Asymmetric swimming pattern of Vibrio alginolyticus cells with single polar flagella

The swimming pattern of bacteria with single polar flagella has usually been described as "run and reverse". We observed the swimming traces of monotrichously flagellated Vibrio alginolyticus cells and examined the relationship between the swimming pattern and the sense of progress. Swimming in regions other than a solid surface was confirmed to be linear run and reverse. Near a solid surface, the traces consisted of "run and arc"; the cells were found to curve sharply during backward swimming, while they progressed linearly during forward swimming. The "run and arc" swimming pattern may play an important role in the chemotaxis strategy of marine bacteria at solid surfaces.     

An Element of Determinism in a Stochastic Flagellar Motor Switch

Marine bacterium Vibrio alginolyticus uses a single polar flagellum to navigate in an aqueous environment. Similar to Escherichia coli cells, the polar flagellar motor has two states; when the motor is counter-clockwise, the cell swims forward and when the motor is clockwise, the cell swims backward. V. alginolyticus also incorporates a direction randomization step at the start of the forward swimming interval by flicking its flagellum. To gain an understanding on how the polar flagellar motor switch is regulated, distributions of the forward Δ_f and backward Δ_b intervals are investigated herein. We found that the steady-state probability density functions, P(Δ_f) and P(Δ_b), of freely swimming bacteria are strongly peaked at a finite time, suggesting that the motor switch is not Poissonian. The short-time inhibition is sufficiently strong and long lasting, i.e., several hundred milliseconds for both intervals, which is readily observed and characterized. Treating motor reversal dynamics as a first-passage problem, which results from conformation fluctuations of the motor switch, we calculated P(Δ_f) and P(Δ_b) and found good agreement with the measurements.     

A fluid-dynamic interpretation of the asymmetric motion of singly flagellated bacteria swimming close to a boundary

The singly flagellated bacterium, Vibrio alginolyticus, moves forward and backward by alternating the rotational direction of its flagellum. The bacterium has been observed retracing a previous path almost exactly and swimming in a zigzag pattern. In the presence of a boundary, however, the motion changes significantly, to something closer to a circular trajectory. Additionally, when the cell swims close to a wall, the forward and backward speeds differ noticeably. This study details a boundary element model for the motion of a bacterium swimming near a rigid boundary and the results of numerical analyses conducted using this model. The results reveal that bacterium motion is apparently influenced by pitch angle, i.e., the angle between the boundary and the swimming direction, and that forward motion is more stable than backward motion with respect to pitching of the bacterium. From these results, a set of diagrammatic representations have been created that explain the observed asymmetry in trajectory and speed between the forward and backward motions. For forward motion, a cell moving parallel to the boundary will maintain this trajectory. However, for backward motion, the resulting trajectory depends upon whether the bacterium is approaching or departing the boundary. Fluid-dynamic interactions between the flagellum and the boundary vary with cell orientation and cause peculiarities in the resulting trajectories.     

Two different stator systems drive a single polar flagellum in Shewanella oneidensis MR-1

The Gram-negative metal ion-reducing bacterium Shewanella oneidensis MR-1 is motile by means of a single polar flagellum. We identified two potential stator systems, PomAB and MotAB, each individually sufficient as a force generator to drive flagellar rotation. Physiological studies indicate that PomAB is sodium-dependent while MotAB is powered by the proton motive force. Flagellar function mainly depends on the PomAB stator; however, the presence of both stator systems under low-sodium conditions results in a faster swimming phenotype. Based on stator homology analysis we speculate that MotAB has been acquired by lateral gene transfer as a consequence of adaptation to a low-sodium environment. Expression analysis at the single cell level showed that both stator systems are expressed simultaneously. An active PomB–mCherry fusion protein effectively localized to the flagellated cell pole in 70–80% of the population independent of sodium concentrations. In contrast, polar localization of MotB–mCherry increased with decreasing sodium concentrations. In the absence of the Pom stator, MotB–mCherry localized to the flagellated cell pole independently of the sodium concentration but was rapidly displaced upon expression of PomAB. We propose that selection of the stator occurs at the level of protein localization in response to sodium concentrations.     

