19-ERACoBioTech SYNBIOGAS: Synthetic landfill microbiomes for enhanced anaerobic digestion to biogas
Lead Research Organisation:
Bangor University
Department Name: Sch of Natural Sciences
Abstract
Lignocellulosic plant biomass is the most abundant waste product generated by society, agriculture and industry. By 2025, global cities will
generate approximately 2.2 billion tonnes of solid waste biomass per year, with significant impacts upon health and the economy at both
local and global scales. Natural communities of microorganisms (microbiomes) convert waste biomass to methane-rich biogas that can be
used as a sustainable and renewable green-energy source to generate electricity, heat and power, and biomethane for injection into the
national gas grid and production of transport fuels. Anaerobic digestion (AD) plants and landfill sites are engineered environments where
these microbial processes are harnessed for waste decomposition and biogas production. The EU is the largest global producer of biogas
from biomass, with over 17,000 AD plants, and consequently, the microbiological conversion of solid waste residues to biogas in AD plants
and landfill sites presents an unprecedented opportunity to leverage key enabling technologies for a sustainable bio-based economy for
green-energy production. In turn, conversion of waste biomass to biomethane will mitigate the escalating environmental and social impacts
of waste residues. However, the metabolic function of microorganisms responsible for anaerobic digestion is poorly understood, and most
previous studies have focused on animal gut microorganisms that are typically used to incoluate microorganisms into anaerobic digestion
plants as slurry. One of the major bottlenecks to industrial application of microorganisms for biomass-conversion is low substrate specificity,
low temperature tolerance, and an inability to perform optimally under reaction conditions. Natural microorganisms found in landfill sites
represent an unexplored repository of biomass-degrading enzyme diversity with the potential to enhance existing industrial
biomass-conversion processes. Landfill microorganisms are already adapted to engineered environments, mineralise diverse solid waste
types, produce methane-rich biogas, and are therefore good candidates for the bioaugmentation of anaerobic digestion processes. The SYNBIOGAS consortium is an academic-industry partnership that will integrate diverse and cutting-edge technological, analytical,
engineering and computational approaches for characterisation of the landfill biomass-degrading microbiome. Microbial isolations, DNA
sequencing, enzyme characterisation and computational modelling of landfill microbial biomass-conversion processes will inform the design
and validation of optimised synthetic landfill microbiomes (SLMs) for enhanced waste biomass-conversion in AD plants and landfill sites, and
to develop applications of the SLM that can be readily adopted by industry. Engineering biomass-degrading microbiomes is a new research
frontier with many novel applications, including bioaugmentation and optimisation of biomass conversion in AD and landfill systems towards
an enhanced bio-based economy for waste management, environmental protection, and sustainable intensification of renewable energy
generation.
generate approximately 2.2 billion tonnes of solid waste biomass per year, with significant impacts upon health and the economy at both
local and global scales. Natural communities of microorganisms (microbiomes) convert waste biomass to methane-rich biogas that can be
used as a sustainable and renewable green-energy source to generate electricity, heat and power, and biomethane for injection into the
national gas grid and production of transport fuels. Anaerobic digestion (AD) plants and landfill sites are engineered environments where
these microbial processes are harnessed for waste decomposition and biogas production. The EU is the largest global producer of biogas
from biomass, with over 17,000 AD plants, and consequently, the microbiological conversion of solid waste residues to biogas in AD plants
and landfill sites presents an unprecedented opportunity to leverage key enabling technologies for a sustainable bio-based economy for
green-energy production. In turn, conversion of waste biomass to biomethane will mitigate the escalating environmental and social impacts
of waste residues. However, the metabolic function of microorganisms responsible for anaerobic digestion is poorly understood, and most
previous studies have focused on animal gut microorganisms that are typically used to incoluate microorganisms into anaerobic digestion
plants as slurry. One of the major bottlenecks to industrial application of microorganisms for biomass-conversion is low substrate specificity,
low temperature tolerance, and an inability to perform optimally under reaction conditions. Natural microorganisms found in landfill sites
represent an unexplored repository of biomass-degrading enzyme diversity with the potential to enhance existing industrial
biomass-conversion processes. Landfill microorganisms are already adapted to engineered environments, mineralise diverse solid waste
types, produce methane-rich biogas, and are therefore good candidates for the bioaugmentation of anaerobic digestion processes. The SYNBIOGAS consortium is an academic-industry partnership that will integrate diverse and cutting-edge technological, analytical,
engineering and computational approaches for characterisation of the landfill biomass-degrading microbiome. Microbial isolations, DNA
sequencing, enzyme characterisation and computational modelling of landfill microbial biomass-conversion processes will inform the design
and validation of optimised synthetic landfill microbiomes (SLMs) for enhanced waste biomass-conversion in AD plants and landfill sites, and
to develop applications of the SLM that can be readily adopted by industry. Engineering biomass-degrading microbiomes is a new research
frontier with many novel applications, including bioaugmentation and optimisation of biomass conversion in AD and landfill systems towards
an enhanced bio-based economy for waste management, environmental protection, and sustainable intensification of renewable energy
generation.
