Wetting of dehydrated Pseudomonas f luorescens biofilms grown on glass substrates by an external liquid is employed as a means to investigate the complex morphology of these biofilms along with their capability to interact with external fluids. The porous structure left behind after dehydration induces interesting droplet spreading on the external surface and imbibition into pores upon wetting. Static contact angles and volume loss by imbibition measured right upon droplet deposition indicate that biofilms of higher incubation times show a higher porosity and effective hydrophilicity. Furthermore, during subsequent rotation tests, using Kerberos device, these properties dictate a peculiar forced wetting/spreading behavior. As rotation speed increases a long liquid tail forms progressively at the rear part of the droplet, which stays pinned at all times, while only the front part of the droplet depins and spreads. Interestingly, the experimentally determined retention force for the onset of droplet sliding on biofilm external surface is lower than that on pure glass. An effort is made to describe such complex forced wetting phenomena by presenting apparent contact angles, droplet length, droplet shape contours, and edges position as obtained from detailed image analysis.
Biofilms consist of a complex microbial community adhering to biotic or abiotic surfaces and enclosed within a protein/polysaccharide self-produced matrix. The formation of this structure represents the most important adaptive mechanism that leads to antibacterial resistance, and therefore, closely connected to pathogenicity. Antimicrobial peptides (AMPs) could represent attractive candidates for the design of new antibiotics because of their specific characteristics. AMPs show a broad activity spectrum, a relative selectivity towards their targets (microbial membranes), the ability to act on both proliferative and quiescent cells, a rapid mechanism of action, and above all, a low propensity for developing resistance. This article investigates the effect at subMIC concentrations of Temporin-L (TL) on biofilm formation in Pseudomonas fluorescens (P. fluorescens) both in static and dynamic conditions, showing that TL displays antibiofilm properties. Biofilm formation in static conditions was analyzed by the Crystal Violet assay. Investigation of biofilms in dynamic conditions was performed in a commercial microfluidic device consisting of a microflow chamber to simulate real flow conditions in the human body. Biofilm morphology was examined using Confocal Laser Scanning Microscopy and quantified via image analysis. The investigation of TL effects on P. fluorescens showed that when subMIC concentrations of this peptide were added during bacterial growth, TL exerted antibiofilm activity, impairing biofilm formation both in static and dynamic conditions. Moreover, TL also affects mature biofilm as confocal microscopy analyses showed that a large portion of preformed biofilm architecture was clearly perturbed by the peptide addition with a significative decrease of all the biofilm surface properties and the overall biomass. Finally, in these conditions, TL did not affect bacterial cells as the live/dead cell ratio remained unchanged without any increase in damaged cells, confirming an actual antibiofilm activity of the peptide.
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