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Revision as of 00:06, 18 October 2018
Design
Engineering fungal mycelium to create viable building materials for Mars is not a small challenge. This project, which was inspired by conversations with Lynn J. Rothschild from NASA, and the Stanford-Brown-RISD iGEM team, whom we owe thanks both for the inspiration and subsequent sparring on ideas, has relied on several scientific fields to try to tackle the problems we have found along the way. Being inspired by the idea of simple and cost-effective construction on Mars, our design focuses on the fungal aspect of how a cyanobacteria/fungal production system can be used to create building materials. As we explored this idea, we were presented with many problems to be solved and even more questions to be answered.
The first and most pressing one, how does one cultivate fungus on Mars? Moreover, which fungal species should we use? Can we determine and model growth characteristics of our fungi? Is the strength of fungal materials determined by any distinct genes and is it possible to regulate these in a way that will make our materials even tougher? And not least, how do we make the system cost-effective? We set out to answer these questions to guide the creation of our final design.
Growing Fungus on Mars
Growing live fungus on Mars will be challenging as the environment on Mars is extremely hostile compared to Earth. In addition to the extreme cold, the atmosphere is very thin at less than 1% of the density of Earth’s atmosphere and consists of 96% carbon dioxide and virtually no oxygen (1). Moreover, our fungus needs organic compounds as a source of carbon and energy. Bringing substrates from Earth is out of the question due to the costs of transportation, thus in situ production of materials is vital. To solve the problem of available carbon sources, one can introduce cyanobacteria to the production system. Cyanobacteria can utilize the carbon dioxide in the Martian atmosphere, albeit, like the fungus, they, too, are unable to grow directly in the extreme conditions of Mars. To solve the problem of the Martian hostility it is necessary to have a production facility. Such a production facility for cyanobacteria has been theorized by researchers at NASA (2). Using covered thermally insulated ponds with compressed air and water from the Martian ice, it would be possible to grow cyanobacteria on Mars. Supporting this idea is the fact that all of the essential elements need for cyanobacterial and even plant growth can be found on Mars (2). Based on this system theorized by NASA, we suggest appending it with the capabilities of using the cyanobacterial biomass to produce fungal materials.
However, non-destructive ways of harvesting nutrients could possibly lead to a more effective process. So instead of using lysed-dead cyanobacteria there is also the possibility of using biomass produced by them as a secreted substrate. Cyanobacteria can provide heterotrophic organisms with both nitrogen and oxygen through their photosynthetic and nitrogen-fixing pathways. Specifically, they can convert carbon dioxide, the most abundant gas in Mars’ atmosphere, into oxygen and at the same time provide NH4+ to the feeding organisms by utilizing N fixation (2). These problems are beyond the scope of our project, but provides an interesting starting point for future projects.
Choice of Organism
The choice of organism used in our project was based on an inductive strategy. We had two variables needed to be fulfilled in our attempt to achieve the goals we had set. The first was to find a fungus species that, given the right substrate, would have the ability to form firm brick structures. The second was to use a safe microorganism with a sequenced genome and established genetic methods, thus enabling us to do genetic modifications. The project started with three candidate-organisms: Pleurotus ostreatus, Schizophyllum commune and Aspergillus oryzae. These organisms have previously been grown on a variety of substrates and some have been used for creating bricks. We were also recommended to use a Fusarium by researchers at Novozymes, but opted to focus on the ones we had multiple recommendations on. For various reasons, our choice ended up being A. oryzae; A. oryzae is comparatively easy to grow on various substrates, it grows fast, gives consistently good results and is a well-established model organism. Though the other fungal species have previously shown to have desirable characteristics, when it comes to creating fungal materials, the results of our initial experiments made us decide on doing our research in A. oryzae with the possibility of transferring the discoveries to other fungal species at a later point in the project.
How to Build Structures on Mars
In determining material properties of our fungal materials, experiments are critical. Depending on how many parameters you want to examine, the number of experiments tend to exponentially increase. Using the Fedorov exchange algorithms, we were able to reduce the number of experiments in the first tests from 216 to 64 and in the second run from 648 to 68, which was a huge time saver. It became clear to us that we needed to produce a lot of fungal sample cubes. Initially, we fabricated some acrylic boxes. These were fast and easy to make. It turns out that acryl does not withstand the pressures and temperature when the form gets autoclaved. We then turned to metal. These metal boxes took a long time to make, and after having fabricated 11, it was decided that the effort would not be worth it. After having done some research, silicone ice cube trays turned out to be the perfect alternative to our metal boxes. Silicone can easily withstand autoclaving and allowed for very uniformly shaped samples. They can also be purchased prefabricated cheaply. The sample cubes were all prepared in slightly different ways according to our experiment optimization such as baking times, compound composition, substrates etc. All the fungal samples were subjected to a destructive compressional test, where the stress/strain-curve was determined for compressional strength. In order to obtain the Young’s modulus of the fungus, the tensile strength would also have to be measured. If Young’s modulus is known, the entire materials’ properties and characteristics can be inferred.
