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Microbial glycoside hydrolases as antibiofilm agents with cross-kingdom activity

Brendan D. Snarr, Perrin Baker, Natalie C. Bamford, Yukiko Sato, Hong Liu, Melanie Lehoux, Fabrice N. Gravelat, Hanna Ostapska, Shane R. Baistrocchi, Robert P. Cerone, Elan E. Filler, Matthew R. Parsek, Scott G. Filler, P. Lynne Howell, Donald C. Sheppard
doi: https://doi.org/10.1101/113696
Brendan D. Snarr
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Perrin Baker
bProgram in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario, M5G 1X8, Canada
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Natalie C. Bamford
bProgram in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario, M5G 1X8, Canada
cDepartment of Biochemistry, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
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Yukiko Sato
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Hong Liu
dLos Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, CA, USA
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Melanie Lehoux
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Fabrice N. Gravelat
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Hanna Ostapska
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Shane R. Baistrocchi
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Robert P. Cerone
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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Elan E. Filler
dLos Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, CA, USA
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Matthew R. Parsek
eDepartment of Microbiology, University of Washington, Seattle, Washington, USA
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Scott G. Filler
dLos Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, CA, USA
fDavid Geffen School of Medicine at University of California, Los Angeles, CA, USA
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P. Lynne Howell
bProgram in Molecular Structure & Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario, M5G 1X8, Canada
cDepartment of Biochemistry, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
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  • For correspondence: don.sheppard@mcgill.ca howell@sickkids.ca
Donald C. Sheppard
aDepartments of Medicine, Microbiology and Immunology; Infectious Diseases in Global Health Program, Centre for Translational Biology, McGill University Health Centre. Montréal, Québec, H4A 3J1, Canada
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  • For correspondence: don.sheppard@mcgill.ca howell@sickkids.ca
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Abstract

Galactosaminogalactan and Pel are cationic heteropolysaccharides produced by the opportunistic pathogens, Aspergillus fumigatus and Pseudomonas aeruginosa, respectively. These exopolysaccharides both contain 1,4-linked N-acetyl-D-galactosamine and play an important role in biofilm formation by these organisms. Proteins containing glycoside hydrolase domains have recently been identified within the biosynthetic pathway of each exopolysaccharide. Recombinant hydrolase domains from these proteins (Sph3h from A. fumigatus and PelAh from P. aeruginosa) were found to degrade their respective polysaccharides in vitro. We therefore hypothesized that these glycoside hydrolases could exhibit anti-biofilm activity, and further, given the chemical similarity between galactosaminogalactan and Pel, that they might display cross-species activity. Treatment of A. fumigatus with Sph3h disrupted A. fumigatus biofilms with an EC50 of 0.4 nM. PelAh treatment also disrupted pre-formed A. fumigatus biofilms with EC50 values similar to those obtained for Sph3h. In contrast, Sph3h was unable to disrupt P. aeruginosa Pel-based biofilms, despite being able to bind to the exopolysaccharide. Treatment of A. fumigatus hyphae with either Sph3h or PelAh significantly enhanced the activity of the antifungals posaconazole, amphotericin B and caspofungin, likely through increasing antifungal penetration of hyphae. Both enzymes were non-cytotoxic and protected A549 pulmonary epithelial cells from A. fumigatus-induced cell damage for up to 24 hours. Intratracheal administration of Sph3h was well tolerated, and reduced pulmonary fungal burden in a neutropenic mouse model of invasive aspergillosis. These findings suggest that glycoside hydrolases can exhibit activity against diverse microorganisms and may be useful as therapeutic agents by degrading biofilms and attenuating virulence.

Significance The production of biofilms is an important strategy used by both bacteria and fungi to colonize surfaces and to enhance resistance to killing by immune cells and antimicrobial agents. We demonstrate that glycoside hydrolases derived from the opportunistic fungus Aspergillus fumigatus and Gram-negative bacterium Pseudomonas aeruginosa can be exploited to disrupt pre-formed fungal biofilms and reduce virulence. Additionally, these glycoside hydrolases can be utilized to potentiate antifungal drugs by increasing their hyphal penetration, to protect human cells from fungal-induced injury and to attenuate virulence of A. fumigatus in a mouse model of invasive aspergillosis. The findings of this study identify recombinant microbial glycoside hydrolases as promising therapeutics with the potential for anti-biofilm activity against pathogens across different taxonomic kingdoms.

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Posted March 04, 2017.
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Microbial glycoside hydrolases as antibiofilm agents with cross-kingdom activity
Brendan D. Snarr, Perrin Baker, Natalie C. Bamford, Yukiko Sato, Hong Liu, Melanie Lehoux, Fabrice N. Gravelat, Hanna Ostapska, Shane R. Baistrocchi, Robert P. Cerone, Elan E. Filler, Matthew R. Parsek, Scott G. Filler, P. Lynne Howell, Donald C. Sheppard
bioRxiv 113696; doi: https://doi.org/10.1101/113696
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Microbial glycoside hydrolases as antibiofilm agents with cross-kingdom activity
Brendan D. Snarr, Perrin Baker, Natalie C. Bamford, Yukiko Sato, Hong Liu, Melanie Lehoux, Fabrice N. Gravelat, Hanna Ostapska, Shane R. Baistrocchi, Robert P. Cerone, Elan E. Filler, Matthew R. Parsek, Scott G. Filler, P. Lynne Howell, Donald C. Sheppard
bioRxiv 113696; doi: https://doi.org/10.1101/113696

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