Tracking the cellulolytic activity of Clostridium thermocellum biofilms

Biotechnology for Biofuels

Alexandru Dumitrache³  Gideon M Wolfaardt⁴  David Grant Allen³  Steven N Liss¹  Lee R Lynd² 

[1] School of Environmental Studies and Department of Chemical Engineering, Queen’s University, 99 University Ave., Kingston, ON K7L 3N6, Canada

[2] Thayer School of Engineering, Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA

[3] Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College St., Toronto, ON M5S 3E1, Canada

[4] Stellenbosch Institute for Advanced Study Wallenberg Research Centre at Stellenbosch University, Stellenbosch 7600, South Africa

关键词: Clostridium thermocellum;    CO₂ production;    Cellulose conversion;    Biofilm kinetics;   

Others  :  794440
DOI  :  10.1186/1754-6834-6-175

 received in 2013-07-11, accepted in 2013-10-02,  发布年份 2013

【 摘 要 】

Background

Microbial cellulose conversion by Clostridium thermocellum 27405 occurs predominantly through the activity of substrate-adherent bacteria organized in thin, primarily single cell-layered biofilms. The importance of cellulosic surface exposure to microbial hydrolysis has received little attention despite its implied impact on conversion kinetics.

Results

We showed the spatial heterogeneity of fiber distribution in pure cellulosic sheets, which made direct measurements of biofilm colonization and surface penetration impossible. Therefore, we utilized on-line measurements of carbon dioxide (CO₂) production in continuous-flow reactors, in conjunction with confocal imaging, to observe patterns of biofilm invasion and to indirectly estimate microbial accessibility to the substrate’s surface and the resulting limitations on conversion kinetics. A strong positive correlation was found between cellulose consumption and CO₂ production (R² = 0.996) and between surface area and maximum biofilm activity (R² = 0.981). We observed an initial biofilm development rate (0.46 h^-1, 0.34 h^-1 and 0.33 h^-1) on Whatman sheets (#1, #598 and #3, respectively) that stabilized when the accessible surface was maximally colonized. The results suggest that cellulose conversion kinetics is initially subject to a microbial limitation period where the substrate is in excess, followed by a substrate limitation period where cellular mass, in the form of biofilms, is not limiting. Accessible surface area acts as an important determinant of the respective lengths of these two distinct periods. At end-point fermentation, all sheets were digested predominantly under substrate accessibility limitations (e.g., up to 81% of total CO₂ production for Whatman #1). Integration of CO₂ production rates over time showed Whatman #3 underwent the fastest conversion efficiency under microbial limitation, suggestive of best biofilm penetration, while Whatman #1 exhibited the least recalcitrance and the faster degradation during the substrate limitation period.

Conclusion

The results showed that the specific biofilm development rate of cellulolytic bacteria such as C. thermocellum has a notable effect on overall reactor kinetics during the period of microbial limitation, when ca. 20% of cellulose conversion occurs. The study further demonstrated the utility of on-line CO₂ measurements as a method to assess biofilm development and substrate digestibility pertaining to microbial solubilization of cellulose, which is relevant when considering feedstock pre-treatment options.

【 授权许可】

   
2013 Dumitrache et al.; licensee BioMed Central Ltd.

【 图 表 】

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