Characterization of a novel air-liquid interface biofilm of Pseudomonas fluorescens SBW25

Pseudomonads are able to form a variety of biofilms that colonize the air-liquid (A-L) interface of static liquid microcosms, and differ in matrix composition, strength, resilience and degrees of attachment to the microcosm walls. From Pseudomonas fluorescens SBW25, mutants have evolved during prolonged adaptation-evolution experiments which produce robust biofilms of the physically cohesive class at the A-L interface, and which have been well characterized. In this study we describe a novel A-L interface biofilm produced by SBW25 that is categorized as a viscous mass (VM)-class biofilm. Several metals were found to induce this biofilm in static King's B microcosms, including copper, iron, lead and manganese, and we have used iron to allow further examination of this structure. Iron was demonstrated to induce SBW25 to express cellulose, which provided the matrix of the biofilm, a weak structure that was readily destroyed by physical disturbance. This was confirmed in situ by a low (0.023-0.047 g) maximum deformation mass and relatively poor attachment as measured by crystal violet staining. Biofilm strength increased with increasing iron concentration, in contrast to attachment levels, which decreased with increasing iron. Furthermore, iron added to mature biofilms significantly increased strength, suggesting that iron also promotes interactions between cellulose fibres that increase matrix interconnectivity. Whilst weak attachment is important in maintaining the biofilm at the A-L interface, surface-interaction effects involving cellulose, which reduced surface tension by approximately 3.8 mN m(-1), may also contribute towards this localization. The fragility and viscoelastic nature of the biofilm were confirmed by controlled-stress amplitude sweep tests to characterize critical rheological parameters, which included a shear modulus of 0.75 Pa, a zero shear viscosity of 0.24 Pa s(-1) and a flow point of 0.028 Pa. Growth and morphological data thus far support a non-specific metal-associated physiological, rather than mutational, origin for production of the SBW25 VM biofilm, which is an example of the versatility of bacteria to inhabit optimal niches within their environment.