Role of Gradient Microenvironments in Supporting Termite Gut Community Diversity

Termites can metabolize woody materials for energy, but no conventional biofuel production process has been able to emulate this functionality with comparable efficiency. The lower termite Reticulitermes flavipes is able to utilize 74-99% of the cellulose and 65-87% of the hemicellulose from lignocellulose with the help of microbes living symbiotically in the gut. If these microbes can be cultured in vitro, then they may be used in an engineered bioprocess to convert woody waste materials into useful products including biofuels and chemical manufacturing feedstocks. The hindgut of the lower termite Reticulitermes flavipes is a diverse microenvironment containing both aerobic and anaerobic regions in close physical proximity. Here we describe development of microhabitat engineering systems to measure survival of constituents of the termite gut microbiome as a function of oxygen gradient conditions. Replicate microhabitats were designed with either varying or constant oxygen conditions within the range from 0-10% equilibrium oxygen saturation in water. Various oxygen saturations were prepared using a microfluidic gradient mixer, and gradients were created by diffusion of oxygen from different concentration feeds through PDMS. Devices were loaded with a mixed microbial community collected from the hindgut of Reticulitermes flavipes, then viable protists were enumerated over for 24 hours. Oxygen concentrations were monitored optically using an oxygen-quenching fluorescent reporting molecule embedded in a polystyrene film. This work provides a basis for future developments in gradient bioengineering using the Reticulitermes flavipes microbiome, and may contribute to the sustainable production of biofuels from waste biomass.