Imaging and Analysis of Pseudomonas aeruginosa Swarming and Rhamnolipid Production

Morris, J. D.; Hewitt, Jessica L.; Wolfe, Lawrence G.; Kamatkar, Nachiket G.; Chapman, Sarah M.; Diener, Justin; Courtney, Andrew J.; Leevy, W. Matthew; Shrout, Joshua D.

doi:10.1128/aem.06644-11

Cited by 57 publications

(70 citation statements)

References 55 publications

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“…Images of swarming bacteria expressing GFP or mCherry were obtained using confocal microscopy and entire swarms were imaged using a Carestream multispectral FX or In-Vivo Xtreme imaging station as described previously (3). Specific detail is provided in SI Materials and Methods.…”

Section: Methodsmentioning

confidence: 99%

“…Even the untrained eye would note the coordinated swarming behavior of fish, birds, and insects. Many bacteria also exhibit collective motion by swarming over surfaces in a coordinated manner to move unimpeded at the same time (2)(3)(4). Our knowledge of the specific actions used by individual cells during collective motion is limited; the behavior of single cells within a dense population is difficult to discern experimentally.…”

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confidence: 99%

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Type IV pili interactions promote intercellular association and moderate swarming of Pseudomonas aeruginosa

Anyan¹,

Amiri²,

Harvey³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Pseudomonas aeruginosa is a ubiquitous bacterium that survives in many environments, including as an acute and chronic pathogen in humans. Substantial evidence shows that P. aeruginosa behavior is affected by its motility, and appendages known as flagella and type IV pili (TFP) are known to confer such motility. The role these appendages play when not facilitating motility or attachment, however, is unclear. Here we discern a passive intercellular role of TFP during flagellar-mediated swarming of P. aeruginosa that does not require TFP extension or retraction. We studied swarming at the cellular level using a combination of laboratory experiments and computational simulations to explain the resultant patterns of cells imaged from in vitro swarms. Namely, we used a computational model to simulate swarming and to probe for individual cell behavior that cannot currently be otherwise measured. Our simulations showed that TFP of swarming P. aeruginosa should be distributed all over the cell and that TFP−TFP interactions between cells should be a dominant mechanism that promotes cell−cell interaction, limits lone cell movement, and slows swarm expansion. This predicted physical mechanism involving TFP was confirmed in vitro using pairwise mixtures of strains with and without TFP where cells without TFP separate from cells with TFP. While TFP slow swarm expansion, we show in vitro that TFP help alter collective motion to avoid toxic compounds such as the antibiotic carbenicillin. Thus, TFP physically affect P. aeruginosa swarming by actively promoting cell−cell association and directional collective motion within motile groups to aid their survival.he bacterium Pseudomonas aeruginosa is a ubiquitous organism that is a known opportunistic pathogen, causing both chronic and acute infections in susceptible populations, including individuals with cystic fibrosis or burn wounds, or Intensive Care Unit patients (1). Among questions that remain unanswered for nonobligate pathogens like P. aeruginosa is how these bacteria initiate infections after entering the host from the environment. Given that P. aeruginosa is among many bacteria that grow as a biofilm during infection, there is a need to understand how individual cells coordinate in space with each other to colonize new surfaces and subsequently transition to stationary biofilms.Many organisms coordinate their movement as a population, emerging as self-organized swarming groups. Even the untrained eye would note the coordinated swarming behavior of fish, birds, and insects. Many bacteria also exhibit collective motion by swarming over surfaces in a coordinated manner to move unimpeded at the same time (2-4). Our knowledge of the specific actions used by individual cells during collective motion is limited; the behavior of single cells within a dense population is difficult to discern experimentally. Previous attempts to study bacterial collective behavior have used computational models to test mechanisms hypothesized to influence collective motion, including directional r...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Type IV pili interactions promote intercellular association and moderate swarming of Pseudomonas aeruginosa

Anyan¹,

Amiri²,

Harvey³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…Optical images of cells within swarms were acquired using fluorescence microscopy or confocal laser scanning microscopy as previously described (6,36) and as detailed in SI Experimental Procedures. CRM imaging was performed as previously described (16) and as detailed in SI Experimental Procedures.…”

Section: Methodsmentioning

confidence: 99%

“…P. aeruginosa is an opportunistic pathogen and one of many bacteria that displays numerous community behaviors, including an ability to readily form surface-attached biofilms. Prior to establishing stationary biofilm communities, P. aeruginosa is known in vitro to exhibit swarming (1,6,7), a group motility behavior employed by some bacteria to explore and expand during surface colonization. While many studies have addressed biofilm development and the transition to static bacterial biofilms, the community behaviors exhibited by motile bacteria are less understood.…”

Section: Introductionmentioning

confidence: 99%

Spatially dependent alkyl quinolone signaling responses to antibiotics in Pseudomonas aeruginosa swarms

