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, 182 (21), 5990-6

Swarming of Pseudomonas Aeruginosa Is Dependent on Cell-To-Cell Signaling and Requires Flagella and Pili

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Swarming of Pseudomonas Aeruginosa Is Dependent on Cell-To-Cell Signaling and Requires Flagella and Pili

T Köhler et al. J Bacteriol.

Abstract

We describe swarming in Pseudomonas aeruginosa as a third mode of surface translocation in addition to the previously described swimming and twitching motilities. Swarming in P. aeruginosa is induced on semisolid surfaces (0.5 to 0.7% agar) under conditions of nitrogen limitation and in response to certain amino acids. Glutamate, aspartate, histidine, or proline, when provided as the sole source of nitrogen, induced swarming, while arginine, asparagine, and glutamine, among other amino acids, did not sustain swarming. Cells from the edge of the swarm were about twice as long as cells from the swarm center. In both instances, bacteria possessing two polar flagella were observed by light and electron microscopy. While a fliC mutant of P. aeruginosa displayed slightly diminished swarming, a pilR and a pilA mutant, both deficient in type IV pili, were unable to swarm. Furthermore, cells with mutations in the las cell-to-cell signaling system showed diminished swarming behavior, while rhl mutants were completely unable to swarm. Evidence is presented for rhamnolipids being the actual surfactant involved in swarming motility, which explains the involvement of the cell-to-cell signaling circuitry of P. aeruginosa in this type of surface motility.

Figures

FIG. 1
FIG. 1
Swarming in P. aeruginosa is induced by agar concentrations below 0.7% (A) and by specific amino acids (B). Colonies of wild-type strain PT5 from a fresh LB agar plate were inoculated by toothpick into the middle of the swarm plates. In panel A the medium contained 0.2% glucose as the carbon source and 0.05% (wt/vol) glutamate as the nitrogen source. Plates in panel B contained 0.02% glucose, a 0.05% concentration of the indicated amino acid as the nitrogen source, and 0.6% agar. Plates were incubated for 24 h at 37°C.
FIG. 2
FIG. 2
Swarming is dependent on the carbon source. M8 swarm plates with aspartate as the nitrogen source were supplemented with either glucose, glycerol, or succinate at a final concentration of 100 mM each. PT5 was inoculated in the center, followed by 24 h of incubation at 37°C.
FIG. 3
FIG. 3
Electron microscopy of PT5 cells taken from the swarm edge and the swarm center. (A) Elongated cells of approximately 3 to 4 μm were observed at the periphery of the swarming colony. (B) Smaller cells, of about 2 μm, were observed at the swarm center. A cell expressing two polar flagella is indicated by the arrow. A few elongated cells from the swarm edge were also found to possess two flagella. Magnification is ×8,500.
FIG. 4
FIG. 4
Swimming and swarming motilities in P. aeruginosa wild-type (WT) PT5 and its fliC and pilA mutant derivatives. Swimming plates were made of LB agar with 0.3% agar. After inoculation, plates were incubated at room temperature for 24 h. Swarm plates were M8-glucose-glutamate plates containing 0.5% agar. Incubation was done at 37°C for 24 h.
FIG. 5
FIG. 5
Swarming requires the las and rhl cell-to-cell signaling systems. The lasI, lasR, rhlI, and rhlR mutants were inoculated on a swarm plate which was incubated at 37°C for 24 and 48 h at room temperature. As a comparison, the wild-type strain, PT5, would have covered the whole plate by that time.
FIG. 6
FIG. 6
Rhamnolipids are the biosurfactant required for swarming in P. aeruginosa. The wild-type (wt) strain, PT5, and rhlA mutant PT712 were inoculated on a swarm plate and incubated at 37°C for 24 h.

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