Technical Summary
The landfill microbiome represents an unexplored repository of biomass-degrading enzyme diversity to enhance existing industrial
biomass-conversion processes and identify new hydrolase enzymes of relevance for industrial biotechnology processes. This project aims to utilise a systems biology analysis of biomass-conversion by landfill microbiota, combining novel technological, analytical and computational approaches for microbiome characterisation, in silico discovery and validation of novel enzymes, and process modelling for the design of optimal synthetic biomass-converting microbiomes. Ultimately, the research will generate synthetic landfill microbiomes (SLMs) designed for bioaugmentation of landfill sites and anaerobic digestion plants for enhanced biomass conversion and biogas generation, enabling a progression in TRL in this sector. In addition, we will utilise life cycle assessment and cost benefit analysis approaches to demonstrate the potential industrial benefits of our process model and synthetic microbiome, and will generate a road map for industry adoption of the new technology. The research leverages new approaches to understanding fundamental questions regarding the ecological factors that drive syntrophic interactions between anaerobic biomass-degrading microbiota.
biomass-conversion processes and identify new hydrolase enzymes of relevance for industrial biotechnology processes. This project aims to utilise a systems biology analysis of biomass-conversion by landfill microbiota, combining novel technological, analytical and computational approaches for microbiome characterisation, in silico discovery and validation of novel enzymes, and process modelling for the design of optimal synthetic biomass-converting microbiomes. Ultimately, the research will generate synthetic landfill microbiomes (SLMs) designed for bioaugmentation of landfill sites and anaerobic digestion plants for enhanced biomass conversion and biogas generation, enabling a progression in TRL in this sector. In addition, we will utilise life cycle assessment and cost benefit analysis approaches to demonstrate the potential industrial benefits of our process model and synthetic microbiome, and will generate a road map for industry adoption of the new technology. The research leverages new approaches to understanding fundamental questions regarding the ecological factors that drive syntrophic interactions between anaerobic biomass-degrading microbiota.
Planned Impact
The landfill microbiome represents an unexplored repository of biomass-degrading enzyme diversity to enhance existing industrial
biomass-conversion processes and identify new hydrolase enzymes of relevance for industrial biotechnology processes (Ransom-Jones et al.,
2017). This project will provide the first systems biology analysis of biomass-conversion by landfill microbiota (WP1-3) combining novel
technological, analytical and computational approaches for microbiome characterisation (WP1), in silico discovery and validation of novel
enzymes (WP2) and process modelling for the design of optimal synthetic biomass-converting microbiomes (WP3). Ultimately, the research
in WP's 1-3 will generate synthetic landfill microbiomes (SLMs) designed for bioaugmentation of landfill sites and anaerobic digestion
plants for enhanced biomass conversion and biogas generation in WP4, enabling a progression from TRL2 to TRL6 through the project
(Figure 2). In WP5, we subsequently utilise life cycle assessment and cost benefit analysis approaches to demonstrate the potential
industrial benefits of our process model and synthetic microbiome, and will generate a road map for industry adoption of the new
technology. The research leverages new approaches to understanding fundamental questions regarding the ecological factors
that drive syntrophic interactions between anaerobic biomass-degrading microbiota.
Synthetic biology approaches for the engineering of biomass-degrading microbiomes is a new research frontier with many
novel applications, including bioaugmentation and optimisation of biomass conversion in AD systems towards an enhanced bio-based
economy for waste management, environmental protection and sustainable intensification of renewable energy generation.
Previously, laboratory-based landfill bioreactor experiments undergoing bioaugmentation with natural compost microorganisms demonstrated improved biomass degradation from 65% to 99%, and increased biogas generation (Kinet et al., 2016). The success of
bioaugmentation for biomass conversion with relatively undefined microbial populations from natural environments in laboratory reactors is
encouraging; however, such approaches have not been extensively validated and demonstrated in relevant industrial environments (i.e. AD
plants and landfills), and SLMs have not previously been designed and tested. Consequently, the opportunity to (i) directly manipulate
biomass-converting microbiota through characterisation, process modelling and the design of more sophisticated synthetic microbiomes,
and (ii), industrial application/validation of SLMs for enhanced biogas generation, in the SYNBIOGAS project is a tantalising challenge, with
significant potential to provide a paradigm shift in waste biomass conversion, green-energy production and waste management.
