Team:Exeter/HP/Bioreactor
Below, you will find a timeline of design steps that were taken in order to reach our final bioreactor.
1: Initial Designs
The iGEM Leiden 2016 team’s project was similar to ours, and so we took inspiration from their work. Our first bioreactor design was along the same lines as Leiden 2016’s. It was very simple design, shown above, with mobilised bacteria being mixed with perchlorate in a single chamber. However, it was only a starting point.
Specialists in DPRB (disperate perchlorate reducing bacteria), John D Coates and Ouwei Wang had drawn up loose designs for bioreactors using their research, shown above. In their design, hydrogen atoms would pass from an anode to a cathode through a cation selective membrane, aiding the reaction in the diagram below. This would speed up the conversion of perchlorate to chlorite, a rate limiting step. The design also switched to immobilised bacteria which reduces the chances of E. coli escaping.
From them we got the idea of using a cathode system to aid perchlorate reduction. After contacting them, we also gained insight into the different parts we needed to insert into our chassis, and the existing parts of E. coli that could be harnessed. They had been working on disparate perchlorate reducing bacteria, and offered a lot of input on how to apply their research to synthetic biology, bringing up the idea of bioreactors for bioremediation on Earth and on Mars.
Dr Clive Butler helped us to understand the needs and properties of bacteria that reduce compounds. From him we realised that we would have to supply haemin, molybdenum, and iron sulphate to our E. coli as the enzymes that reduce perchlorate are metalloenzymes.
2: Earth versus Mars
Our initial research into bioreactors suggested that there is a need for a smaller, more efficient bioreactor in comparison to the large fluid and solid bed bioreactors currently reducing perchlorate on Earth. However, Dr Ceri Lewis, an ecotoxicologist, informed us that perchlorate in water sources is not a prominent environmental issue. More information about the spread and severity of perchlorate needs to be known before it can be determined whether this is a sufficiently useful project, and this information wouldn’t be available within the timescale of the project.
Professor Mike Allen of Plymouth Marine Laboratory, a biochemist with experience in bioreactor design, advised us that if there was a real effort to remediate perchlorate contamination, it would occur on site and not at water sources. This is due to the soluble nature of perchlorate, meaning it would spread through water sources very quickly. In order to bioremediate it, you would have to run the entire water source through the bioreactor, which is infeasible. He told us that he didn’t see much demand for a novel perchlorate reducing bioreactor on Earth.
3: Deciding on A Martian Bioreactor
We contacted iGEM teams in areas where perchlorate contamination seemed to be problem. Many of the teams had never heard of perchlorate and were unaware it was a problem, indicating that it is not a major problem, even to those with higher than normal levels. We hoped that they could test their tap water themselves, as our attempts to get in contact with American cities’ water and power departments ended due to bureaucratic restriction. We managed to collaborate with an American team in Virginia, working with them on a perchlorate assay. We did not find significant level of perchlorate in their water.
Looking closer to home, we also got in touch with South West Water and spoke to one of their Water Quality Specialists, Tim Coates, asking for a water quality report for our local area of Devon. They informed us that perchlorate is not a pollutant they are concerned about, and that they do not test for perchlorate in their water.
It was concluded that perchlorate was not such a large issue as previously thought, and there was little to no demand for a bioreactor that could filter it out of existing water systems. Thanks to all of this stakeholder input, we decided to focus on a Martian bioreactor rather than a Earth one.
4: Swirl Flow Bioreactor
After we had decided that our bioreactor would be designed for Martian use, we spoke again with Professor Mike Allen. He introduced us to the concept of Swirl Flow Bioreactors (SFB), which would allow us to mix the perchlorate with the E. coli and separate the oxygen, all in one chamber. We took the design of a typical swirl flow bioreactor and adapted it to fit our needs of having immobilized E. coli and a cathode, shown above.
However, as Professor Allen pointed out, having an electric current in the bioreactor would provide opportunity for unexpected and unwanted reactions, while not contributing significantly to the rate of reaction. Therefore it was decided that the design should be simplified and the cathode removed. He introduced the concept of alginate beads for containing the E. coli to us. Plymouth Marine Laboratory were kind enough to lend us one of their Swirl Flow bioreactors, which we used to test whether the beads would be suitable.
