Consolidated Bioprocessing of Cellulose to Itaconic Acid by a Co-Culture of Trichoderma Reesei and Ustilago Maydis.

Abstract

Background: Itaconic acid is a bio-derived platform chemical with uses ranging from polymer synthesis to biofuel production. The efficient conversion of cellulosic waste streams into itaconic acid could thus enable the sustainable production of a variety of substitutes for fossil oil based products. However, the realization of such a process is currently hindered by an expensive conversion of cellulose into fermentable sugars. Here, we present the stepwise development of a fully consolidated bioprocess (CBP), which is capable to directly convert recalcitrant cellulose into itaconic acid without the need for separate cellulose hydrolysis including the application of commercial cellulases. The process is based on a synthetic microbial consortium of the cellulase producer Trichoderma reesei and the itaconic acid producing yeast Ustilago maydis. Results: The efficiency of the process was compared to a simultaneous hydrolysis and fermentation setup (SSF). Because of the additional substrate consumption of T. reesei in the CBP, the itaconic acid yield was significantly lower in the CBP than in the SSF. In order to increase yield and productivity of itaconic acid in the CBP, the population dynamics was manipulated by varying the inoculation delay between T. reesei and U. maydis. Surprisingly, neither inoculation delay nor inoculation density significantly affected the population development or the CBP performance. Instead, the substrate availability was the most important parameter. U. maydis was only able to grow and to produce itaconic acid when the cellulose concentration and thus, the sugar supply rate, was high. Finally, the metabolic processes during fed-batch CBP were analyzed in depth by online respiration measurements. A method was developed to estimate cellulose consumption, itaconic acid formation as well as the actual itaconic acid production yield online. Again, substrate availability was the key factor also controlling itaconic acid yield. In summary, an itaconic acid titer of 34 g/L with a total productivity of up to 0.07 g/L/h and a yield of 0.16 g/g could be reached during fed-batch cultivation. Conclusion: This study demonstrates the feasibility of consortium based CBP for itaconic acid production and also lays the fundament for the development and improvement of similar microbial consortia for cellulose based organic acid production.

Defined Microbial Mixed Culture for Utilization of Polyurethane Monomers

“…Defined mixed cultures can be organized as presented here, where the catalytic tasks are distributed to the individual members, that is, the conversion of the different monomers are fulfilled by dedicated strainswith a single objective, that is, to synthesize rhamnolipids. Such a mixed culture can be also designed such that the individual members depend on each other; for example, in the study of Schlembach et al, cellulose hydrolysis and product synthesis (itaconic acid as an example) were distributed to Trichoderma reesei and an engineered Ustilago maydis strain, respectively . Finally, the tasks of substrate conversion and product formation can be consolidated into a single strain.…”

“…Such a mixed culture can be also designed such that the individual members depend on each other; for example, in the study of Schlembach et al, cellulose hydrolysis and product synthesis (itaconic acid as an example) were distributed to Trichoderma reesei and an engineered Ustilago maydis strain, respectively. 53 Finally, the tasks of substrate conversion and product formation can be consolidated into a single strain. While, for decades, such strains were aimed in lignocellulosic biotechnology; for example, for bioethanol production, no such strain has been competitive until now.…”

Tunable population dynamics in a synthetic filamentous coculture

“…It serves as an excellent indicator of substrate consumption dynamics and can be measured noninvasively and online. It has been proven a reliable method for evaluating cellulase formation and cellulose consumption in axenic cultures, as well as defined cocultures (Antonov et al, 2016(Antonov et al, , 2017Schlembach et al, 2020). Although the OTR only delivers a sum signal of the coculture respiration activity, the metabolic contribution of each partner can be estimated by comparison to axenic cultures.…”