The SYNBIOGAS project is directly relevant to the CoBioTech call, and will develop a biotechnological application (synthetic landfill
microbial communities for enhanced biomass conversion to biogas) with potential for strong economic and social impacts towards at
sustainable bio-based economy. Our integration and development of the approach with industry partners ensures that our research has high
potential for commercialisation and industry adoption through achieving high levels of technology readiness (TRL 2-6). Social impacts
include sustainable options for waste management, reduce environmental impact of waste biomass, and the development of key enabling
technologies for sustainable green-energy generation with key economic benefits. Tangible outputs would include: novel CAZYmes with
enhanced catalytic activity and substrate specificity; high resolution datasets on SLM activity; world-leading metabolic process models of
anaerobic digestion processes; validated biotechnological applications of SLMs for AD bioaugmentation; life cycle assessment and a
stakeholder roadmap for technology implementation.
biomass-conversion processes and identify new hydrolase enzymes of relevance for industrial biotechnology processes (Ransom-Jones et al.,
2017). This project will provide the first systems biology analysis of biomass-conversion by landfill microbiota (WP1-3) combining novel
technological, analytical and computational approaches for microbiome characterisation (WP1), in silico discovery and validation of novel
enzymes (WP2) and process modelling for the design of optimal synthetic biomass-converting microbiomes (WP3). Ultimately, the research
in WP's 1-3 will generate synthetic landfill microbiomes (SLMs) designed for bioaugmentation of landfill sites and anaerobic digestion
plants for enhanced biomass conversion and biogas generation in WP4, enabling a progression from TRL2 to TRL6 through the project
(Figure 2). In WP5, we subsequently utilise life cycle assessment and cost benefit analysis approaches to demonstrate the potential
industrial benefits of our process model and synthetic microbiome, and will generate a road map for industry adoption of the new
technology. The research leverages new approaches to understanding fundamental questions regarding the ecological factors
that drive syntrophic interactions between anaerobic biomass-degrading microbiota.
Synthetic biology approaches for the engineering of biomass-degrading microbiomes is a new research frontier with many
novel applications, including bioaugmentation and optimisation of biomass conversion in AD systems towards an enhanced bio-based
economy for waste management, environmental protection and sustainable intensification of renewable energy generation.
Previously, laboratory-based landfill bioreactor experiments undergoing bioaugmentation with natural compost microorganisms demonstrated improved biomass degradation from 65% to 99%, and increased biogas generation (Kinet et al., 2016). The success of
bioaugmentation for biomass conversion with relatively undefined microbial populations from natural environments in laboratory reactors is
encouraging; however, such approaches have not been extensively validated and demonstrated in relevant industrial environments (i.e. AD
plants and landfills), and SLMs have not previously been designed and tested. Consequently, the opportunity to (i) directly manipulate
biomass-converting microbiota through characterisation, process modelling and the design of more sophisticated synthetic microbiomes,
and (ii), industrial application/validation of SLMs for enhanced biogas generation, in the SYNBIOGAS project is a tantalising challenge, with
significant potential to provide a paradigm shift in waste biomass conversion, green-energy production and waste management.
The SYNBIOGAS project is directly relevant to the CoBioTech call, and will develop a biotechnological application (synthetic landfill
microbial communities for enhanced biomass conversion to biogas) with potential for strong economic and social impacts towards at
sustainable bio-based economy. Our integration and development of the approach with industry partners ensures that our research has high
potential for commercialisation and industry adoption through achieving high levels of technology readiness (TRL 2-6). Social impacts
include sustainable options for waste management, reduce environmental impact of waste biomass, and the development of key enabling
technologies for sustainable green-energy generation with key economic benefits. Tangible outputs would include: novel CAZYmes with
enhanced catalytic activity and substrate specificity; high resolution datasets on SLM activity; world-leading metabolic process models of
anaerobic digestion processes; validated biotechnological applications of SLMs for AD bioaugmentation; life cycle assessment and a
stakeholder roadmap for technology implementation.