Our first SFB without cathodes involved four chambers, one to mix the regolith with water to extract the perchlorate, and three to consecutively reduce the perchlorate into oxygen until none is left. It is shown above. However, this had high energy requirements due to four impellers being powered at once, and could be simplified further.
5: Mars Design
We contacted Nick Musgrove, commercial director of Infors HT to help us understand the basics of bioreactor design. Using the information gathered from this talk we drafted the features that we envisaged our bioreactor having to optimise it for perchlorate reduction.
In order to develop our idea further, we needed to ascertain how regolith would be entered into our bioreactor. Dr Ben Reeve, CTO of CustoMem, a company specialising in biomembranes, discussed with us whether a biomembrane would be an effective measure for getting perchlorate into our reactor. As a result of this talk it was decided that a biomembrane would be an unnecessary complication to our bioreactor and could not achieve anything that a simple sieve-like filter couldn’t.
Understanding how our bioreactor would be used and who would operate it was a key consideration in our design process and Libby Jackson, a flight coordinator the UK Space Agency, illuminated this for us. We learned how bacteria are stored and experimented with in spacecraft, the priorities that astronauts have when experimenting, and the legal precedent of taking modified E. coli into space. She raised valuable questions of biosecurity that we had to answer.
6: Tailoring the Bioreactor For Mars
With a solid base, we began to tweak the design based on the needs of a bioreactor on Mars, with the help of our stakeholders. We determined the most important aspects of a Martian bioreactor:
- 3D printable and light. This is due to limitations on what we could take to the planet. Michael Curtis-Rowse informed us that our procedures on Mars could be automated and that no human interaction with the bioreactor should be required. He helped determine the kind of materials we would have to make the bioreactor out of (printable plastic, Ultem, shown above).
- Resource efficient. Power consumption has to be low, in order not to drain the resources of the rest of the biodome. By reducing the bioreactor to one chamber and one impeller, it is possible to drastically reduce the energy requirements, since that is the main use of power.
- Shielded from radiation. Without protection, our bacteria will die on the Martian surface. The shielding also needs to be light enough to take to Mars. High Z Steel-Steel Composite Metal Foam (HZ S-S CMF) can block UV rays and neutrinos as well as pure lead (Shuo Chen et. al., 2015), while being much lighter and environmentally friendly.
- Safe. There must be a way to kill the bacteria in the bioreactor in the case of unwanted mutations or other unforeseen events. Fortunately, in the event of escape, the UV on the surface would kill any bacteria which made it out of the bioreactor. In the case of unwanted mutation, running copper-alginate beads through the bioreactor has been shown to kill E. coli (Simon F. Thomas et. al., 2014).
We also thought briefly about where we could place the bioreactor on Mars. Alex Price informed us of which areas are protected, and which would be viable landing spots. Professor Charles Cockell introduced us to the idea of the planetary park system, which could protect certain areas from contamination while allowing others to become potentially habitable.
7: Integrating into Existing Infrastructure
8: A Future for the Bioreactor
After talking closely with stakeholders throughout the entire design process, we settled on a final design. The design features a single impeller chamber in which the reaction will take place. On the Martian surface, a rover would shovel regolith into the bioreactor with water. Then a filter would remove the solids rocks and soil, leaving only a perchlorate solution. This solution would then flow into the reaction chamber with alginate beads containing E.coli, where the perchlorate will react with the bacteria. The filtered soil will be deposited into the baffle system by the vortex, since it will be the densest material, and then removed. Oxygen produced by the reaction will be the least dense, and so the vortex will concentrate it in the centre of the chamber, to be carried out via a manifold. The remaining water will be recycled back into the chamber for the next reaction. It is a simple but efficient design.
If we had more time, it would be valuable to build and test a prototype, however, we were lucky enough to experiment with an existing, similar bioreactor which gave us an idea of how our design would work. In the future, we would like to keep communicating with our stakeholders and develop our design into a fully functioning bioreactor, producing oxygen on Mars.
