title: Exopolysaccharide biosynthetic glycoside hydrolases can be utilized to disrupt and prevent Pseudomonas aeruginosa biofilms.
abstract: Bacterial biofilms present a significant medical challenge because they are recalcitrant to current therapeutic regimes. A key component of biofilm formation in the opportunistic human pathogen Pseudomonas aeruginosa is the biosynthesis of the exopolysaccharides Pel and Psl, which are involved in the formation and maintenance of the structural biofilm scaffold and protection against antimicrobials and host defenses. Given that the glycoside hydrolases PelAh and PslGh encoded in the pel and psl biosynthetic operons, respectively, are utilized for in vivo exopolysaccharide processing, we reasoned that these would provide specificity to target P. aeruginosa biofilms. Evaluating these enzymes as potential therapeutics, we demonstrate that these glycoside hydrolases selectively target and degrade the exopolysaccharide component of the biofilm matrix. PelAh and PslGh inhibit biofilm formation over a 24-hour period with a half maximal effective concentration (EC50) of 69.3 ± 1.2 and 4.1 ± 1.1 nM, respectively, and are capable of disrupting preexisting biofilms in 1 hour with EC50 of 35.7 ± 1.1 and 12.9 ± 1.1 nM, respectively. This treatment was effective against clinical and environmental P. aeruginosa isolates and reduced biofilm biomass by 58 to 94%. These noncytotoxic enzymes potentiated antibiotics because the addition of either enzyme to a sublethal concentration of colistin reduced viable bacterial counts by 2.5 orders of magnitude when used either prophylactically or on established 24-hour biofilms. In addition, PelAh was able to increase neutrophil killing by ~50%. This work illustrates the feasibility and benefits of using bacterial exopolysaccharide biosynthetic glycoside hydrolases to develop novel antibiofilm therapeutics.
authors.affiliation: Program in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
authors.forename: Perrin
authors.initials: P
authors.lastname: Baker
authors.affiliation: Departments of Microbial Infection and Immunity, Microbiology, Center for Microbial Interface Biology, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Preston J
authors.initials: PJ
authors.lastname: Hill
authors.affiliation: Departments of Medicine, Microbiology, and Immunology, McGill University, Montréal, Québec H3A 2B4, Canada.; Infectious Diseases and Immunity in Global Health Program, Centre for Translational Biology, McGill University Health Centre, Montréal, Québec H4A 3J1, Canada.
authors.forename: Brendan D
authors.initials: BD
authors.lastname: Snarr
authors.affiliation: Program in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.; Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
authors.forename: Noor
authors.identifier: 0000-0001-8930-6185
authors.initials: N
authors.lastname: Alnabelseya
authors.affiliation: Departments of Microbial Infection and Immunity, Microbiology, Center for Microbial Interface Biology, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Pestrak
authors.affiliation: Departments of Medicine, Microbiology, and Immunology, McGill University, Montréal, Québec H3A 2B4, Canada.; Infectious Diseases and Immunity in Global Health Program, Centre for Translational Biology, McGill University Health Centre, Montréal, Québec H4A 3J1, Canada.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Lee
authors.affiliation: Department of Microbiology, University of Washington, Seattle, WA 98195, USA.
authors.forename: Laura K
authors.initials: LK
authors.lastname: Jennings
authors.affiliation: Program in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
authors.forename: John
authors.identifier: 0000-0002-2475-6465
authors.initials: J
authors.lastname: Tam
authors.affiliation: Program in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.; Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
authors.forename: Roman A
authors.initials: RA
authors.lastname: Melnyk
authors.affiliation: Department of Microbiology, University of Washington, Seattle, WA 98195, USA.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Parsek
authors.affiliation: Departments of Medicine, Microbiology, and Immunology, McGill University, Montréal, Québec H3A 2B4, Canada.; Infectious Diseases and Immunity in Global Health Program, Centre for Translational Biology, McGill University Health Centre, Montréal, Québec H4A 3J1, Canada.
authors.forename: Donald C
authors.initials: DC
authors.lastname: Sheppard
authors.affiliation: Departments of Microbial Infection and Immunity, Microbiology, Center for Microbial Interface Biology, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Daniel J
authors.identifier: 0000-0003-4592-4816
authors.initials: DJ
authors.lastname: Wozniak
authors.affiliation: Program in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.; Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
authors.forename: P Lynne
authors.initials: PL
authors.lastname: Howell
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D018441:Biofilms / Q000187:drug effects / Q000254:growth & development*; D002384:Catalysis; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D004783:Environmental Microbiology; D004789:Enzyme Activation; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D009504:Neutrophils / Q000276:immunology / Q000382:microbiology; D011135:Polysaccharides, Bacterial / Q000096:biosynthesis*; D054730:Protein Interaction Domains and Motifs; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000302:isolation & purification / Q000502:physiology*
chemical_list: D000900:Anti-Bacterial Agents; D011135:Polysaccharides, Bacterial; D006026:Glycoside Hydrolases
keywords: Pel; Pseudomonas aeruginosa; Psl; biofilm; exopolysaccharide; glycoside hydrolase; therapeutic