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Pel exopolysaccharide biosynthetic loci are phylogenetically widespread biofilm matrix determinants in bacteria. In Pseudomonas aeruginosa, Pel is crucial for cell-to-cell interactions and reducing susceptibility to antibiotic and mucolytic treatments. While genes encoding glycoside hydrolases have long been linked to biofilm exopolysaccharide biosynthesis, their physiological role in biofilm development is unclear. Here we demonstrate that the glycoside hydrolase activity of P. aeruginosa PelA decreases adherent biofilm biomass and is responsible for generating the low molecular weight secreted form of the Pel exopolysaccharide. We show that the generation of secreted Pel contributes to the biomechanical properties of the biofilm and decreases the virulence of P. aeruginosa in Caenorhabditis elegans and Drosophila melanogaster. Our results reveal that glycoside hydrolases found in exopolysaccharide biosynthetic systems can help shape the soft matter attributes of a biofilm and propose that secreted matrix components be referred to as matrix associated to better reflect their influence., (© 2023. The Author(s).)

Bacterial colonization and biofilm formation on surfaces are typically mediated by the deposition of exopolysaccharides and conditioning protein layers. Pseudomonas aeruginosa is a nosocomial opportunistic pathogen that utilizes strain-specific exopolysaccharides such as Psl, Pel or alginate for both initial surface attachment and biofilm formation. To generate surfaces that resist P. aeruginosa colonization, we covalently bound a Psl-specific glycoside hydrolase (PslG h ) to several, chemically-distinct surfaces using amine functionalization (APTMS) and glutaraldehyde (GDA) linking. In situ quartz crystal microbalance (QCM) experiments and fluorescence microscopy demonstrated a complete lack of Psl adsorption on the PslG h -bound surfaces. Covalently-bound PslG h was also found to significantly reduce P. aeruginosa surface attachment and biofilm formation over extended growth periods (8 days). The PslG h surfaces showed a ∼99.9% (∼3-log) reduction in surface associated bacteria compared to control (untreated) surfaces, or those treated with inactive enzyme. This work demonstrates a non-eluting 'bioactive' surface that specifically targets a mechanism of cell adhesion, and that surface-bound glycoside hydrolase can significantly reduce surface colonization of bacteria through local, continuous enzymatic degradation of exopolysaccharide (Psl). These results have significant implications for the surface design of medical devices to keep bacteria in a planktonic state, and therefore susceptible to antibiotics and antimicrobials., (Copyright © 2018 Elsevier Ltd. All rights reserved.)