Stakeholders
Professor John D Coates
Professor of Microbiology, Chair Plant and Microbial Biology, and Academic Director, Energy Biosciences Institute. He has done research in Environmental microbiology encompassing the fields of bioremediation, alternative energy production, and biogeochemistry. The Coates Lab focuses on environmental microbiology: applied microbiology and bioremediation. They investigate removal of radioactive toxic metals, carcinogenic petroleum-based hydrocarbon contaminants, and toxic munitions byproducts from the environment. Recently, they identified dominant groups of bacteria that can transform perchlorate wastes into innocuous chloride, isolated and characterized more than 40 such bacteria, and identified the common biochemical pathway and genetic systems involved.
http://plantandmicrobiology.berkeley.edu/profile/coates
Ouwei Wang
Ouwei Wang is a Graduate Student at the Coates Lab at the University of California. The Coates Lab focuses on environmental microbiology: applied microbiology and bioremediation. They investigate removal of radioactive toxic metals, carcinogenic petroleum-based hydrocarbon contaminants, and toxic munitions byproducts from the environment. Recently, they identified dominant groups of bacteria that can transform perchlorate wastes into innocuous chloride, isolated and characterized more than 40 such bacteria, and identified the common biochemical pathway and genetic systems involved.
http://plantandmicrobiology.berkeley.edu/profile/owang
Dr Clive Butler
Associate Professor of Microbial Biochemistry. His research interests lie in the study of the microbiology and biochemistry of important mineral cycles, with specific focus on the microbial interaction with selenium and nitrogen. The main objective of his work is to characterize the biochemical reactions of these cycles in whole cells and cell fractions. He studies gene expression and protein synthesis in response to environmental change; investigating electron-transfer reactions between proteins involved in respiratory pathways; and purifying and characterizing proteins and enzymes to reveal their structure and function.
https://biosciences.exeter.ac.uk/staff/index.php?web_id=clive_butler
Dr Ceri Lewis
Senior Lecturer in Marine Biology. She is a marine biologist interested in understanding how marine invertebrates adapt and survive in a changing and increasingly polluted marine environment. She’s currently working on a number of ocean acidification and microplastics projects and has a particular interest in the reproductive ecology of marine invertebrates. She is also a member of the Environmental Biology research group. In addition to her research she’s very active in public and educational outreach, teaming up with an educational charity to get her research findings fed into UK and international schools and working to increase public understanding of our oceans.
https://biosciences.exeter.ac.uk/staff/index.php?web_id=ceri_lewis
Professor Mike Allen
https://www.pml.ac.uk/People/Science_Staff/Professor_Mike_Allen
University of Virginia iGEM Team
Their project, Quorus, explores quorum sensing and how it can be applied to biomanufacturing. Heterogeneity of cell populations caused by quorum sensing leads to variability in gene expression that is hard to predict. During biomanufacturing, elevating quorum-induced protein expression will lead to gain of profit. Decreasing this expression can also be beneficial in situations where undesirable biofilms may form on medical equipment or controlling virulence in bacteria. Their team will modify the existing bacterial quorum sensing system controlled by the Lsr operon by upregulating the synthesis and excretion of Autoinducer-2 , a universal quorum molecule. This will increase population-scale AI-2 intake and phosphorylation after the initial AI-2 threshold concentrations have been reached to reduce variability in induced gene expression.
https://2018.igem.org/Team:Virginia
South West Water Company
South West Water is the water and wastewater service provider for a population of c. 1.7 million in Cornwall, Devon, and parts of Somerset and Dorset. Since 2016 it has also been providing water services in the Bournemouth Water region to a population of c. 0.5 million. They provide reliable, efficient and high quality drinking water and waste water services throughout these areas. South West Water is the only water supplier in the region, therefore there is a very strict system of regulation in place to safeguard the best interests of its customers and the environment.
https://www.southwestwater.co.uk/
Nick Musgrove
Nick Musgrove Commercial Director Nick has a degree in Pharmacology obtained whilst working as a research scientist at Beecham Pharmaceuticals. He has 28 years sales experience and has been in his current position for almost 19 years. He has helped develop a viable and sustainable business and his philosophy is to always put the customer first; offering leading world class products with relevant scientific knowledge and excellent back-up and support.
http://www.infors-ht.co.uk/en/ihr-team-in-deutschland/
Dr Ben Reeve
Bioengineer and entrepreneur, making bio-based materials to help tackle the most urgent problems in water treatment. Co-founding scientist and chief technology officer at CustoMem Ltd. Leads technology development. He completed his PhD at Imperial College London in 2016 where his research focused on toolkit development and biomaterial production in non-model organisms. Ben did his MA at Cambridge university in biological sciences.