Multiple CheY Homologs Control Swimming Reversals and Transient Pauses in Azospirillum brasilense

Chemotaxis, together with motility, helps bacteria foraging in their habitat. Motile bacteria exhibit a variety of motility patterns, often controlled by chemotaxis, to promote dispersal. Motility in many bacteria is powered by a bidirectional flagellar motor. The flagellar motor has been known to briefly pause during rotation because of incomplete reversals or stator detachment. Transient pauses were previously observed in bacterial strains lacking CheY, and these events could not be explained by incomplete motor reversals or stator detachment. Here, we systematically analyzed swimming trajectories of various chemotaxis mutants of the monotrichous soil bacterium, Azospirillum brasilense. Like other polar flagellated bacterium, the main swimming pattern in A. brasilense is run and reverse. A. brasilense also uses run-pauses and putative run-reverse-flick-like swimming patterns, although these are rare events. A. brasilense mutant derivatives lacking the chemotaxis master histidine kinase, CheA4, or the central response regulator, CheY7, also showed transient pauses. Strikingly, the frequency of transient pauses increased dramatically in the absence of CheY4. Our findings collectively suggest that reversals and pauses are controlled through signaling by distinct CheY homologs, and thus are likely to be functionally important in the lifestyle of this soil organism.     

Difference in bacterial motion between forward and backward swimming caused by the wall effect

A bacterial cell that has a single polar flagellum alternately repeats forward swimming, in which the flagellum pushes the cell body, and backward swimming, in which the flagellum pulls the cell body. We have reported that the backward swimming speeds of Vibrio alginolyticus are on average greater than the forward swimming speeds. In this study, we quantitatively measured the shape of the trajectory as well as the swimming speed. The trajectory shape in the forward mode was almost straight, whereas that in the backward mode was curved. The same parameters were measured at different distances from a surface. The difference in the motion characteristics between swimming modes was significant when a cell swam near a surface. In contrast, the difference was indistinguishable when a cell swam >60 microm away from any surfaces. In addition, a cell in backward mode tended to stay near the surface longer than a cell in forward mode. This wall effect on the bacterial motion was independent of chemical modification of the glass surface. The macroscopic behavior is numerically simulated on the basis of experimental results and the significance of the phenomenon reported here is discussed.     

A Non-Poissonian Flagellar Motor Switch Increases Bacterial Chemotactic Potential

We investigate bacterial chemotactic strategies using run-tumble and run-reverse-flick motility patterns. The former is typically observed in enteric bacteria such as Escherichia coli and Salmonella and the latter was recently observed in the marine bacteria Vibrio alginolyticus and is possibly exhibited by other polar flagellated species. It is shown that although the three-step motility pattern helps the bacterium to localize near hot spots, an exploitative behavior, its exploratory potential in short times can be significantly enhanced by employing a non-Poissonian regulation scheme for its flagellar motor switches.     

Difference between forward and backward swimming speeds of the single polar-flagellated bacterium, Vibrio alginolyticus

The forward and backward swimming speeds and periods of a Vibrio alginolyticus strain that has a single polar flagellum were measured. The backward swimming speeds were 1.5 times greater than the forward ones on average and the average period of backward swimming was shorter than forward swimming. However, the swimming speed and period were not correlated. Similar results were obtained for a mutant that has a 1.6 times longer flagellum on average.     

Transitioning to confined spaces impacts bacterial swimming and escape response

Symbiotic bacteria often navigate complex environments before colonizing privileged sites in their host organism. Chemical gradients are known to facilitate directional taxis of these bacteria, guiding them toward their eventual destination. However, less is known about the role of physical features in shaping the path the bacteria take and defining how they traverse a given space. The flagellated marine bacterium Vibrio fischeri, which forms a binary symbiosis with the Hawaiian bobtail squid, Euprymna scolopes, must navigate tight physical confinement during colonization, squeezing through a tissue bottleneck constricting to ～2 μm in width on the way to its eventual home. Using microfluidic in vitro experiments, we discovered that V. fischeri cells alter their behavior upon entry into confined space, straightening their swimming paths and promoting escape from confinement. Using a computational model, we attributed this escape response to two factors: reduced directional fluctuation and a refractory period between reversals. Additional experiments in asymmetric capillary tubes confirmed that V. fischeri quickly escape from confined ends, even when drawn into the ends by chemoattraction. This avoidance was apparent down to a limit of confinement approaching the diameter of the cell itself, resulting in a balance between chemoattraction and evasion of physical confinement. Our findings demonstrate that nontrivial distributions of swimming bacteria can emerge from simple physical gradients in the level of confinement. Tight spaces may serve as an additional, crucial cue for bacteria while they navigate complex environments to enter specific habitats.     