Publications
El Houari A
(2023)
Lutispora saccharofermentans sp. nov., a mesophilic, non-spore-forming bacterium isolated from a lab-scale methanogenic landfill bioreactor digesting anaerobic sludge, and emendation of the genus Lutispora to include species which are non-spore-forming and mesophilic.
in International journal of systematic and evolutionary microbiology
Hornung B
(2023)
An objective criterion to evaluate sequence-similarity networks helps in dividing the protein family sequence space
in PLOS Computational Biology
| Description | 1. In this project we have identified several new species of bacteria associated with waste decomposition and biogas generation and have a collection of more than 60 pure culture isolates and low diversity enruchment cultures comprising biomass-converting microorganisms. 2. We have also generated large DNA sequence datasets to describe the microorganisms in landfill sites that can be used to decompose different waste types in landfill sites and anaerobic digester plants. 3. We have conducted bioreactor experiments to assess teh efficiency of waste biomass conversion to biogas using natural microbial communities form landfills and anaerobic degestion plants, but also combinations of isoalted bacteria and enrichments comtaining Archaea. WP1. Characterising landfill microbiomes. 1.1 Construction of landfill microbiome bioreactors (COMPLETED); 18 lab-scale bioreactors were established, comprising six treatments in triplicate; (1) AD sludge, (2) maize, (3) food waste, (4) chicken manure, (5) municipal solid waste and (6) a control (landfill leachate without feedstock). Bioreactors were incubated for 28 days and sampled at days 0, 7, 14, 21, and 28 for physicochemical, DNA-based microbiome analysis (16S rRNA gene and shotgun metagenomics), biogas production and microbial isolation. 1.2 Multi-omic sequencing of biomass-degrading microbiomes (COMPLETED). DNA from 90 bioreactor samples underwent 16S rRNA gene Illumina MiSeq sequencing, and Shotgun Metagenomic sequencing (Illumina NovaSeq 6000) and have been delivered to the partners for enzyme discovery (WP2) and for network modelling (WP3). 1.3 Isolation of landfill biomass-converting microorganisms (ONGOING). anaerobic biomass-converting microorganisms were isolated from bioreactors using High throughput isolation in microplates or Hungate roll-tubes. Over 500 strains of landfill microorganisms have been isolated, belonging to 60 different species. Fifteen of these isolates are novel and represent members of new species, genera, and families. In addition, low diversity enrichments of cellulose-degrading communities (Fibrobacters and Spirochaetes) and methanogens have been generated for use in WP4. 1.4 Genomics of landfill biomass-degrading isolates (ONGOING). Whole genome sequencing is complete for three of the new species isolates (named m5, m25 and meth-B3). They are also deposited in two culture collections (DSMZ and ATTC) and will soon be formally taxonomically described in upcoming publications. WP2. Computational enzyme discovery and lab characterisation. Task 2.1 Computational enzyme discovery (COMPLETED). The metagenome datasets have been annotated through the CAZy pipeline to identify carbohydrate-active enzymes. Task 2.2 Bacteroidetes polysaccharide utilization loci (PULs) (COMPLETED). Based on the presence of SusD genes, we predicted PULs that are bacterial operons gathering complementary enzymes for the deconstruction of target substrates. Both gene and operon annotations for tasks 2.1 and 2.2 will soon be assigned to each time point and treatment from WP1 to allow specific interpretation of key enzymes involved in AD. Enriched enzyme families and those with similar substrate specificity will be selected for biochemical characterization in task 2.3. Hence, to benefit from the most precise knowledge and predictive power, a literature review has been performed and a method for the creation of subfamilies has been developed in Task 2.1. A manuscript presenting an objective criterion to select optimal thresholds in sequence similarity networks has been written and will soon be submitted. This work package will provide subfamilies for 3 glycoside hydrolase families, while only 5 families have benefited from such detailed analysis in the past 15 years. Task 2.3 'Enzyme screening and biochemical characterization' will commence in the next few weeks. WP3. Network modelling and synthetic microbiome design. Task 3.1 Community-wide metabolic network modelling (ONGOING). Computer code was developed to model community-wide metabolic networks and to identify choke-points and levels of redundancy for desired conversion pathways. However, after methodological considerations, this task was rescheduled to be completed after task 3.2. Task 3.2 Reconstruction of individual metabolic network models (ONGOING). A pipeline for the reconstruction of metabolic models using CarveMe was developed. CarveMe uses ORFs from MAGs as input, and our pipeline modified the tool to receive the taxonomy of the modeled organism, allowing to rank reactions annotated by KEGG and BioCyc as more likely to be present in the model. To produce high-quality models, we created a database that unifies metabolic reactions from KEGG, BioCyc, and BIGG. Additionally, we developed a web application for the curation and comparison of metabolic models. The tool identifies essential reactions and maps them to KEGG and BIGG pathways, and facilitates the automatic inclusion and deletion of reactions according to the pathway in which they are present. The tool is available online. Task 3.3 Dynamic community simulations (ONGOING). Scripts for identifying metabolite exchange between organisms were developed and will be used for the community simulations. |
