Confinement discerns swarmers from planktonic bacteria

Powered by flagella, many bacterial species exhibit collective motion on a solid surface commonly known as swarming. As a natural example of active matter, swarming is also an essential biological phenotype associated with virulence, chemotaxis, and host pathogenesis. Physical changes like cell elongation and hyper flagellation have been shown to accompany the swarming phenotype. Less studied, however, are the contrasts of collective motion between the swarming cells and their counterpart planktonic cells of comparable cell density. Here, we show that confining bacterial movement in circular microwells allows distinguishing bacterial swarming from collective swimming. On a soft agar plate, a novel bacterial strain Enterobacter sp. SM3 in swarming and planktonic states exhibited different motion patterns when confined to circular microwells of a specific range of sizes. When the confinement diameter was between 40 μm and 90 μm, swarming SM3 formed a single swirl motion pattern in the microwells whereas planktonic SM3 formed multiple swirls. Similar differential behavior is observed across several other species of gram-negative bacteria. We also observed 'rafting behavior' of swarming bacteria upon dilution. We hypothesize that the rafting behavior might account for the motion pattern difference. We were able to predict these experimental features via numerical simulations where swarming cells are modeled with stronger cell-cell alignment interaction. Our experimental design using PDMS microchip disk arrays enabled us to observe bacterial swarming on murine intestinal surface suggesting a new method for characterizing bacterial swarming under complex environments, such as in polymicrobial niches, and for in vivo swarming exploration.

Rhamnolipids and their 3-(3-hydroxyalkanoyloxy)alkanoic acid precursors activate Arabidopsis innate immunity through two independent mechanisms

Plant innate immunity is activated upon perception of invasion pattern molecules by plant cell-surface immune receptors. Several bacteria of the genera Pseudomonas and Burkholderia produce rhamnolipids (RLs) from L-rhamnose and (R)-3-hydroxyalkanoate precursors (HAAs). RL and HAA secretion is required to modulate bacterial swarming motility behavior. The bulb-type lectin receptor kinase LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION/S-DOMAIN-1-29 (LORE/SD1-29) mediates medium-chain 3-hydroxy fatty acid (mc-3-OH-FA) sensing in the plant Arabidopsis thaliana. Here, we show that the lipidic secretome from Pseudomonas aeruginosa comprising RLs, HAAs and mc-3-OH-FAs stimulates Arabidopsis immunity. HAAs, like mc-3-O-FAs, are sensed by LORE and induce canonical immune signaling and local resistance to plant pathogenic Pseudomonas infection. By contrast, RLs trigger an atypical immune response and resistance to Pseudomonas infection independent of LORE. Thus, the glycosyl moieties of RLs, albeit abolishing sensing by LORE, do not impair their ability to trigger plant defense. In addition, our results show that RL-triggered immune response is affected by the sphingolipid composition of the plasma membrane. In conclusion, RLs and their precursors released by bacteria can both be perceived by plants but through distinct mechanisms.

Confinement Discerns Swarmers from Planktonic Bacteria

Powered by flagella, many bacterial species exhibit collective motion on a solid surface commonly known as swarming. As a natural example of active matter, swarming is also an essential biological phenotype associated with virulence, chemotaxis, and host pathogenesis. Physical changes like cell elongation and hyper flagellation have been shown to accompany the swarming phenotype. However, less noticeable, are the contrasts of collective motion between the swarming cells and the planktonic cells of comparable cell density. Here, we show that confining bacterial movement in designed dimensions allows distinguishing bacterial swarming from collective swimming. We found that on a soft agar plate, a novel bacterial strain Enterobacter sp. SM3 exhibited different motion patterns in swarming and planktonic states when confined to circular microwells of a specific range of sizes. When the confinement diameter was between 40 μm and 90 μm, swarming SM3 formed a single swirl motion pattern in the microwells whereas planktonic SM3 showed multiple swirls. Similar differential behavior is observed across a range of randomly selected gram-negative bacteria. We hypothesize that the “rafting behavior” of the swarming bacteria upon dilution might account for the motion pattern difference. We verified our conjectures via numerical simulations where swarming cells are modeled with lower repulsion and more substantial alignment force. The novel technical approach enabled us to observe swarming on a non-agar tissue surface for the first time. Our work provides the basis for characterizing bacterial swarming under more sophisticated environments, such as polymicrobial swarmer detection, and in vivo swarming exploration.

Remodeling of Chemotaxis is a Cornerstone of Bacterial Swarming

Many bacteria use flagella-driven motility to swarm or move collectively over a surface terrain. Bacterial adaptations for swarming can include cell elongation, hyper-flagellation, recruitment of special stator proteins and surfactant secretion, among others. We recently demonstrated another swarming adaptation in Escherichia coli, wherein the chemotaxis pathway is remodeled to increase run durations (decrease tumble bias), with running speeds increased as well. We show here that the modification of motility parameters during swarming is not unique to E. coli, but shared by a diverse group of bacteria we examined – Proteus mirabilis, Serratia marcescens, Salmonella enterica, Bacillus subtilis, and Pseudomonas aeruginosa – suggesting that altering the chemosensory physiology is a cornerstone of swarming.ImportanceBacteria within a swarm move characteristically in packs, displaying an intricate swirling motion where hundreds of dynamic packs continuously form and dissociate as the swarm colonizes increasing expanse of territory. The demonstrated property of E. coli to reduce its tumble bias and hence increase its run duration during swarming is expected to maintain/promote side-by-side alignment and cohesion within the bacterial packs. Here we observe a similar low tumble bias in five different bacterial species, both Gram positive and Gram negative, each inhabiting a unique habitat and posing unique problems to our health. The unanimous display of an altered run-tumble bias in swarms of all species examined here suggests that this behavioral adaptation is crucial for swarming.

Optimizing a human fecal assay that elicits bacterial swarming

A distinct property of many bacteria is swarming: swift movement across a surface through flagella propulsion. Early research indicates that bacterial swarming can be a protective host response to intestinal inflammation. Central to the further study of bacterial swarming in human health is an effective and replicable assay for swarming that can accommodate complex material, such as fecal matter. To date, nearly all swarming assays described in the literature are specific for bacteria grown in culture, most often Pseudomonas. In this paper, we describe a protocol for discerning swarming of bacteria from frozen human fecal samples. Moreover, we tested 4 variables that may influence the effectiveness of the assay: the method by which frozen samples were thawed, the concentration of agar used in the Lysogenic broth (LB) agar plate, the volume of LB agar poured in the plate, and the volume of sample inoculated. We found that while the type of thaw and volume of LB agar had little to no effect on swarming, greater concentrations of agar were negatively correlated with swarming and greater volumes of the sample were positively correlated with swarming.