Deciphering the network of microbial, environmental and physical factors underlying aggregation and enhanced performance in syntrophic microorganism communities

Project Information

SYNAG

Grant agreement ID: 948138

DOI

10.3030/948138

Project closed

EC signature date 23 October 2020

Start date 1 March 2021

End date 28 February 2026

Funded under

• EXCELLENT SCIENCE - European Research Council (ERC)

Total cost

€ 992 622,00

EU contribution

€ 992 622,00

Coordinated by

SVERIGES LANTBRUKSUNIVERSITET
Sweden

Periodic Reporting for period 3 - SYNAG (Deciphering the network of microbial, environmental and physical factors underlying aggregation and enhanced performance in syntrophic microorganism communities)

Reporting period: 2024-03-01 to 2025-08-31

Summary of the context and overall objectives of the project

Syntrophic microorganisms perform unique and crucial tasks in microbial processes used for sustainable energy and nutrient recovery from various organic waste streams. These syntrophic microorganisms cooperate to convert central intermediate compounds (acids) in the degradation to methane and these microbes are due to their restricted activity often a bottleneck that restricts productivity and process stability.

The main motivational factor for the research conducted in SYNAG is to contribute to the combat against climate change and to find paths for sustainable productions that will help us to pass on a healthier world to the next generation. Specifically, SYNAG has a clear purpose to improve the technologies needed for sustainable renewable energy, green products and sustainable fertiliser production. For this purpose, environmental conditions evaluated in the experiments conducted in SYNAG to high degree mimic the habitat residing in the biogas process, with the overall goal to find paths to improve the microbial degradation and the methane formation within the biogas process.

The aim of this project is to overcome the restrictions related to syntrophic activity, by bringing microorganisms into close proximity in multicellular aggregates (flocs). Syntrophic aggregation (SYNAG) has fundamental importance for syntrophic activity, but there is still much to learn in this area. To overcome the current knowledge gap, we study the regulatory processes that underlie aggregate formation and the acid degradation rate and evaluate how these two aspects are affected by surrounding environmental factors. The long-term overarching goal of SYNAG is to form a general model for aggregate development in syntrophic communities and to create a basis for novel process-design that will support key microorganisms and improve the productivity in biomethane processes.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The work performed in SYNAG has so far resulted in identification of a novel thermophilic syntrophic propionate oxidizing bacterium (SPOB ‘Candidatus Thermosyntrophopropionicum ammoniitolerans’) and a novel thermophilic syntrophic acetate oxidizing bacterium (SAOB). The expression-based analyses indicated use of both H2 and formate for electron transfer between the syntrophic bacteria and their cooperating methanogens. Furthermore, both the SPOB and the SAOB expressed genes for metallophosphoesterase characterised for its role in nanotube and biofilm formation and intercellular molecular exchange and cysteine synthase/O-acetylserine sulfhydrolase and sporulation proteins, which can be involved in biofilm formation. Expression of flagella proteins was not detected and the result instead indicated that the bacteria use pilus appendages for establishing physical contact with each other and the syntrophic methanogenic partner. The expression of quorum sensing and signalling mechanism-related genes by the syntrophic species were also profiled. Furthermore, in this syntrophic community it was found that the electron removal associated with syntrophic propionate and acetate oxidation was mediated by the hydrogen/formate-utilising methanogens Methanoculleus sp. and Methanothermobacter sp., with the latter observed to be critical for efficient propionate degradation. Similar dependence on Methanothermobacter was not seen for acetate degradation. This work has been published in The ISME Journal https://doi.org/10.1093/femsre/fuab057. A study, including cultivation and transcriptome-resolved analyses of syntrophic propionate and acetate conversion to methane at mesophilic and high ammonia conditions have been conducted and revealed activities connected to floc-formation. We also study the impact by acid-degradation rates by the syntrophic communities by the addition of supportive material. The manuscripts of these two studies are under preparation. In SYNAG, a review has also been published, summarizing basic and applied understanding of syntrophic propionate-oxidizing bacteria and highlighting knowledge gaps within the area. In this review we revealed distinctions in gene repertoire and organization for the pathway used for syntrophic propionate oxidation, hydrogenases and formate dehydrogenases, electron transfer mechanisms and environmental conditions governing propionate oxidation. This review has been published in the FEMS Microbiology Reviews https://www.nature.com/articles/s41396-023-01504-y.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The work conducted in SYNAG has so far resulted in identification of three novel species involved in syntrophic acid-degradation under high ammonia conditions, including two thermophilic syntrophic bacteria and a novel mesophilic methanogen. Hence, the results obtained in SYNAG such as the expression-based analyses conducted in both thermophilic and mesophilic syntrophic communities have taken us several steps closer to answering the question whether taxonomically distinct microorganisms can encode the same syntrophic metabolic function, or if this capacity is a metabolic niche occupied by a few distinct species. Another issue of relevance for SYNAG, is whether syntrophic communities share common mechanisms for aggregation or if the mechanisms are distinct depending on species. By using expression-based analyses, we have discovered divergent as well as reciprocal activities related to nanotube-formation, flagella, pilus appendages and quorum sensing in the thermophilic and the mesophilic syntrophic communities. Furthermore, the indication that Methanothermobacter sp. was observed to be critical for efficient propionate degradation by the thermophilic SPOB is also highly interesting and will be studied further. All the above-mentioned insights are highly beneficial for future research progress in SYNAG and will guide in the on-going and yet-to be started work to further uncover the genetic networks and mechanisms that trigger cell aggregation.

Progress has also been made in the construction of the fluidic system for cultivation of anaerobic microorganisms and a motorised camera system has been developed to monitor floc-formation during growth in anaerobic flasks. This system has revealed intriguing movements during growth of the syntrophic cultures, a time-laps video is available on this site: https://www.slu.se/en/departments/molecular-sciences/research-groups/microbial_biotechnology/project-synag/

At the end of the project, the expectation is that within SYNAG we have been able to obtain enough knowledge in order to pinpoint essential activities related to floc-formation in syntrophic communities active under high ammonia conditions. The prospect is also to increase the knowledge of the impact on acid-degrading capacities and methane-forming rate by the addition of supportive material and/or of varying environmental conditions of relevance for the biogas process.

Visualisation of the syntrophic interactions and work performed within SYNAG