| Exploitation Route | 1. The identification of bacteria and archaea that are potentially new to science enables further analysis of their biology and further work will facilitate a greater understanding of the processes associated with biogas production. 2. Enhanced understanding of the composition and activity of microorganisms that degrdade waste biomass has applicatiosn in future research focussed on enhancing biogas production from waste. |
| Sectors | Agriculture Food and Drink Energy Environment Manufacturing including Industrial Biotechology |
| URL | http://www.synbiogas.com |
| Description | A pipeline for high-throughput microbial isolation, sorting, screening and synthetic community assembly |
| Amount | £932,339 (GBP) |
| Funding ID | BB/X01942X/1 |
| Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 07/2023 |
| End | 07/2024 |
| Description | FuturEnzyme: Enzymes for more environment-friendly consumer products |
| Amount | € 6,000,000 (EUR) |
| Funding ID | 101000327 |
| Organisation | European Commission H2020 |
| Sector | Public |
| Country | Belgium |
| Start | 05/2021 |
| End | 03/2025 |
| Title | Analysis of the whole genome of of the SLM isolate |
| Description | Raw reads generated by MiSeq for each isolate were were adapter trimmed using Trimmomatic 0.30 with a sliding window quality cutoff of Q15 (Bolger et al., 2014). De novo assembly was then performed using SPAdes version 3.12 (Bankevich et al., 2012), and finally the contigs were annotated using Prokka 1.14 (Seemann, 2014). Scaffolds below to 1500 bp were filtered and the final assembled genome for each each isolate is generated to be used for downstream analysis of annotation and characterization. |
| Type Of Material | Physiological assessment or outcome measure |
| Year Produced | 2021 |
| Provided To Others? | Yes |
| Impact | These MAGS serve to the French and German partners for CAZymes characterization and for metabolic network modelling. |
| Title | Bioreactors design |
| Description | In order to design synthetic landfill microbiomes, lab-scale bioreactors for lignocellulose biomass conversion using 1 litter DURAN bioreactors were set up. The goal is to simulate what's happen in landfill sites in order to design and validate synthetic landfill microbiomes for enhanced biogas production. Five feedstocks rich in cellulose (Municipal solid waste, Maize, Chicken manure/paper/cow slurry mixture, Food waste and AD sludge), were tested as energy source for the microbial communities obtained from Hafod landfill site and drilled waste obtained from Rouaben site. The feedstocks were frozen dried to eliminate liquid phase and keep only the solid material. Then 2% of total solid of each feedstock was added anaerobically to the bioreactors under N2-free oxygen gas, then he bioreactors were sterilized by autoclaving and inoculated anaerobically using Leachate/drilled waste mixture. Finally, Bioreactors were incubated for 28 days under agitation at 150 rpm and at 37 °C. The microbial growth in bioreactors was followed during incubation and a sampling of both gas and liquid phases each 7 days was performed from each bioreactor for physiochemical (pH, fibres analysis, biodegradation products), molecular (DNA based approaches) and cultural (microorganisms isolation) downstream analyses. |
| Type Of Material | Biological samples |
| Year Produced | 2020 |
| Provided To Others? | No |
| Impact | The samples taken out from these bioreactors set up using this tool will be used for the purpose of the characterisation of microbial communities originated and involved in lignocellulose biomass conversion in Landfill sites. In addition, this tools allows to simulate what's happen in landfill sites in order to characterize, design and validate synthetic landfill microbiomes for enhanced biogas production. |
| URL | https://www.duran-bottle-system.com/files/Downloads/order_info_caps_closure/DURAN_GLS80_BottleCaps-C... |
| Title | Microorganisms Isolation Work |
| Description | The targeted microorganisms and groups of interest were first isolated/enriched using specific medium, and by using the key substrates known to be used by them, respectively. For example, cellulose degrading microorganisms (Fibrobacters and Spirochaete) were enriched on Avicel (a microcrystalline cellulose) as unique energy source. However, methanogens were enriched in the presence of the six key substrates known to be used by Archaea (H2, acetate, methanol, methylamine, formate and CO2). Once, enriched, the microorganisms were isolated using High-throughput isolation in microplates or Hungate tubes techniques. For isolation using microplates, to ensure anoxic conditions and avoid oxygen, the entire isolation procedure was carried out in an anaerobic and the microplates were sealed as soon as possible and then incubated inside the anaerobic chamber. However, for isolation using Hungate tubes were performed using a solid anaerobic medium previously dispatched in Hungate tubes. The entire procedure of sampling and inoculations were performed using sterile syringes and needles under a flow of free-oxygen nitrogen. Once isolated, strains will be identified by amplification and sequencing of the 16S rRNA gene using Sanger sequencing.. |
