Flagellar polymorphism-dependent bacterial swimming motility in a structured environment

Abstract

Most motile bacteria use supramolecular motility machinery called bacterial flagellum, which converts the chemical energy gained from ion flux into mechanical rotation. Bacterial cells sense their external environment through a two-component regulatory system consisting of a histidine kinase and response regulator. Combining these systems allows the cells to move toward favorable environments and away from their repellents. A representative example of flagellar motility is run-and-tumble swimming in Escherichia coli, where the counter-clockwise (CCW) rotation of a flagellar bundle propels the cell forward, and the clockwise (CW) rotation undergoes cell re-orientation (tumbling) upon switching the direction of flagellar motor rotation from CCW to CW. In this mini review, we focus on several types of chemotactic behaviors that respond to changes in flagellar shape and direction of rotation. Moreover, our single-cell analysis demonstrated back-and-forth swimming motility of an original E. coli strain. We propose that polymorphic flagellar changes are required to enhance bacterial movement in a structured environment as a colony spread on an agar plate.

Keywords: TIRFM; bacterial flagellum; chemotaxis; colony spreading; flagellar polymorphism.

Figure 1

Structure of bacterial flagellum. (A) Electron micrograph of E. coli cell. (B) Current structural and functional models of BFM. OM, outer membrane; PG, peptidoglycan layer; CM, cytoplasmic membrane. (C) Schematics of chemosensory systems. (D) Polymorphic flagellar changes depend on the number of protofilaments; the R-type protofilaments are shown in red. These figures were reproduced with permission from “Seibutsubutsuri, 61 (5), 2021” for Fig. 1A, “Front. Microbiol., 13, 948383, 2022” for Fig. 1B, and “Q. Rev. Biophys., 30 (1), 1–65, 1997” for Fig. 1D, with modifications.

Figure 2

Chemotactic behaviors with relation to flagellation pattern. (A) Flagellation patterns and representative bacterial species. (B) Effect of flagellar shape and rotational direction on cell swimming direction. The rotational direction of the motor was defined as viewed from the filament towards the motor. In left-handed filaments, cells move forward and backward by counterclockwise (CCW) and clockwise (CW) flagellar rotation, respectively. In right-handed filaments, cells move forward and backward by CW and CCW flagellar rotation, respectively. (C) Run-and-tumble behavior of E. coli: between 0 and 0.7 s, a single cell moved forward by CCW rotation of left-handed flagellar filaments; at 0.8 s, the flagella bundle was disrupted by motor switching, causing the cell to change its swimming direction (tumble). During tumbling, the flagellar filament transformed from a left-handed normal filament into a right-handed curly filament. (D) Forward-flick-backward swimming behavior of A. fischeri. (E) Wrapping motion of B. insecticola at 0.625 s intervals. A cell moves forward by the CCW rotation of the left-handed flagellar filament. The flagellar filament transformed into a right-handed coiled filament owing to motor switching from the CCW to CW direction. Consequently, the cells changed their swimming direction without reorientation of the cell body. Images in Fig. 2C-E are fluorescent images; arrows indicate the direction of swimming. Scale bars=2 μm. These figures were reused and modified with permission from “Seibutsubutsuri, 61 (5), 2021” for Fig. 2C and 2D, and “ISME J, 12(3): 838–848, 2018, Springer Nature” for Fig. 2E with modifications.

Figure 3

Distinct chemotactic behaviors in the original E. coli K-12 ATCC10798. (A) Motility of the original E. coli K-12 strain on soft agar plates. Scale bar=1 cm. (B) Sequential phase-contrast images taken at 50-ms intervals for 10 s were integrated using an intermittent color code: “red → yellow → green → cyan → blue.” Scale bar=20 μm. (C) Capillary assay of ATCC10798 cells. In the presence of 1 μM Serine, ATCC10798 cells gathered near the tip, denoted by orange color, indicating that the cells showed chemotactic responses. Scale bar=200 μm. (D) Imaging of fluorescently labeled single filaments using TIRFM. A filament of ATCC10798 was oriented from the 2^nd to the 4^th quadrant relative to the major axis of the filament, indicating right-handed helicity. (E) Sequential TIRFM images of W3110 cells taken at 2.5-ms intervals. The orientation of the flagellar filament(s) is from the 1^st to the 3^rd quadrant relative to the major axis of the filament, indicating left-handed helicity of the filaments. From 47.5 to 65.0 ms, the wave of flagella propagates in a direction away from the hook end toward the flagellar tip, indicating that the flagella rotate in the CCW direction. The direction of rotation was switched at 67.5 ms, and their helicity changed from left- to right-handed within 70 ms. Scale bar=2 μm. (F) Sequential fluorescent images of the original E. coli K-12 ATCC10798 cells, taken at 0.15-s intervals. From 0 to 0.9 s, the cell exhibits forward movement by CCW rotation of the right-handed flagellar filament. At 0.9 s, the motor is switched from the CCW to CW direction, resulting in a change in the swimming direction. This behavior is similar to the back-and-forth movement observed in Vibrio alginolyticus, rather than the run-and-tumble movement observed in the standard motility of E. coli. Scale bar=5 μm. These figures were reused with permission from the “Scientific Reports, 28; 10(1), 15887, 2020, Springer Nature” for Fig. 3A-F, with modifications.

Figure 4

Flagellar polymorphisms required for moving in a structured environment. (A) Sequential-phase contrast images of cellular migration of E. coli W3110 cells on 0.2% agarose. The arrows indicate the swimming direction after a reversal, with an angle change of approximately 180°. Scale bar=5 μm. (B) Diagram showing E. coli escape a dead end within a structured environment. (C) Sequential fluorescence images of Burkholderia insecticola. From 0 to 0.5 s, the cell showed forward movement through CCW rotation of left-handed flagellar filaments before becoming trapped in a dead end. The rotational direction of the motor was then switched from CCW to CW, transforming the filament into a right-handed coiled state. This allowed continuous CW rotation to reorient the swimming direction of the cell. Scale bar=5 μm. (D) Flagella contact the surface, and its point of contact transiently changes. Still image (left), sum-type projection (middle), and kymograph of the middle portion of the left image (right). The green lines indicate the edges of the cell body. Scale bars=2 μm (left and middle) and 3 μm (right). (E) Diagram showing how the wrapping motion occurs in B. insecticola. (D) The kymograph illustrates that the flagellar wave propagation moves in the opposite direction of the flagellar end. This suggests that the flagellar helix instantaneously interacts with the surface and subsequently transiently changes its interaction point by continuous flagellar rotation around its cell body. These figures were reproduced with permission from “Scientific Reports, 28; 10(1), 15887, 2020, Springer Nature” for Fig. 4A and “ISME J, 12(3): 838–848, 2018, Springer Nature” for Fig. 4D with modifications.