Bacterial chemotaxis in an optical trap

An optical trapping technique is implemented to investigate the chemotactic behavior of a marine bacterial strain Vibrio alginolyticus. The technique takes the advantage that the bacterium has only a single polar flagellum, which can rotate either in the counter-clock-wise or clock-wise direction. The two rotation states of the motor can be readily and instantaneously resolved in the optical trap, allowing the flagellar motor switching rate S(t) to be measured under different chemical stimulations. In this paper the focus will be on the bacterial response to an impulsive change of chemoattractant serine. Despite different propulsion apparati and motility patterns, cells of V. alginolyticus apparently use a similar response as Escherichia coli to regulate their chemotactic behavior. Specifically, we found that the switching rate S(t) of the bacterial motor exhibits a biphasic behavior, showing a fast initial response followed by a slow relaxation to the steady-state switching rate S0. The measured S(t) can be mimicked by a model that has been recently proposed for chemotaxis in E. coli. The similarity in the response to the brief chemical stimulation in these two different bacteria is striking, suggesting that the biphasic response may be evolutionarily conserved. This study also demonstrated that optical tweezers can be a useful tool for chemotaxis studies and should be applicable to other polarly flagellated bacteria.     

Kinematics of swimming in two burrowing anguilliform fishes

Anguilliform or eel-like fishes are typically bottom dwellers, some of which are specialized burrowers. Although specializations for burrowing are predicted to affect the kinematics of swimming, it remains unknown to what extent this is actually the case. Here we examine swimming kinematics and efficiency of two burrowing anguilliform species, Pisodonophis boro and Heteroconger hassi, with different degrees of specialization for burrowing. Our data suggest that differences in the swimming kinematics may indeed be related to the differences in burrowing specialization and style between both species. The resemblance between the swimming kinematics of P. boro and previously published data for Anguilla anguilla and Anguilla rostrata may be linked with the relatively limited burrowing specialization of P. boro and suggests an overall stereotypy in anguilliform forward-swimming patterns. The body of H. hassi, in contrast, is more specialized for tail-first burrowing and backward swimming bears a striking resemblance to the backward burrowing motions observed in this species. These motions differ significantly from backward swimming in Anguilla and in P. boro. The kinematics of forward swimming are, however, comparable across species. Thus, our data suggest that specializations for burrowing may affect swimming kinematics in anguilliform fishes, but also that forward swimming and burrowing are not necessarily incompatible. Future studies comparing the kinematics and mechanics of burrowing in these and other anguilliform fishes are needed to better understand how specializations for burrowing constrain backward swimming in H. hassi.     

Kinematic Comparison of Forward and Backward Swimming and Maneuvering in a Self-Propelled Sub-Carangiform Robotic Fish

We make a thorough kinematic comparison of forward and backward swimming and maneuvering on a self-propelled robot platform that uses sub-carangifbrm swimming as the primary propulsor. An improved Central Pattern Generator （CPG） model allowing free adjustment of phase relationship and directional bias is employed to achieve flexible swimming and smooth transition. Considering the characteristics of forward swimming in carangiform fish and backward swimming in anguilliform fish, various backward swimming patterns for the sub-carangiform robotic fish are suitably created by reversing the direction of propagating propulsive waves. Through a combined use of the CPG control and closed-loop swimming direction control strategy, flexible and precise turning maneuvers in both forward and backward swimming are implemented and compared. By contrast with forward swimming, backward swimming requires a higher frequency or an increased lateral displacement to reach the same relative swimming speed. Noticeably, the phase difference shows a greater impact on forward swimming than on backward swimming. Our observations also indicate that the robotic fish achieves a larger turning rate in forward maneuvering than in backward maneuvering, yet these two maneuvers display comparable turning precision.     