| Type Of Material | Physiological assessment or outcome measure |
| Year Produced | 2013 |
| Provided To Others? | No |
| Impact | The isolates of interest will be used to construct the synthetic landfill microbiome. This microbiome will be challenged in order demonstrate its robustness and efficiency through its application for bioaugmentation in both lab scale landfill designs, as well as global scale in landfill and anaerobic digestion sites. |
| Title | Whole genome sequencing of the landfill isolates |
| Description | For the whole genome sequencing, the extracted DNA of each isolate, extracted using DNeasy UltraClean Microbial Kit (Qiagen) according to the supplier's recommendations, was eluted in nuclease-free EB buffer (10 mM Tris-HCl pH 8.0). The integrity of the genomic DNA was performed in 1% agarose gel with a 1 Kb molecular weight ladder. DNA quantification was carried out using dsDNA Quantification Kits following the manufacture recommendations (Invitrogen™ Qubit™ 3 Fluorometer - Fisher Scientific) and diluted to a final concentration of 10 ng/µL. DNA Libraries were sequenced using Illumina sequencing platform (NovaSeq) with 2x250 bp paired-end reads using MicrobesNG facilities (Birmingham Research Park, UK). |
| Type Of Material | Physiological assessment or outcome measure |
| Year Produced | 2020 |
| Provided To Others? | Yes |
| Impact | These genomes were analyses and used to build the metabolic pathways of the Synthetic Landfill Microbiome. |
| Title | Description and characterization of novel species |
| Description | Three strains (designed m25, m5, and meth-B3) among the 17 novel species isolated from the SLM bioreactors were deeply described and characterized in term of ecophysiology, and taxogenomics. These three strains are subject of three publications to be submitted to the International Journal of Systematic and Evolutionary Microbiology (IJSEM) for novel species description isolated for the first time from the landfill microbiota. |
| Type Of Material | Data handling & control |
| Year Produced | 2021 |
| Provided To Others? | No |
| Impact | The description of these strains will be a source of information to fill in the lack of knowledge missed in term of the diversity of the biomass converting microorganisms in landfill ecosystems. |
| Title | Extraction of Genomic DNA and High Throughput Sequencing |
| Description | Total DNA extractions were performed in triplicate using CTAB/phenol Cloroform method following Griffiths instructions for DNA-RNA coextraction (Griffiths et al., 2000). The DNA was stored at -80 °C until further use. The V3-V4 hypervariable region of the 16S rRNA gene targeting Bacteria and Archaea was amplified using the primers set 341F and 805R. Forward and reverse primers contained the adapter sequences (5'GTGYCAGCMGCCGCGGTA) and (3'ACTYAAAKGAATTGRCGGGG), respectively, used in Illumina sequencing technology. The amplicons were sequenced using an Illumina MiSeq (300 bp paired-end reads) at Centre for Environmental Biotechnology (CEB) of Bangor University. |
| Type Of Material | Data handling & control |
| Year Produced | 2021 |
| Provided To Others? | No |
| Impact | Raw datasets of Miseq sequencing was analysed using QIIME2 pipeline and was used to characterize the single gene communities profiling coming from Bioreactors design. |
| Title | Metagenome sequencing and analysis of Landfill bioreactors |
| Description | Total DNA extracted from the the bioreactors design was utilized to generate 32 sequencing libraries in Total. The genomic DNA was randomly fragmented by sonication, then DNA fragments were end polished, A-tailed, and ligated with the full-length adapters of Illumina sequencing, and followed by further PCR amplification with P5 and indexed P7 oligos. The PCR products as the final construction of the libraries were purified with AMPure XP system. Then libraries were checked for size distribution by Agilent 2100 Bioanalyzer (Agilent Technologies, CA, USA), and quantified by real-time PCR (to meet the criteria of 3 nM). The qualified libraries are fed into Illumina sequencers after pooling according to its effective concentration and expected data volume. The 32 DNA extracts were sequenced using Shotgun metagenomics service (NovaSeq 6000 PE150) generating 40 million paired reads sequencing per sample) at Novogene platform (Cambridge, United Kingdom). 12 G of raw data per sample were generated. Adapter sequences were removed using Cutadapt (version 1.2.1) (45) and trimmed via Sickle (version 1.2) (46) with a minimum window quality score of 20 and reads shorter than 10 bp removed. The three sequence libraries were combined and assembled via Ray Meta (31) (version 2.3.1, k-mer = 31) using the HPC Wales computing network. Raw reads and assembled contigs were uploaded as separate data sets to One Codex and classified against the One Codex database (32). The raw reads obtained were assembled using megahit software and analysed using using the HPC Wales computing network. The assembled metagenome contigs were subjected to taxonomic assignment. Gene prediction was performed on both the whole-metagenome data set via Prodigal v2.6 and annotated via the dbCAN. |