Improving the fungal materials through synthetic biology
When using fungal mycelium and organic materials as the basis for building materials, a new venue of possibilities suddenly appear. Fungi are genetically modifiable. This allows for tuning of different external parameters or even adding properties such as biosensors etc. Thus, the interesting question to answer in this aspect is; what genetic modifications are beneficial when it comes to construction on Mars?
Like any other organism, any fungus is optimized for the survival of its species in its niche. No fungal strain is solely optimized for creating strong materials, as that wouldn’t be an advantageous evolutionary strategy. By making the fungus a little worse at surviving, we aim to make it better for a building. In our project, we ended up focusing primarily on two genes; melA and gfaA.
Protection from UV radiation and Melanin
One part of what makes the surface of Mars a hostile environment is its radiation. Unlike the terrestrial atmosphere, the Martian atmosphere does not shield well against cosmic radiation, making radiation a serious concern on the red planet (10). Part of the radiation issue is short wavelength UV radiation such as UVB and UVC, which is not just harmful for living tissue, but also causes radiation damage to solid materials (11). Nature has evolved systems that give the protection against UV radiation, most well-known is the pigment melanin. Overexpressing melanin might make fungal materials more resilient against radiation damage. Studies have shown that melanotic bacteria experience increased protection from the full spectrum of UV compared to counterparts without melanin (12). To slow down the potential erosion from UV radiation, we wanted to express the gene melA from Rhizobium etli CFN42 (13) in our fungal host. A previous iGEM team from the Technical Institute of Tokyo have shown that overexpression of melanin in E. coli can been achieved using the melA gene. melA encodes this for a tyrosinase that catalyses the enzymatic steps converting tyrosine into dopaquinone. We investigated the biosynthetic pathway of melanin in A. oryzae and discovered the same metabolic step as encoded for by melA (14). Heterologous expression of MelA in A. oryzae may thus also increase the production of melanin.
Melanin (Codon optimized for A. oryzae)
Cell wall structure and GfaA
Colors and selectable markers - AmilCP
amilCP gene. amilCP encodes a blue chromoprotein and exhibits strong blue color when expressed in E. coli, and we wanted to use amilCP constructs as a reporter system to test transformation success of different fungi.
Mission Architecture
(1)Williams M. 2017. What is the Weather like on Mars?.
(2)Verseux C, Baqué M, Lehto K, De Vera JPP, Rothschild LJ, Billi D. 2016. Sustainable life support on Mars - The potential roles of cyanobacteria. Int J Astrobiol 15:65–92.
(3)Egan M, Kreso J. 2011. Expediting Factors in Developing a Successful Space Colony. Worcester Polytechnic Institute, Massachusetts.
(4)Wolchover N. 2012. What are the Limits of Human Survival?. Live Science.
(5)Walter T. Oxygen and Human Requirements.
(6)Williams DR. 2018. Mars Fact Sheet.
(7)Andre M, Massimino D. 1992. Growth of Plants at Reduced Pressures: Experiments in Wheat - Technological Advantages and Constraints. Adv Space Res 12:97-106.
(8)Caceres PG. 2009. IMoM-6A. PDF.
(9)Zip Tie Domes. 2018. What is Geodesic Dome Frequency? An Explanation.
(10)Williams M. 2016. How bad is the radiation on Mars?. Universe Today.
(11)Catling DC, Cockell CS, McKay CP. Ultraviolet Radiation on the Surface of Mars. NASA Ames Research Center.
(12)Geng J, Yu SB, Wan X, Wang XJ, Shen P, Zhou P, Chen XD. 2008. Protective action of bacterial melanin against DNA damage in full UV spectrums by a sensitive plasmid-based noncellular system. J Biochem Biophys Methods 70:1151–1155.
(13)Tokyo Tech iGEM Team. 2009. Protection from UV ray.
(14)Kanehisa Laboratories. 2018. Tyrosine Metabolism - Aspergillus oryzae.
(15)Zhou Z, Dang Y, Zhou M, Li L, Yu C, Fu J, Chen S, Liu Y. 2016. Codon usage is an important determinant of gene expression levels largely through its effects on transcription. Proc Natl Acad Sci 113:E6117–E6125.
(16)Yang Z (Joey), Zhang F, Still B, White M, Amstislavski P. 2017. Physical and Mechanical Properties of Fungal Mycelium-Based Biofoam. J Mater Civ Eng 29:04017030.
(17)Ram AFJ, Arentshorst M, Damveld RA, vanKuyk PA, Klis FM, van den Hondel CAMJJ. 2004. The cell wall stress response in Aspergillus niger involves increased expression of the glutamine: Fructose-6-phosphate amidotransferase-encoding gene (gfaA) and increased deposition of chitin in the cell wall. Microbiology 150:3315–3326.