FliL emerged as a modulator of flagellar motor function in several bacterial species, but its function in Pseudomonas aeruginosa was unknown. Here, by performing single-motor studies using a bead assay, we elucidated its effects on the flagellar motor in P. aeruginosa .
ABSTRACT 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. IMPORTANCE 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.
Bacteria inhabit complex environments rich in macromolecular polymers that exhibit viscoelastic properties. While the influence of viscoelasticity on bacterial swimming is recognized, its impact on chemotaxis-a critical behavior for bacterial survival and colonization-remains elusive. In this study, we employed a microfluidic device to establish attractant gradients and observed the chemotactic behavior of
Photoactivatable fluorescent proteins (PA-FPs) are widely used in photoactivated localization microscopy (PALM), and information about their blinking properties is important for precise assembling of PALM images and for quantitative molecular counting with PALM. As the photophysical properties of fluorescent proteins depend sensitively on the local physiochemical environment, we developed a series of procedures to characterize the properties of the PA-FPs in fixed cells. As an example, we characterized the properties of Dendra2 in Escherichia coli and found that they are appreciably different from those in vitro. This highlights the importance of characterizing the photophysical properties of PA-FPs under the same experimental conditions as subsequent PALM imaging or molecular counting experiments. Other PA-FPs could be characterized using this strategy.
Cells of Escherichia coli are able to swim up gradients of chemical attractants by modulating the direction of rotation of their flagellar motors, which spin alternately clockwise (CW) and counterclockwise (CCW). Rotation in either direction has been thought to be symmetric and exhibit the same torques and speeds. The relationship between torque and speed is one of the most important measurable characteristics of the motor, used to distinguish specific mechanisms of motor rotation. Previous measurements of the torque–speed relationship have been made with cells lacking the response regulator CheY that spin their motors exclusively CCW. In this case, the torque declines slightly up to an intermediate speed called the “knee speed” after which it falls rapidly to zero. This result is consistent with a “power-stroke” mechanism for torque generation. Here, we measure the torque–speed relationship for cells that express large amounts of CheY and only spin their motors CW. We find that the torque decreases linearly with speed, a result remarkably different from that for CCW rotation. We obtain similar results for wild-type cells by reexamining data collected in previous work. We speculate that CCW rotation might be optimized for runs, with higher speeds increasing the ability of cells to sense spatial gradients, whereas CW rotation might be optimized for tumbles, where the object is to change cell trajectories. But why a linear torque–speed relationship might be optimum for the latter purpose we do not know.
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
The flagellar motor drives the rotation of flagellar filaments, propeling the swimming of flagellated bacteria. The maximum torque the motor generates, the stall torque, is a key characteristics of the motor function. Direct measurements of the stall torque carried out three decades ago suffered from large experimental uncertainties, and subsequently there were only indirect measurements. Here, we applied magnetic tweezer to directly measure the stall torque in E. coli . We precisely calibrated the torsional stiffness of the magnetic tweezer, and performed motor resurrection experiments at stall, accomplishing a precise determination of the stall torque per torque-generating unit (stator unit). From our measurements, each stator passes 2 protons per step, indicating a tight coupling between motor rotation and proton flux.
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