Visualization of Flagella during Bacterial Swarming

Abstract

When cells of Escherichia coli are grown in broth and suspended at low density in a motility medium, they swim independently, exploring a homogeneous, isotropic environment. Cell trajectories and the way in which these trajectories are determined by flagellar dynamics are well understood. When cells are grown in a rich medium on agar instead, they elongate, produce more flagella, and swarm. They move in coordinated packs within a thin film of fluid, in intimate contact with one another and with two fixed surfaces, a surfactant monolayer above and an agar matrix below: they move in an inhomogeneous, anisotropic environment. Here we examine swarm-cell trajectories and ways in which these trajectories are determined by flagellar motion, visualizing the cell bodies by phase-contrast microscopy and the flagellar filaments by fluorescence microscopy. We distinguish four kinds of tracks, defining stalls, reversals, lateral movement, and forward movement. When cells are stalled at the edge of a colony, they extend their flagellar filaments outwards, moving fluid over the virgin agar; when cells reverse, changes in filament chirality play a crucial role; when cells move laterally, they are pushed sideways by adjacent cells; and when cells move forward, they are pushed by flagellar bundles in the same way as when they are swimming in bulk aqueous media. These maneuvers are described in this report.

Single-cell analysis in situ in a Bacillus subtilis swarming community identifies distinct spatially separated subpopulations differentially expressing hag (flagellin), including specialized swarmers

“…Thus, as illustrated in Supplementary Movie Files S1 and S2 (14 frames s 21 ), the cells formed many short-lived rafts or packs composed of around three to eight aligned cells. As shown in Supplementary Movie File S3, when cells at dendrite tips were filmed at 40 frames s 21 , we saw highly mobile swirling packs and large streams of cells, behaviour resembling the 'acrobatics' shown by E. coli (Copeland et al, 2010;Darnton et al, 2010;Turner et al, 2010;McCarter, 2010). Such behaviour has been shown to be characteristic of bacteria suspended in a thin fluid film .…”

“…Serratia liquefaciens swarmers, however, appear to be only moderately elongated, although profusely flagellated (Eberl et al, 1999). In contrast, in Escherichia coli and Salmonella typhimurium Turner et al, 2010;Harshey, 2003), migration is restricted to softer agar (around 0.5 %), and involves smaller clusters (packs) of swarmers that are only moderately elongated and flagellated (two-to threefold in each case). In Pseudomonas aeruginosa, swarming takes the form of unbranched dendrites or tendrils, rather than confluent fields, and swarmers are around twofold larger than vegetative cells, with two flagella rather than one (Rashid & Kornberg, 2000;Caiazza et al, 2005).…”

Water reservoir maintained by cell growth fuels the spreading of a bacterial swarm

“…The agar was autoclaved and stored at room temperature. Before use, it was melted in a microwave oven, cooled to approximately 60°C, and pipetted in 25-mL aliquots into 150 × 15-mm polystyrene petri plates (8). Antibiotics (for E. coli HCB1668 and MG1655-ASV) and arabinose (for E. coli HCB1668) were added to the liquefied swarm agar before pipetting at the concentrations used in liquid cultures.…”

“…The strain used for studies of fluid motion was E. coli HCB1668 (FliC S353C), an AW405 derivative that swarms well, developed for flagellar visualization (8). The strain used for studies of cell growth and cell density was E. coli MG1655-ASV (a gift from Kim Lewis, Northeastern University, Boston, MA), which expresses a short-lived derivative of GFP (ASV) under control of the ribosomal rrnBP1 promoter, developed for studies of cell growth (21,22).…”

“…Much has been learned about the genetics and biochemistry of bacterial swarming as well as its relevance to biofilm formation and pathogenic infections (1)(2)(3). More recent advances have been made at the single-cell level (6)(7)(8)(9). However, relatively little is known about the thin layer of fluid that supports flagellar motility and allows swarm cells to maintain a distinct physiological state (10,11).…”