https://www.linkedin.com/in/ben-reeve/
Libby Jackson
Libby Jackson is currently the Human Spaceflight and Microgravity Programme Manager for the UK Space Agency, so she is responsible for the UK's Human Spaceflight and Microgravity programmes on the International Space Station (ISS). She used to be a flight director on space missions, ensuring that everyone worked together and everything went according to plan. Flight directors talk to everyone involved, from the astronauts to the scientists on Earth to the people who are responsible for making the rockets work. A flight director's most important tasks are to keep the astronauts safe, to check that all the planned experiments take place in space, and, of course, to sort out any problems that crop up.
http://www.destinationspace.uk/meet-space-crew/libby-jackson/
Alex Price
Currently working on a PhD in Microbiology, Open University, UK, Project: 'Biogeochemistry in the deep sub-surface environment: the key for finding potential life on Mars'. He is working with the OU’s Astrobiology research group and has taken data from various Mars missions to reconstruct aspects of the early Mars environment, when it was warmer and wetter, so that he can assess the ability of certain microbes to have thrived there.
http://www.open.ac.uk/people/abp87#tab1
Michael Curtis-Rowse
Lead Manufacturing Technologist at Satellite Applications Catapult. An innovative and versatile engineer with substantial experience in the exploitation of advanced materials and additive manufacturing across the technology readiness spectrum including applications ranging from spacecraft to medical devices.
https://uk.linkedin.com/in/mike-curtis-rouse-0a82629
Professor Charles Cockell
Professor of Astrobiology. His research group is interested in Astrobiology. As a discipline, it seeks to understand the origin, evolution and distribution of life in the Universe. Their particular research focus lies in the study of life in extreme environments and understanding the the diversity, processes and biosignatures of life in extremes and the potential habitability of extraterrestrial environments. Their work is conducted within the UK Centre for Astrobiology, a virtual astrobiology centre they established in 2011 that is affiliated with the NASA Astrobiology Institute.
https://www.ph.ed.ac.uk/people/charles-cockell
Melanie Pickett
Graduate Researcher at NASA - National Aeronautics and Space Administration, currently working to address the issue of water regeneration for long-duration space missions. She is also in pursuit of her Ph.D in Environmental Engineering at the University of South Florida. She is most interested in sustainable water, wastewater, and solid waste treatment practices coupled with potential biofuel generation opportunities. Her current doctoral research is focused on transitioning a proof-of-concept, lab-scale algal membrane photobioreactor technology to a functional prototype for the sustainable production of algal bioproducts and water quality enhancement.
https://www.linkedin.com/in/melaniepickett1/
Professor Claude Gilles Dussap
Author of more than 150 international publications, his research focuses on the engineering of biosystems and understanding the coupling between the conduct of a process and the operation of living cells in a bioreactor. He is a member of several editorial committees of international journals and symposia. He has also worked on the ESA’s Micro-Ecological Life Support System Alternative programme (MELiSSA). Within the Polytech network, and particularly in Clermont-Ferrand, he works for the balanced and synergistic development of engineering courses at universities. His strong commitment to the principles of equal opportunities, for the recruitment and professional integration of students, and his concern for the international opening of training led him to take on many national responsibilities such as the presidency of the e3a contest at the Class level. Preparations for the Grandes Ecoles, or in the program IDEFI - AVOSTTI (Initiatives of Excellence in Innovative Training).
https://www.linkedin.com/in/claude-gilles-dussap-a3bba09b/
- Françoise Bichai, Benoit Barbeau (2006) Assessing the Disinfecting power of Chlorite in Drinking Water (DOI: 10.2166/wqrj.2006.041)
- Ouwei Wang, John D. Coates (2017) Biotechnological Applications of Microbial (Per)chlorate Reduction (DOI: 10.3390/microorganisms5040076)
- Shuo Chen, Mohamed Bourham, Afsane hRabiei (2015) Attenuation efficiency of X-ray and comparison to gamma ray and neutrons in composite metal foams (DOI: 10.1016/j.radphyschem.2015.07.003)
- Simon F. Thomas, Paul Rooks, Fabian Rudin, Sov Atkinson, Paul Goddard, Rachel Bransgrove, Paul T. Mason, Michael J. Allen (2014) The Bactericidal Effect of Dendritic Copper Microparticles, Contained in an Alginate Matrix, on Escherichia colis (DOI: 10.1371/journal.pone.0096225)