ERA-NET: Engineering microbial communities for the conversion of lignocellulose into medium-chain carboxylates

Microbial-based production of chemical compounds used in our daily life is a sustainable alternative to more traditional industrial processes like petrochemical production or to the extraction from natural resources. Microbes like bacteria and fungi have already been used for hundreds of years in the production of fermented foods like cheese, wine, and beer. Currently, the focus of microbial biotechnology is to find the best microbe that can convert a simple substrate, like sugar, into a chemical compound with economic value. To take this concept one step further, and make it even more sustainable, the next challenge is to replace the use of refined substrates with raw materials such as forest and agricultural residues. The aim of the Cell4Chem project was to design synthetic microbial consortia that can degrade lignocellulosic material and convert it into a class of compounds called medium-chain carboxylates. One of the main sources of such compounds is the palm oil industry, which is well-known for its harmful impact on natural ecosystems. Since this job is too demanding for a single microbe, used computational models to design an optimal team of microbes that can work together and divide all the required tasks. Similarly, our team consisted of five European partners, including computational biologists and experimentalists that worked together, exchanging data, models, and tools to reach this shared goal.

The aim of the Cell4Chem project was to convert sustainable resources such as lignocellulosic residues into useful chemicals, in particular the medium-chain carboxylates caproate and caprylate. Palm and coconut oil are currently the major sources of these two chemicals, which poses a significant environmental and social impact. We developed a genome-scale metabolic model for Ruminiclostridium cellulotyticum, one of the few species known to fully degrade both cellulose and hemicellulose. The model was extensively curated with experimental data obtained from the literature, covering approximately 25 years of research on this organism. Simulations of this model showed good agreement with experimental data for the fermentation of mixed lignocellulosic polysaccharides. Therefore, this organism is a promising microbial cell factory for sustainable transformation of lignocellulosic residues into valuable industrial products. Our experimental partners performed enrichment cultures from different isolation sources (manure, compost, soil) on lignocellulosic substrates. The enrichment was performed through serial passages over the course of several weeks, followed by metabolomics analysis and genome sequencing. Using these data, we built genome-scale metabolic models for all the members present in each enrichment culture. We used these models to simulate metabolic cross-feeding interactions within each community. These allowed to better understand the role of each community member and to find the most promising candidates for the assembly of a synthetic microbial community optimized for the production of our target compounds. Finally, we found serious limitations in all the community simulation methods that have been previously published. Therefore, we developed a new improved simulation method that showed good results for the enrichment cultures discussed above and for other datasets taken from the literature. The method was implemented into an open-source tool that is now publicly available for the scientific community.

One of the major challenges of today’s society is the shift from fossil-based industry towards renewable resources. The Cell4Chem project addresses this challenge by harnessing the power of microbial communities and enable transformation processes that result in high-value products from sustainable feedstocks. Medium-chain carboxylates (MCC) such as caproate and caprylate are specialty chemicals with broad application that can be produced by anaerobic fermentation of complex biomass. Currently, the utilization of sustainable feedstocks is mostly limited to biomass with high ethanol or lactate content, as such electron donors are crucial for reaching efficient MCC production. The exploitation of more abundant lignocellulosic biomass has the potential of greatly expanding the application of this new anaerobic fermentation technology, however, it harbors two major bottlenecks, the poor hydrolysis of cellulose and low internal production of lactate. Cell4Chem tackles these issues on three engineering levels. First, different strains will be genetically modified to create metabolic specialists for cellulose hydrolysis and lactate production. Second, these specialized strains will be combined into synthetic consortia with chain-elongating bacteria that can convert lactate into MCC. Third, anaerobic bioreactors will be operated with tailored upscaling strategies for the most promising designed consortia. The communities will be monitored over time using multi-omics methods in order to follow community dynamics and performance. These data will be further processed by bioinformatic tools to construct mechanistic microbial community models to elucidate metabolic interactions and screen for optimal community compositions.