Developing synthetic consortia for industrial biofuel production from lignocellulose

Abstract

This report describes work initiated at the Shell Biodomain Houston to evaluate candidate species for use as consolidated bioprocessing (CBP) hosts for conversion of lignocellulose (LC) to fuel. A systematic literature review regarding desirable characteristics for LC CBP hosts identified eight potential species: Bacillus subtilis 168, Cellulomonas fimi, Clostridium cellulolyticum, Clostridium thermocellum, Escherichia coli, Lactobacillus plantarum, Thermoanaerobacterium saccharolyticum and Thermobifida fusca. Commercially available API CH 50 kits were shown to be a rapid convenient method of testing the ability of all the species to ferment a wide range of carbohydrates. Analysis of batches cultured in the most important LC-derived sugars (D-glucose, cellobiose, D-xylose, L-arabinose) for biomass, ethanol, acetate and lactate production prompted T. fusca to be discarded as a potential host. This highlighted the high efficiency to which L. plantarum could achieve high product (lactic acid) yields. Physiological characterisation by imaging flow cytometry (IFC) established that it is possible to separate populations of different bacteria for the purpose of future sorting of complex consortia. Based on all these results a two-member microbial consortium was designed in which the biomass-to-model fuel product (lactate) pathway was shared among two species in co-culture with each species being specialised for specific processes i.e. C. fimi as the cellulolytic, and L. plantarum as the biofuel-producing specialist. For population monitoring of consortia IFC technology was utilised to establish a microbial demographic of C. fimi and L. plantarum separately and co-cultured together. Subsequently, a novel method to determine species abundance in synthetic consortia was developed allowing high-throughput testing. This could be used in industry for rapid process optimisation of synthetic consortia, and as an online monitoring and management system to detect consortium population-balance in industrial fermentations.