Submitted to: Applied and Environmental Microbiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 10/2/2006
Publication Date: 12/7/2006
Citation: Weimer, P.J., Price, N.P., Kroukamp, O., Joubert, L.M., Wolfaardt, G.M., Van Zyl, W.H. 2006. Studies of the extracellular glycocalyx of the anaerobic ruminal bacterium Ruminococcus albus 7. Applied and Environmental Microbiology. 72(12):7559-7566. Interpretive Summary: Bacteria that degrade forage fiber in the rumen (the main fermentation chamber in the digestive tract of cattle) produce a substance that allows the bacteria to attach to the fiber for more effective degradation, and this substance has been shown to have novel properties that may allow its use as a commercial wood adhesive. Use of this material as a commercial adhesive would open new markets for materials derived from forages, but to do so would require that we develop methods to produce the material in larger amounts, and in a form with maximum adhesive performance properties. We used microscopic and chemical methods to characterize the adhesive produced by one of the most important fiber degrading bacterial species in the rumen. The adhesive material contains large amounts of different types of sugar molecules arranged in a unique fashion to form a polymer that connects individual bacterial cells to one another and to the fiber surface. Production of the adhesive material requires only a modest energy expenditure by the bacterium. These results will benefit researchers by providing clues on how the bacteria produce the adhesive, which will allow us to increase the amount and quality of the adhesive that the bacteria make.
Technical Abstract: Anaerobic cellulolytic bacteria are thought to adhere to cellulose via several mechanisms, one of which is the production of a glycocalyx containing extracellular polymeric substances (EPS). As the composition and structure of these glycocalyces have not been elucidated, a combination of scanning electron microscopy (SEM) and chemical analysis was used to characterize the glycocalyx of Ruminococcus albus strain 7. Variable-pressure SEM revealed that growth of this strain was accompanied by formation of thin, pilus-like cellular extensions that adhered 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 [B] + glycocalyx [G]+ 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 (ND) removed almost all of the adherent cells and protein while retaining a residual network of adhering noncellular material. Subsequent treatment of this residue (G + C) with 2 N trifluoroacetic acid (120 oC, 90 min) resulted in complete removal of the glycocalyx. This treatment released primarily glucose, along with substantial amounts of xylose and mannose, but only traces of galactose, the most abundant sugar in the majority of characterized bacterial exopolysaccharides. Linkage analysis and characterization by NMR suggested that most of the glucosyl units were not present as partially degraded cellulose. Calculations, based on the amount of carbohydrate and protein removed by these chemical treatments, suggest that the energetic demand for synthesis of the non-protein fraction of EPS by this organism represents only a small fraction (< 4%) of the anabolic ATP expenditure of the culture.