aeruginosa. We found that an increase in swimming speed led to an increase in the radius of curvature and a decrease in surface detention time. These effects were explained through hydrodynamic modeling as a result of an increase in the cell-surface distance with increasing swimming speed. We also observed distinct surface-escaping mechanisms between the two bacterial species. Our study suggests that a decrease in swimming speed could enhance the likelihood of cells adhering to the surface, promoting biofilm formation. This sheds light on the role of reduced swimming speed in the transition from motile to sedentary bacterial lifestyles.

Type IVa pili (T4aP) are ubiquitous cell surface filaments important for surface motility, adhesion to surfaces, DNA uptake, biofilm formation, and virulence. T4aP are built from thousands of copies of the major pilin subunit and tipped by a complex composed of minor pilins and in some systems also the PilY1 adhesin. While major pilins of structurally characterized T4aP have lengths of <165 residues, the major pilin PilA of Myxococcus xanthus is unusually large with 208 residues. All major pilins have a conserved N-terminal domain and a variable C-terminal domain, and the additional residues of PilA are due to a larger C-terminal domain. We solved the structure of the M. xanthus T4aP (T4aPMx) at a resolution of 3.0 Å using cryo-EM. The T4aPMx follows the structural blueprint of other T4aP with the pilus core comprised of the interacting N-terminal α1-helices, while the globular domains decorate the T4aP surface. The atomic model of PilA built into this map shows that the large C-terminal domain has more extensive intersubunit contacts than major pilins in other T4aP. As expected from these greater contacts, the bending and axial stiffness of the T4aPMx is significantly higher than that of other T4aP and supports T4aP-dependent motility on surfaces of different stiffnesses. Notably, T4aPMx variants with interrupted intersubunit interfaces had decreased bending stiffness, pilus length, and strongly reduced motility. These observations support an evolutionary scenario whereby the large major pilin enables the formation of a rigid T4aP that expands the environmental conditions in which the T4aP system functions.

The environmental dissemination of extracellular antibiotic resistance genes (eARGs) in wastewater and natural water bodies has aroused growing ecological concerns. The coexisting chemical pollutants in water are known to markedly affect the eARGs transfer behaviors of the environmental microbial community, but the detailed interactions and specific impacts remain elusive so far. Here, we revealed a concentration-dependent impact of dimethyl phthalate (DMP) and several other types of phthalate esters (common water pollutants released from plastics) on the natural transformation of eARGs. The DMP exposure at an environmentally relevant concentration (10 μg/L) resulted in a 4.8-times raised transformation frequency of Acinetobacter baylyi but severely suppressed the transformation at a high concentration (1000 μg/L). The promotion by low-concentration DMP was attributed to multiple mechanisms, including increased bacterial mobility and membrane permeability to facilitate eARGs uptake and improved resistance of the DMP-bounded eARGs (via noncovalent interaction) to enzymatic degradation (with suppressed DNase activity). Similar promoting effects of DMP on the eARGs transformation were also found in real wastewater and biofilm systems. In contrast, higher-concentration DMP suppressed the eARGs transformation by disrupting the DNA structure. Our findings highlight a potentially underestimated eARGs spreading in aquatic environments due to the impacts of coexisting chemical pollutants and deepen our understanding of the risks of biological-chemical combined pollution in wastewater and environmental water bodies.

We investigate the motility of B. subtilis under different degrees of confinement induced by transparent porous hydrogels. The dynamical behavior of the bacteria at short times is linked to characteristic parameters describing the hydrogel porosity. Mean squared displacements (MSDs) reveal that the run-and-tumble dynamics of unconfined B. subtilis progressively turns into sub-diffusive motion with increasing confinement. Correspondingly, the median instantaneous velocity of bacteria decreases and becomes more narrowly distributed, while the reorientation rate increases and reaches a plateau value. Analyzing single-trajectories, we show that the average dynamical behavior is the result of complex displacements, in which active, diffusive and sub-diffusive segments coexist. For small and moderate confinements, the number of active segments reduces, while the diffusive and sub-diffusive segments increase. The alternation of sub-diffusion, diffusion and active motion along the same trajectory can be described as a hopping ad trapping motion, in which hopping events correspond to displacements with an instantaneous velocity exceeding the corresponding mean value along a trajectory. Different from previous observations, escape from local trapping occurs for B. subtilis through active runs but also diffusion. Interestingly, the contribution of diffusion is maximum at intermediate confinements. At sufficiently long times transport coefficients estimated from the experimental MSDs under different degrees of confinement can be reproduced using a recently proposed hopping and trapping model. Finally, we propose a quantitative relationship linking the median velocity of confined and unconfined bacteria through the characteristic confinement length of the hydrogel matrix. Our work provides new insights for the bacterial motility in complex media that mimic natural environments and are relevant to important problems like sterilization, water purification, biofilm formation, membrane permeation and bacteria separation.

