title: Extracellular enzymes affect biofilm formation of mucoid Pseudomonas aeruginosa.
abstract: Pseudomonas aeruginosa secretes a variety of hydrolases, many of which contribute to virulence or are thought to play a role in the nutrition of the bacterium. As most studies concerning extracellular enzymes have been performed on planktonic cultures of non-mucoid P. aeruginosa strains, knowledge of the potential role of these enzymes in biofilm formation in mucoid (alginate-producing) P. aeruginosa remains limited. Here we show that mucoid P. aeruginosa produces extracellular hydrolases during biofilm growth. Overexpression of the extracellular lipases LipA and LipC, the esterase EstA and the proteolytic elastase LasB from plasmids revealed that some of these hydrolases affected the composition and physicochemical properties of the extracellular polymeric substances (EPS). While no influence of LipA was observed, the overexpression of estA and lasB led to increased concentrations of extracellular rhamnolipids with enhanced levels of mono-rhamnolipids, elevated amounts of total carbohydrates and decreased alginate concentrations, resulting in increased EPS hydrophobicity and viscosity. Moreover, we observed an influence of the enzymes on cellular motility. Overexpression of estA resulted in a loss of twitching motility, although it enhanced the ability to swim and swarm. The lasB-overexpression strain showed an overall enhanced motility compared with the parent strain. Moreover, the EstA- and LasB-overproduction strains completely lost the ability to form 3D biofilms, whereas the overproduction of LipC increased cell aggregation and the heterogeneity of the biofilms formed. Overall, these findings indicate that directly or indirectly, the secreted enzymes EstA, LasB and LipC can influence the formation and architecture of mucoid P. aeruginosa biofilms as a result of changes in EPS composition and properties, as well as the motility of the cells.
authors.affiliation: University of Duisburg-Essen, Faculty of Chemistry, Biofilm Centre, Department of Aquatic Microbiology, Geibelstrasse 41, D-47057 Duisburg, Germany.
authors.forename: Petra
authors.initials: P
authors.lastname: Tielen
authors.affiliation: Heinrich-Heine-University of Duesseldorf, Institute for Molecular Enzyme Technology, Research Centre Juelich, Stetternicher Forst, D-52425 Juelich, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Rosenau
authors.affiliation: Heinrich-Heine-University of Duesseldorf, Institute for Molecular Enzyme Technology, Research Centre Juelich, Stetternicher Forst, D-52425 Juelich, Germany.
authors.forename: Susanne
authors.initials: S
authors.lastname: Wilhelm
authors.affiliation: Heinrich-Heine-University of Duesseldorf, Institute for Molecular Enzyme Technology, Research Centre Juelich, Stetternicher Forst, D-52425 Juelich, Germany.
authors.forename: Karl-Erich
authors.initials: KE
authors.lastname: Jaeger
authors.affiliation: University of Duisburg-Essen, Faculty of Chemistry, Biofilm Centre, Department of Aquatic Microbiology, Geibelstrasse 41, D-47057 Duisburg, Germany.
authors.forename: Hans-Curt
authors.initials: HC
authors.lastname: Flemming
authors.affiliation: University of Duisburg-Essen, Faculty of Chemistry, Biofilm Centre, Department of Aquatic Microbiology, Geibelstrasse 41, D-47057 Duisburg, Germany.
authors.forename: Jost
authors.initials: J
authors.lastname: Wingender
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D018441:Biofilms; D005110:Extracellular Space / Q000201:enzymology* / Q000235:genetics; D006025:Glycosaminoglycans / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D006025:Glycosaminoglycans; C046129:mucoid exopolysaccharide (P aeruginosa)