| Type Of Material | Data analysis technique |
| Year Produced | 2021 |
| Provided To Others? | No |
| Impact | The assembled metagenome contigs will be used in WP2 and WP3 of SYNBIOGAS project by our French and German partners for Computational enzymes discovery and network modelling in order to validate the optimal synthetic microbiomes for anaerobic digestion to enhance biogas production. |
| Title | Sanger sequencing for identification of isolates |
| Description | All genomic DNA extractions from isolated strains cultures were carried out using a kit (UltraCleanTM Soil DNA isolation kit, MoBio Laboratories) according to the supplier's recommendations. The enrichments were incubated at 37 ° C for 1 week; 1.8 ml of culture is centrifuged, The bacterial pellet is used for extraction. The gel electrophoresis technique is used to visualize nucleic acids. The DNAs are mixed with a buffer solution (TBA 1 X) applied to a 1% agarose gel; then molecules pushed back on one side by the electric current produced by one electrode are simultaneously attracted by the current produced by the other electrode. A marker of known size making it possible to determine the size of the DNAs more easily is used. The DNA samples to be loaded on the agarose gel are first mixed with a loading buffer which increases the density of the sample. At the end of the migration, the DNA fragments are easily detected on the gel using a fluorescent dye, safeview. Amplifications of the full gene encoding 16S rRNA was carried out using with the primers (0.8 µM each) 27F (5'-TGAGCCATGATCAAACTCT-3') /1487R (5'-GGWTACCTTGTTACG-3') and MyTaq™ Red Mix (Bioline). The PCR products are purified using the commercial QIAquick PCR Purification Kit (QIAGEN). The purpose of this purification is to separate the amplified DNA fragment from the rest of the components of the reaction medium (proteins, salts) and in particular from other nucleic acids (unused primers, amplification of unwanted sequences less than 100 base pairs, etc. ). The size and quantity of amplicons were checked by electrophoresis on a 1% agarose gel and then Sanger sequencing of the amplicons obtained is carried out by the Genwiz platform (https://www.genewiz.com/en-GB/Public/Services/Sanger-Sequencing/). |
| Type Of Material | Data analysis technique |
| Year Produced | 2021 |
| Provided To Others? | No |
| Impact | The isolation work/sanger identification allowed to isolate more than 600 isolates. Among them, 7 new strains were isolated for the for the first time. These new strain represent a novel species, genera/family and will be subject of characterisation publication. In addition, the the resulting datasets were used for the identification of isolates. The isolates of interest will be challenged in order demonstrate its robustness and efficiency through its application for bioaugmentation in both lab scale landfill designs, as well as global scale in landfill and anaerobic digestion sites for biogas production. |
| Title | Single Gene Community Profiling Analysis |
| Description | The datasets from MiSeq sequencing were processed following the QIIME2 pipeline as described by Bolyen et al., (2019). Amplicon Sequence Variants (ASVs) were grouped at an identity threshold of 97%. From the taxonomic affiliation, a representative sequence of each ASV was extracted. Finally, a normalization of the abundances found for each ASV was carried out by the DESeq method. Diversity and evenness indices were calculated using the bacterial ASVs data after normalization. The evolutionary history was inferred using the neighbor joining method. An alignment was then carried out between ASVs; the alignment was used to build a phylogenetic tree with FastTree algorithm. The statistical calculations and multivariate analyses were conducted using R software (https://cran.r-project.org/) with the following libraries: phyloseq, vegan, ape, Venn diagram, heatmap2, gclus, ggplot2, GUniFrac, grid, and optparse. A one-way analysis of variance (ANOVA) was used to compare the mean values with respect to sampling sites. A principal component analysis (PCA) was performed to evaluate the microbial community composition of the samples. |
| Type Of Material | Data analysis technique |
| Year Produced | 2021 |
| Provided To Others? | No |
| Impact | This analysis, using metabarcoding approaches, allowed to investigate the diversity profiling of the microbial communities in each bioreactors treatment with a focus on a specific groups of interest such cellulose degrading microorganisms and Archaea. |
| Description | AIX University Marseille |
| Organisation | Aix-Marseille University |
| Country | France |
| Sector | Academic/University |
| PI Contribution | Project partner working on work package 2 of the project |
| Collaborator Contribution | In silico enzyme discovery |
| Impact | In progress |
| Start Year | 2020 |
| Description | Infinis |
| Organisation | Infinis Limited |