Swarming motility is a type of movement used by pathogenic flagellated bacteria as virulence factor to colonize surfaces and cause damage to the host. Vibrio parahaemolyticus is a pathogenic flagellated bacterium that increases its virulence by switching from swimmer to swarming cells. The hosts of pathogenic V. parahaemolyticus include farmed shrimp. Therefore, methods to detect and quantify this movement are important to control shrimp diseases caused by pathogenic V. parahaemolyticus strains. We developed an optimized swarming motility assay by identifying the most optimal type of agar, and drying time of the culture medium, agar concentration and volume of the bacterial culture to achieve the fastest swarming motility during the migration of V. parahaemolyticus on Petri dishes during a 24-hour incubation period. The method includes data analysis that could be used as a tool to identify potential anti-virulence products by comparing the slopes of the linearized diameters of the swarming halos of bacteria treated with the products, as they migrate on Petri dishes over a 24-hour incubation period. Here we report:•A simple method for detection and quantification of swarming motility halos of V. parahaemolyticus bacteria.•A method that could be used as a tool to identify potential anti-virulence products.

Adjusting motility patterns according to environmental cues is important for bacterial survival. Myxococcus xanthus, a bacterium moving on surfaces by gliding and twitching mechanisms, modulates the reversal frequency of its front-back polarity in response to mechanical cues like substrate stiffness and cell-cell contact. In this study, we propose that M. xanthus\'s gliding machinery senses environmental mechanical cues during force generation and modulates cell reversal accordingly. To examine our hypothesis, we expand an existing mathematical model for periodic polarity reversal in M. xanthus, incorporating the experimental data on the intracellular dynamics of the gliding machinery and the interaction between the gliding machinery and a key polarity regulator. The model successfully reproduces the dependence of cell reversal frequency on substrate stiffness observed in M. xanthus gliding. We further propose reversal control networks between the gliding and twitching motility machineries to explain the opposite reversal responses observed in wild type M. xanthus cells that possess both motility mechanisms. These results provide testable predictions for future experimental investigations. In conclusion, our model suggests that the gliding machinery in M. xanthus can function as a mechanosensor, which transduces mechanical cues into a cell reversal signal.

To swim through a viscous fluid, a flagellated bacterium must overcome the fluid drag on its body by rotating a flagellum or a bundle of multiple flagella. Because the drag increases with the size of bacteria, it is expected theoretically that the swimming speed of a bacterium inversely correlates with its body length. Nevertheless, despite extensive research, the fundamental size-speed relation of flagellated bacteria remains unclear with different experiments reporting conflicting results. Here, by critically reviewing the existing evidence and synergizing our own experiments of large sample sizes, hydrodynamic modeling, and simulations, we demonstrate that the average swimming speed of Escherichia coli, a premier model of peritrichous bacteria, is independent of their body length. Our quantitative analysis shows that such a counterintuitive relation is the consequence of the collective flagellar dynamics dictated by the linear correlation between the body length and the number of flagella of bacteria. Notably, our study reveals how bacteria utilize the increasing number of flagella to regulate the flagellar motor load. The collective load sharing among multiple flagella results in a lower load on each flagellar motor and therefore faster flagellar rotation, which compensates for the higher fluid drag on the longer bodies of bacteria. Without this balancing mechanism, the swimming speed of monotrichous bacteria generically decreases with increasing body length, a feature limiting the size variation of the bacteria. Altogether, our study resolves a long-standing controversy over the size-speed relation of flagellated bacteria and provides insights into the functional benefit of multiflagellarity in bacteria.

TmaR, the only known pole-localizer protein in Escherichia coli, was shown to cluster at the cell poles and control localization and activity of the major sugar regulator in a tyrosine phosphorylation-dependent manner. Here, we show that TmaR assembles by phase separation (PS) via heterotypic interactions with RNA in vivo and in vitro. An unbiased automated mutant screen combined with directed mutagenesis and genetic manipulations uncovered the importance of a predicted nucleic-acid-binding domain, a disordered region, and charged patches, one containing the phosphorylated tyrosine, for TmaR condensation. We demonstrate that, by protecting flagella-related transcripts, TmaR controls flagella production and, thus, cell motility and biofilm formation. These results connect PS in bacteria to survival and provide an explanation for the linkage between metabolism and motility. Intriguingly, a point mutation or increase in its cellular concentration induces irreversible liquid-to-solid transition of TmaR, similar to human disease-causing proteins, which affects cell morphology and division.

OBJECTIVE: Dickeya fangzhongdai is a newly identified plant bacterial pathogen with a wide host range. A clear understanding of the cell-to-cell communication systems that modulate the bacterial virulence is of key importance for elucidating its pathogenic mechanisms and for disease control. In this study, we present evidence that putrescine molecules from the pathogen and host plants play an essential role in regulating the bacterial virulence. The significance of this study is in (i) demonstrating that putrescine signaling system regulates D. fangzhongdai virulence mainly through modulating the bacterial motility and production of PCWD enzymes, (ii) outlining the signaling and regulatory mechanisms with which putrescine signaling system modulates the above virulence traits, and (iii) validating that D. fangzhongdai could use both arginine and ornithine pathways to synthesize putrescine signals. To our knowledge, this is the first report to show that putrescine signaling system plays a key role in modulating the pathogenicity of D. fangzhongdai.