Morales-Soto¹,

Dunham

Baig

et al. 2018

Journal of Biological Chemistry

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There is a general lack of understanding about how communities of bacteria respond to exogenous toxins such as antibiotics. Most of our understanding of community-level stress responses comes from the study of stationary biofilm communities. Although several community behaviors and production of specific biomolecules affecting biofilm development and associated behavior have been described for Pseudomonas aeruginosa and other bacteria, we have little appreciation for the production and dispersal of secreted metabolites within the 2D and 3D spaces they occupy as they colonize, spread, and grow on surfaces. Here we specifically studied the phenotypic responses and spatial variability of alkyl quinolones, including the Pseudomonas quinolone signal (PQS) and members of the alkyl hydroxyquinoline (AQNO) subclass, in P. aeruginosa plateassay swarming communities. We found that PQS production was not a universal signaling response to antibiotics as tobramycin elicited an alkyl quinolone response while carbenicillin did not. We also found that PQS and AQNO profiles in response to tobramycin were markedly distinct and influenced these swarms on different spatial scales. The distribution of alkyl quinolones varied by several orders of magnitude within the same swarm. At some tobramycin exposures, P. aeruginosa swarms produced alkyl quinolones in the range of 150 µM PQS and 400 µM AQNO that accumulated as aggregates.

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“…The surface growth rate of each strain was obtained by quantifying fluorescence over time by a fluorescence-imaging method (43). Briefly, M. xanthus colonies were grown on one 150-mm CTT agar (1.5%) plate containing 8 l/100 ml of Syto64 bacterial-staining dye (Life Technologies, Grand Island, NY).…”

Section: Methodsmentioning

confidence: 99%

Cell Division Resets Polarity and Motility for the Bacterium Myxococcus xanthus

et al. 2014

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e Links between cell division and other cellular processes are poorly understood. It is difficult to simultaneously examine division and function in most cell types. Most of the research probing aspects of cell division has experimented with stationary or immobilized cells or distinctly asymmetrical cells. Here we took an alternative approach by examining cell division events within motile groups of cells growing on solid medium by time-lapse microscopy. A total of 558 cell divisions were identified among approximately 12,000 cells. We found an interconnection of division, motility, and polarity in the bacterium Myxococcus xanthus. For every division event, motile cells stop moving to divide. Progeny cells of binary fission subsequently move in opposing directions. This behavior involves M. xanthus Frz proteins that regulate M. xanthus motility reversals but is independent of type IV pilus "S motility." The inheritance of opposing polarity is correlated with the distribution of the G protein RomR within these dividing cells. The constriction at the point of division limits the intracellular distribution of RomR. Thus, the asymmetric distribution of RomR at the parent cell poles becomes mirrored at new poles initiated at the site of division. Many approaches to study cell division utilize traits that readily allow the distinction of two progeny cells. For example, cells displaying "asymmetrical" division traits allow the clear distinction of numerous characteristics that can then be monitored while deciphering other unknowns. Caulobacter bacteria are among the best studied with this distinction (1), but other biological examples include: preneuron neuroblast brain cells, budding Saccharomyces cerevisiae, germ line cells of male Drosophila, endospore development by Bacillus species, and Mycobacterium species subjected to environmental nutrient stress (2, 3). While such explicitly distinct examples may be rare, nearly all types of cells display some asymmetrical properties when functioning properly. There are numerous examples of distinctive asymmetrical and polarized attributes of cells (4). However, one difficulty that remains in characterizing asymmetrical properties in biology is distinguishing the timing and order of those intra-and intercellular attributes that are transient in nature. Alternative to studying asymmetric cell types that can be readily differentiated, other research strategies to probe stages of division often examine stationary or immobilized cells.Myxococcus xanthus is one of many myxobacteria, common soil microbes that grow readily in environments rich in complex organics, such as those containing decaying plants (5) or other bacteria (6). M. xanthus cells exhibit a symmetric morphology. The specific mechanism and dynamics of M. xanthus cell division, like those of most nonmodel organisms, are not entirely known. M. xanthus is among many bacteria lacking a clear MinCD system that drives the recruitment of FtsZ for division. It is known that the middle of M. xanthus cells is marked by PomZ, w...

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Imaging and Analysis of Pseudomonas aeruginosa Swarming and Rhamnolipid Production

Cited by 57 publications

References 55 publications

Type IV pili interactions promote intercellular association and moderate swarming of Pseudomonas aeruginosa

Type IV pili interactions promote intercellular association and moderate swarming of Pseudomonas aeruginosa

Spatially dependent alkyl quinolone signaling responses to antibiotics in Pseudomonas aeruginosa swarms

Cell Division Resets Polarity and Motility for the Bacterium Myxococcus xanthus

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