Both images at 1000× magnification. Scale bar = 10 microns. Chemostat biofilm culture V. paradoxus EPS was inoculated into a Biosurface Technologies CDC biofilm reactor and grown as a batch culture for 20 h (Fig 10A, B). Continuous culture for 2d after this initial batch phase resulted in the formation of a dense,

filamentous biofilm (Fig 10C–H). Staining with the BacLight system (Invitrogen) showed a mixed population of live and dead cells at all stages of development. At higher magnification, the filamentous structures of the developing biofilm are readily apparent, and filaments that stain with propidium iodide, indicating dead cells, are particularly strongly evident. Figure 10 Biofilms cultivated in a CDC stirred Selinexor price biofilm reactor. V. paradoxus EPS was cultured from a broth inoculum for 18 h under stirred batch conditions (A, B), followed by 24 h (C, D) or 48 h (E, F) under continuous flow conditions (2 ml/min). BacLight staining with PI (red, dead cells) beta-catenin phosphorylation and Syto9 (green, live cells). 100×, scale bar = 100 microns (A, C, E). 400×, scale bar = 25 microns (B, D, F). Discussion The environmental bacterium Variovorax

paradoxus is involved in a number of important processes, such as promoting plant growth and remediation of xenobiotics. Our work with the V. paradoxus strain EPS demonstrates that this strain is capable of coordinated surface behaviors in laboratory culture. The behaviors we’ve examined in this report are the development of a swarm on defined high water activity (low agarose content) media and the formation of biofilms on several abiotic surfaces. We have examined the capaCity of this organism to move across a solid surface, and identified the motility demonstrated as swarming. We utilized agarose as the solidifying agent in our media, at 0.5% w/v, based on previous swarming analyses [39] and auxotrophy studies in our lab showing that V. paradoxus EPS utilizes organic components of bacteriological agar as nutrients (not shown). The motility was shown to require flagellar activity (Fig 2, 3),

aminophylline and to involve the production of a chemically uncharacterized wetting agent (Fig 4). The presence of 1–3 flagella per cell on swimming V. paradoxus has been noted in previous work, and is cited as a defining characteristic of this taxon [41]. We identified these flagella in broth cultures of our strain (not shown). In the recently released draft sequence of V. paradoxus S110, genes encoding flagellar components have been identified (Han et al, http://​genome.​ornl.​gov/​microbial/​vpar_​s110). Based on these data along with our experimental results, we feel justified in labeling the surface motility observed as swarming motility. Our experiments allow some insights into the mechanism of V. paradoxus EPS swarming. Swarming is inhibited by Congo Red with a threshold value of 50 μg/L, consistent with the inhibition of the function of a single flagellum.