Scott Hamilton-Brehm, Zhiwu Wang, Jennifer L. Morrell-Falvey, and James G. Elkins. BioEnergy Science Center, Biosciences Division, Oak Ridge National Laboratory, 1 Bethel Valley Road, Oak Ridge, TN 37831
Caldicellulosiruptor obsidiansis is a newly isolated, extremely thermophilic anaerobe from Yellowstone National Park and is capable of hydrolyzing pretreated lignocellulosic biomass (switchgrass or Populus), Avicel, xylan, cardboard, and filter paper. Fermentation experiments demonstrate that C. obsidiansis metabolizes C-5/C-6 sugars and produces small quantities of ethanol without the need to add support enzymes. In addition, this organism exhibits a high tolerance to the microbial inhibitory products created during the pretreatment process of lignocellulosic biomass. C. obsidiansis growth on insoluble substrates is closely associated with surface attachment. However, the mechanism of solid substrate utilization is still unknown. To begin testing this, we observed the spatial and temporal dynamics of microbial colonization of a model cellulose substrate using confocal laser scanning microscopy and 3D image reconstruction. The data indicate that C. obsidiansis colonies quickly expand horizontally and vertically on the cellulose surface, creating depressions in the substrate that eventually puncture a cellulose membrane within 72 hours. The thickness of the spreading biofilm stabilizes at a constant value, indicating carbon source diffusion from the substrate is limiting cell growth. Theoretical 2- and 3-dimensional simulations support this experimental observation. The distance between cells to the substrate surface played a crucial role in the conversion rate of the cellulose and thus the overall fermentation efficiency. By understanding the mechanism of microbial colonization of recalcitrant polymerized sugars, it may be possible to manipulate the fermentation conditions and target genetic modifications to improve the utilization efficiency of substrate carbon to produce desired end-products such as ethanol.
