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Applied and Environmental Microbiology, December 2006, p. 7559-7566, Vol. 72, No. 12
0099-2240/06/$08.00+0     doi:10.1128/AEM.01632-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Studies of the Extracellular Glycocalyx of the Anaerobic Cellulolytic Bacterium Ruminococcus albus 7

Paul J. Weimer,^1,2* Neil P. J. Price,³ Otini Kroukamp,^4, Lydia-Marie Joubert,^4, Gideon M. Wolfaardt,^4, and Willem H. Van Zyl⁴

U.S. Dairy Forage Research Center, Agricultural Research Service, U.S. Department of Agriculture, Madison, Wisconsin,¹ Department of Bacteriology, University of Wisconsin—Madison, Madison, Wisconsin,² National Center for Agricultural Utilization Research, Agricultural Research Service, U.S. Department of Agriculture, Peoria, Illinois,³ Department of Microbiology, University of Stellenbosch, Stellenbosch, South Africa⁴

Received 13 July 2006/ Accepted 2 October 2006

Anaerobic cellulolytic bacteria are thought to adhere to cellulose via several mechanisms, including production of a glycocalyx containing extracellular polymeric substances (EPS). As the compositions and structures of these glycocalyces have not been elucidated, variable-pressure scanning electron microscopy (VP-SEM) and chemical analysis were used to characterize the glycocalyx of the ruminal bacterium Ruminococcus albus strain 7. VP-SEM revealed that growth of this strain was accompanied by the formation of thin cellular extensions that allowed the bacterium to adhere to cellulose, followed by formation of a ramifying network that interconnected individual cells to one another and to the unraveling cellulose microfibrils. Extraction of 48-h-old whole-culture pellets (bacterial cells plus glycocalyx [G] plus residual cellulose [C]) with 0.1 N NaOH released carbohydrate and protein in a ratio of 1:5. Boiling of the cellulose fermentation residue in a neutral detergent solution removed almost all of the adherent cells and protein while retaining a residual network of adhering noncellular material. Trifluoroacetic acid hydrolysis of this residue (G plus C) released primarily glucose, along with substantial amounts of xylose and mannose, but only traces of galactose, the most abundant sugar in most characterized bacterial exopolysaccharides. Linkage analysis and characterization by nuclear magnetic resonance suggested that most of the glucosyl units were not present as partially degraded cellulose. Calculations suggested that the energy demand for synthesis of the nonprotein fraction of EPS by this organism represents only a small fraction (<4%) of the anabolic ATP expenditure of the bacterium.

Published ahead of print on 6 October 2006.

Present address: Department of Chemistry and Biology, Ryerson University, Toronto, Ontario, Canada.

Present address: Cell Sciences Imaging Facility, Stanford University, Stanford, Calif.