| Country | United Kingdom |
| Sector | Private |
| PI Contribution | Research expertise and analysis of samples |
| Collaborator Contribution | Provision of landfill leachate and drilled waste samples for experimental work |
| Impact | In progress |
| Start Year | 2020 |
| Description | Material change - Angelsey energy |
| Organisation | Material Change |
| Country | United Kingdom |
| Sector | Private |
| PI Contribution | Provision of expertise and research relating to operation of the companies AD plants |
| Collaborator Contribution | Provision of expertise and anaerobic digester feedstocks and samples |
| Impact | In progress - tested AD feedstocks for biogas generation |
| Start Year | 2020 |
| Description | Natural Resources Wales |
| Organisation | Natural Resources Wales |
| Country | United Kingdom |
| Sector | Public |
| PI Contribution | Supporting industry by addressing key waste management questions. Providing data from samples analysed. |
| Collaborator Contribution | Introduction to industry waste management partners. Intellectual input into experimental and research design. Provision of samples and materials of rthe project. |
| Impact | New collaborations and partnerships; new research ideas; reseach co-design. |
| Start Year | 2020 |
| Description | UFZ Helmholtz |
| Organisation | Helmholtz Association of German Research Centres |
| Department | Helmholtz Centre for Environmental Research - UFZ |
| Country | Germany |
| Sector | Academic/University |
| PI Contribution | Project leaders - provide data and expertise |
| Collaborator Contribution | Project partner working on work package 3 and 4 of the project |
| Impact | In progress |
| Start Year | 2020 |
| Title | SBMLComp: a web application for curation and comparison of constraint-based metabolic models. |
| Description | a web application for curation and comparison of constraint-based metabolic models. |
| Type Of Technology | Webtool/Application |
| Year Produced | 2021 |
| Open Source License? | Yes |
| Impact | Manuscript currently in preparation |
| Description | Bangor University undergraduate research seminar - February 2021 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | National |
| Primary Audience | Undergraduate students |
| Results and Impact | Research seminar as part of 3rd year benvironmental microiology and biotechnology module at Bangor University |
| Year(s) Of Engagement Activity | 2021 |
| Description | Bangor University undergraduate research seminar - February 2022 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Undergraduate students |
| Results and Impact | Lecture discussing research funded by this project |
| Year(s) Of Engagement Activity | 2022 |
| Description | ERA CoBioTech Hack |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | The third BioTech Research & Innovation Hack serves as a Joint Status Seminar of the research and innovation projects funded under ERA CoBioTech. In 12 online session the research consortia will present their current process, working: to bring new insights about the potential of microbial communities, co-cultures and cascades of microorganisms for new products or biological processes, to synthesise new or to improve existing cell factories (bacteria, archaea, yeasts, fungi, algae, etc.) to isolate new and to improve the performance of existing enzymes (biocatalysts) and enzyme cascades etc. to optimise microbial communities for better performance, to upscale industrial processes and to increase their efficiency and economic feasibility, and to discover new, efficient and sustainable methods for manufacturing bioproducts. |
| Year(s) Of Engagement Activity | 2021 |
| URL | https://www.cobiotech.eu/events?backRef=10&event=BioTech_R_I_Hack_2021 |
| Description | ERA CoBioTech: Invitation to the BioTech Research & Innovation Hack 2020 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | Presented a status seminar on the SYNBIOGAS project for the ERA CoBioTech community at the BioTech Research & Innovation Hack 2020. |
| Year(s) Of Engagement Activity | 2020 |
| Description | ERA Policy Brief Workshop |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | We would like to involve all projects funded under the ERA-CoBioTech umbrella towards the drafting of the Policy Brief. Although our experts have already extracted key information from your reports and (published) results, we would very much appreciate to learn about your thoughts and provide the opportunity to have your say to "underline the importance of your research and its impact" in order to include it in the Policy Brief, where applicable. |
| Year(s) Of Engagement Activity | 2022 |
| Description | Flash talk and poster presentation at ISME18 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | Selected for a flash presentation and poster at the International Society for Microbiology Ecology (ISME) international meeting in Lausanne, Sweden in August 2022. |
| Year(s) Of Engagement Activity | 2022 |
| Description | Invited guest seminar at University of Birmingham |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Other audiences |
| Results and Impact | Invited research seminar focussing on research associated with enviro mental and host-associated microbiomes |
| Year(s) Of Engagement Activity | 2022 |