A Phase Variant of Azospirillum lipoferum Lacks a Polar Flagellum and Constitutively Expresses Mechanosensing Lateral Flagella

Flagellation of a nonswimming variant of the mixed flagellated bacterium Azospirillum lipoferum 4B was characterized by electron microscopy, and polyclonal antibodies were raised against polar and lateral flagellins. The variant cells lacked a polar flagellum due to a defect in flagellin synthesis and constitutively expressed lateral flagella. The variant cells were able to respond to conditions that restricted the rotation of lateral flagella by producing more lateral flagella, suggesting that the lateral flagella, as well as the polar flagellum, are mechanosensing.     

Attachment of Vibrio alginolyticus to Glass Surfaces Is Dependent on Swimming Speed

The attachment of Vibrio alginolyticus to glass surfaces was investigated with special reference to the swimming speed due to the polar flagellum. This bacterium has two types of flagella, i.e., one polar flagellum and numerous lateral flagella. The mutant YM4, which possesses only the polar flagellum, showed much faster attachment than the mutant YM18, which does not possess flagella, indicating that the polar flagellum plays an important role. The attachment of YM4 was dependent on Na⁺ concentration and was specifically inhibited by amiloride, an inhibitor of polar flagellum rotation. These results are quite similar to those for swimming speed obtained under the same conditions. Observations with other mutants showed that chemotaxis is not critical and that the flagellum does not act as an appendage for attachment. From these results, it is concluded that the attachment of V. alginolyticus to glass surfaces is dependent on swimming speed.     

Molecular Adsorption Steers Bacterial Swimming at the Air/Water Interface

Microbes inhabiting Earth have adapted to diverse environments of water, air, soil, and often at the interfaces of multiple media. In this study, we focus on the behavior of Caulobacter crescentus, a singly flagellated bacterium, at the air/water interface. Forward swimming C. crescentus swarmer cells tend to get physically trapped at the surface when swimming in nutrient-rich growth medium but not in minimal salt motility medium. Trapped cells move in tight, clockwise circles when viewed from the air with slightly reduced speed. Trace amounts of Triton X100, a nonionic surfactant, release the trapped cells from these circular trajectories. We show, by tracing the motion of positively charged colloidal beads near the interface that organic molecules in the growth medium adsorb at the interface, creating a high viscosity film. Consequently, the air/water interface no longer acts as a free surface and forward swimming cells become hydrodynamically trapped. Added surfactants efficiently partition to the surface, replacing the viscous layer of molecules and reestablishing free surface behavior. These findings help explain recent similar studies on Escherichia coli, showing trajectories of variable handedness depending on media chemistry. The consistent behavior of these two distinct microbial species provides insights on how microbes have evolved to cope with challenging interfacial environments.     

Chemoreception and the deformationof the active space in freely swimming copepods: a numerical study

A three-dimensional alga-tracking, chemical advection–diffusionmodel was used to calculate the deformation of the active spacesurrounding an alga entrained within the flow field around afreely swimming copepod. From the model, the advance warningtime resulting from the copepod's chemo-reception of the entrainedalga was quantified, and copepod chemoreception capability comparedfor several different swimming behaviors: hovering in the water,swimming slowly (swimming upward, swimming backward and swimmingforward), swimming fast (swimming upward, swimming backwardand swimming forward) and sinking (with the anterior pointingupward or downward). The results show that when it hovers orswims slowly, a copepod can use chemoreception to remotely detectindividual algae entrained by the flow field around itself.In contrast, a fast-swimming copepod is not able to rely onchemoreception to remotely detect individual algae. The possibilityof a free-sinking copepod using chemoreception to detect algalparticles is also indicated. It is shown that advection by thefluid motion dominates over diffusion in transporting the chemicalsignals inside the active space to the location of a copepod'schemoreceptors. The feeding current structure for a hoveringcopepod is described. It is suggested that the feeding currentstructure and re-routing or re-orienting response by a copepodin response to its antennule or other cephalic appendage inputsallow the copepod to capture the food particles that would otherwisepass outside its capture area and increase the amount of foodcaptured.     

