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Analysis of the Aspergillus fumigatus Biofilm Extracellular Matrix by Solid-State Nuclear Magnetic Resonance Spectroscopy

Courtney Reichhardt, Jose A. G. Ferreira, Lydia-Marie Joubert, Karl V. Clemons, David A. Stevens, Lynette Cegelski
Courtney Reichhardt
aDepartment of Chemistry, Stanford University, Stanford, California, USA
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Jose A. G. Ferreira
bDepartment of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, California, USA
cCalifornia Institute for Medical Research, San Jose, California, USA
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Lydia-Marie Joubert
dCell Sciences Imaging Facility, Stanford University School of Medicine, Stanford, California, USA
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Karl V. Clemons
bDepartment of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, California, USA
cCalifornia Institute for Medical Research, San Jose, California, USA
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David A. Stevens
bDepartment of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, California, USA
cCalifornia Institute for Medical Research, San Jose, California, USA
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Lynette Cegelski
aDepartment of Chemistry, Stanford University, Stanford, California, USA
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DOI: 10.1128/EC.00050-15
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ABSTRACT

Aspergillus fumigatus is commonly responsible for lethal fungal infections among immunosuppressed individuals. A. fumigatus forms biofilm communities that are of increasing biomedical interest due to the association of biofilms with chronic infections and their increased resistance to antifungal agents and host immune factors. Understanding the composition of microbial biofilms and the extracellular matrix is important to understanding function and, ultimately, to developing strategies to inhibit biofilm formation. We implemented a solid-state nuclear magnetic resonance (NMR) approach to define compositional parameters of the A. fumigatus extracellular matrix (ECM) when biofilms are formed in RPMI 1640 nutrient medium. Whole biofilm and isolated matrix networks were also characterized by electron microscopy, and matrix proteins were identified through protein gel analysis. The 13C NMR results defined and quantified the carbon contributions in the insoluble ECM, including carbonyls, aromatic carbons, polysaccharide carbons (anomeric and nonanomerics), aliphatics, etc. Additional 15N and 31P NMR spectra permitted more specific annotation of the carbon pools according to C-N and C-P couplings. Together these data show that the A. fumigatus ECM produced under these growth conditions contains approximately 40% protein, 43% polysaccharide, 3% aromatic-containing components, and up to 14% lipid. These fundamental chemical parameters are needed to consider the relationships between composition and function in the A. fumigatus ECM and will enable future comparisons with other organisms and with A. fumigatus grown under alternate conditions.

FOOTNOTES

    • Received 17 March 2015.
    • Accepted 6 July 2015.
    • Accepted manuscript posted online 10 July 2015.
  • Supplemental material for this article may be found at http://dx.doi.org/10.1128/EC.00050-15.

  • Copyright © 2015, American Society for Microbiology. All Rights Reserved.
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Analysis of the Aspergillus fumigatus Biofilm Extracellular Matrix by Solid-State Nuclear Magnetic Resonance Spectroscopy
Courtney Reichhardt, Jose A. G. Ferreira, Lydia-Marie Joubert, Karl V. Clemons, David A. Stevens, Lynette Cegelski
Eukaryotic Cell Oct 2015, 14 (11) 1064-1072; DOI: 10.1128/EC.00050-15

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Analysis of the Aspergillus fumigatus Biofilm Extracellular Matrix by Solid-State Nuclear Magnetic Resonance Spectroscopy
Courtney Reichhardt, Jose A. G. Ferreira, Lydia-Marie Joubert, Karl V. Clemons, David A. Stevens, Lynette Cegelski
Eukaryotic Cell Oct 2015, 14 (11) 1064-1072; DOI: 10.1128/EC.00050-15
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