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

Mikrobe basert produksjon av kjemiske forbindelser som brukes i vårt daglige liv er et bærekraftig alternativ til mer tradisjonelle industrielle prosesser som petrokjemisk produksjon eller utvinning av naturressurser. Mikrober som bakterier og sopp har allerede blitt brukt i hundrevis av år i produksjonen av fermenterte matvarer som ost, vin og øl. Nå er fokuset for mikrobiell bioteknologi å finne den beste mikroben som kan omdanne et enkelt substrat, som sukker, til en kjemisk forbindelse med økonomisk verdi. For å ta dette konseptet et skritt videre, og gjøre det enda mer bærekraftig, er neste utfordring å erstatte bruken av prosseserte materialer med råvarer som skog og jordbruksrester. Cell4Chem-prosjektet designet syntetiske mikrobielle konsortier som kan bryte ned lignocellulose og konvertere det til en klasse forbindelser som kalles mellomstore karboksylater. En av hovedkildene til slike forbindelser er palmeoljeindustrien, som er kjent for sin skadelige innvirkning på naturlige økosystemer. Siden denne jobben er for krevende for en enkelt mikrobe, brukte vi beregningsmodeller for å designe et perfekt team av mikrober som kan jobbe sammen og dele alle nødvendige oppgaver. På samme måte bestod teamet vårt av fem europeiske partnere, inkludert beregnings og eksperimentelle biologer som jobbet sammen, utveksle data, modeller og verktøy for å nå dette delte målet.

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.