Na(+)- and H(+)-dependent motility in the coral pathogen Vibrio shilonii

In this work, we analyzed motility and the flagellar systems of the marine bacterium Vibrio shilonii. We show that this bacterium produces lateral flagella when seeded on soft agar plates at concentrations of 0.5% or 0.6%. However, at agar concentrations of 0.7%, cells become round and lose their flagella. The sodium channel blocker amiloride inhibits swimming of V. shilonii with the sheathed polar flagellum, but not swarming with lateral flagella. We also isolated and characterized the filament–hook–basal body of the polar flagellum. The proteins in this structure were analyzed by MS. Eight internal sequences matched with known flagellar proteins. The comparison of these sequences with the protein database from the complete genome of V. shilonii allows us to conclude that some components of the polar flagellum are encoded in two different clusters of flagellar genes, suggesting that this bacterium has a complex flagellar system, more complex possibly than other Vibrio species reported so far.     

The Measurement of Swimming Velocity of Vibrio cholerae and Pseudomonas aeruginosa Using the Video Tracking Method

The swimming velocities of two monotrichous flagellated bacteria were measured by a computer-assisted video tracking method. Tracing the moving path of the individual bacterium revealed that the bacterial cell did not swim continuously in a straight direction, but frequently changed swimming direction and velocity. The average swimming velocities calculated from the 3-sec path were 75.4 ±9.4 μm/sec in four strains of Vibrio cholerae and 513 ±8.4 μm/sec in five strains of Pseudomonas aeruginosa. These results suggest that V. cholerae swim faster than P. aeruginosa at 30 C in nutrient broth. This method is useful for a detailed analysis of bacterial movement and moving patterns in different environmental conditions.     

Locomotion of Gymnarchus Niloticus： Experiment and Kinematics

In addition to forward undulatory swimming, Gymnarchus niloticus can swim via undulations of the dorsal fin while the body axis remains straight; furthermore, it swims forward and backward in a similar way, which indicates that the undulation of the dorsal fin can simultaneously provide bidirectional propulsive and maneuvering forces with the help of the tail fin. A high-resolution Charge-Coupled Device （CCD） imaging camera system is used to record kinematics of steady swimming as well as maneuvering in G. niloticus. Based on experimental data, this paper discusses the kinematics （cruising speed, wave speed, cycle frequency, amplitude, lateral displacement） of forward as well as backward swimming and maneuvering. During forward swimming, the propulsive force is generated mainly by undulations of the dorsal fin while the body axis remains straight. The kinematic parameters （wave speed, wavelength, cycle frequency, amplitude） have statistically significant correlations with cruising speed. In addition, the yaw at the head is minimal during steady swimming. From experimental data, the maximal lateral displacement of head is not more than 1% of the body length, while the maximal lateral displacement of the whole body is not more than 5% of the body length. Another important feature is that G. niloticus swims backwards using an undulatory mechanism that resembles the forward undulatory swimming mechanism. In backward swimming, the increase of lateral displacement of the head is comparatively significant; the amplitude profiles of the propulsive wave along the dorsal fin are significantly different from those in forward swimming. When G. niloticus does fast maneuvering, its body is first bent into either a C shape or an S shape, then it is rapidly unwound in a travelling wave fashion. It rarely maneuvers without the help of the tail fin and body bending.     

Dephosphorylation of inner arm 1 is associated with ciliary reversals in Tetrahymena thermophila

In many organisms, depolarizing stimuli cause an increase in intraciliary Ca2+, which results in reversal of ciliary beat direction and backward swimming. The mechanism by which an increase in intraciliary Ca2+ causes ciliary reversal is not known. Here we show that Tetrahymena cells treated with okadaic acid or cantharidin to inhibit protein phosphatases do not swim backwards in response to depolarizing stimuli. We also show that both okadaic acid and cantharidin inhibit backward swimming in reactivated, extracted cell models treated with Ca2+. In contrast, treatment of whole cells or extracted cell models with protein kinase inhibitors has no effect on backward swimming. These results suggest that a component of the axonemal machinery is dephosphorylated during ciliary reversal. The phosphorylation state of inner arm dynein 1 (I1) was determined before and after cells were exposed to depolarizing conditions that induce ciliary reversal. An I1 intermediate chain is phosphorylated in forward swimming cells but is dephosphorylated in cells treated with a depolarizing stimulus. Our results suggest that dephosphorylation of Tetrahymena inner arm dynein 1 may be an essential part of the mechanism of ciliary reversal in response to increased intraciliary Ca2+.