Biofilm-Mediated Fragmentation and Degradation of Microcrystalline Cellulose by Cellulomonas flavigena KU (ATCC 53703)

Abstract

Cellulomonas flavigena KU (ATCC 53703) produces an extracellular matrix involved in the degradation of microcrystalline cellulose. This extracellular material is primarily composed of the gel-forming, β-1,3-glucan known as curdlan and associated, cellulose-degrading enzymes. In this study, the effects of various forms of nutrient limitation on cellulose attachment, cellular aggregation, curdlan production, and biofilm formation were investigated throughout a 7-day incubation period by using phase-contrast microscopy. Compared to cultures grown in non-limiting media, nitrogen-limitation promoted early attachment of C. flavigena KU cells to the cellulose surface, and cellulose attachment was congruent with cellular aggregation and curdlan production. Over the course of the experiment, microcolonies of attached cells grew into curdlan-producing biofilms on the cellulose. By contrast, bacterial cells grown on cellulose in non-limiting media remained unattached and unaggregated throughout most of the incubation period. By 7 days of incubation, bacterial aggregation was ninefold greater in N-limited cultures compared to nutritionally complete cultures. In a similar way, phosphorus- and vitamin-limitation (i.e., yeast extract-limitation) also resulted in early cellulose attachment and biofilm formation. Furthermore, nutrient limitation promoted more rapid and efficient fragmentation and degradation of cellulose, with cellulose fragments in low-N media averaging half the size of those in high-N media after 7 days. Two modes of cellulose degradation are proposed for C. flavigena KU, a "planktonic mode" and a "biofilm mode". Similar observations have been reported for other curdlan-producing cellulomonads, and these differing cellulose degradation strategies may ultimately prove to reflect sequential stages of a multifaceted biofilm cycle important in the bioconversion of this abundant and renewable natural resource.