This thesis explored the development of self-healing, sustainable materials using Basidiomycetes fungi, with a particular focus on their ability to withstand high temperatures.

The primary goal of this research was to identify Basidiomycetes species capable of surviving the thermal processes typically applied post-production of mycelium materials. A key finding was that the species S. commune and G. sessile could revive consistently across growth in various media even after drying at temperatures up to 100°C. Other species like T. gibbosa and T. hirsuta showed less consistent revival at these temperatures, whereas T. versicolor and B. adusta only revived at lower temperatures (40°C and 60°C). Further investigations assessed whether stress factors such as nutrient depletion and pH reduction during growth could trigger these survival mechanisms, potentially boosting the resilience of mycelium-based materials. The study found that harsh conditions, like MM and low pH levels, greatly hindered mycelium development. Microscopic analysis identified chlamydospores in G. sessile across all conditions, indicating their importance in mycelium regeneration. In contrast, conditions of N-MM or slightly acidic pH induced the formation of asexual spore structures in S. commune, suggesting their potential role as protective agents under stress.

Catho Vanclooster (2023-2024)

Supervisors: Elise Elsacker, Eveline Peeters

Master of Science in Bio-Engineering Sciences: Cell and Gene Biotechnology -Molecular Biotechnology

Mycelium-Based Composites (MBC) are innovative bio-materials made from fungal mycelium, which acts as a natural glue to bind agricultural or industrial waste fibres. These materials are sustainable, biodegradable and require low energy to produce, making them attractive for green construction and circular economy practices as replacement for fossil-fuel-based materials. The characteristics of MBC can be tuned by adjusting various factors, such as fungal species, substrate, or environmental conditions, to achieve desired properties.

This master’s thesis aims to explore the influence of different production parameters on the growth of MBC. A literature review and patent analysis highlight the need for standardized production processes and detailed descriptions. The first part of the thesis develops a framework to understand the numerous parameters affecting the fungal development. The experimental phase assesses mycelium development through visual assessment, whiteness quantification, dry weight loss, shrinkage percentage, density increase and water absorption. Various initial water contents (20, 30, 60 and 90 wt%) and wheat bran concentrations (0, 5, 20, and 40 wt%) are examined over a period of 15 days.

The proposed methods aim to quickly identify influences and explore the tuneable characteristics of MBC. The results indicate that the methods are useful for observing trends and peaks. However, the variability inherent in working with living organisms introduces heterogeneity, complicating the comparison of values.

In conclusion, the research highlights the importance of describing the production process to enable meaningful comparison of results. The identification of parameters and their interrelations is crucial and represents the first step towards upscaling and industrialising of MBC production.

Florence Van Laethem (2023-2024)

Supervisors: Lars De Laet, Elise Elsacker

Master of Science in Architectural Engineering

The construction industry is a major environmental polluter. Bio-based materials like Mycelium-Based Composite (MBC) offer sustainable solutions. MBC is derived from fungi and is biodegradable and energy-efficient. Permafungi, a Brussels-based company, repurposes waste from oyster mushroom farming into MBC. They aim to produce more of MBC monthly for packaging and construction, using their spent mushroom substrate. Despite its potential, MBC faces challenges like low mechanical strength and slow production, requiring further research and industrial scaling to compete with synthetic materials. This master's thesis aimed to define the physical and compressive mechanical properties and optimize the production process of mycelium-based composites (MBC) using spent mushroom substrate (SMS). A comprehensive literature review was conducted followed by experiments based on different stages of the MBC process. The main findings of this study were the following. Skipping sterilization saves time and resources but results in minimal or no mycelium growth, highlighting its importance. However, current sterilization techniques have high energy costs, raising sustainability issues in the context of industrialization. Adding small amounts of hemp and beechwood to SMS improves mechanical properties without significantly affecting mycelium growth. Specifically, 30% beechwood greatly enhances compressive strength but slightly reduces mycelium growth. SMS with one fructification o (SMS F2) and SMS two (SMS F2) have distinct impacts; SMS F2 shows less mycelium expansion but better mechanical behavior due to its initial higher Pleurotus ostreatus content. Adding new substrate to an inoculated one reduces mycelium growth and results in uneven fungal skin, indicating challenges in uniform colonization. This method, while promising for faster production, proved unsuitable for use. The study concluded that using SMS for MBC is a viable and sustainable alternative to common substrates, with small substrate additions showing significant improvements.

Isabelle Rodrigues Malinowska (2023-2024)

Supervisors: Lars De Laet, Elise Elsacker

Master of Science in Architectural Engineering

Fungal biomatter, a new material in the early stages of development, has gained the interest of various disciplines. Researchers have been exploring the fabrication methods and growth parameters of mycelium-based composites and their potential use as a sustainable material. However, since mycelium research is still in its early stages, there is a gap in the available literature. So far, there has been no research on using bioadhesion in the fabrication process of mycelium based composites. Such a process may offer the potential to upscale the production of fungal biomatter and further expand its range of application. This thesis focuses on understanding the material properties of mycelium-based composites, and explores the possibility of using bioadhesion as a new fabrication method which can be utilised to fine-tune the properties of these composites and enhance their mechanical performance. In order to gain a deeper understanding of this new material and develop a new fabrication method which may be used to upscale production, composites are studied in their most basic form. Mycelium spawn and plant based substrates were the only materials used, in the fabrication of composites, for the experiments conducted in this thesis. No pre-mixed material, nutritional sources or aggregates were added. This strategy was used to ensure that the basic composite behaviour is studied and the mechanical results of samples are analysed in their raw form, creating a baseline from which further research can be developed.

Dana Raslan (2021-2022)

Supervisors: Martyn Dade-Robertson, Elise Elsacker

Master of Architecture Linked Research Project at Newcastle University

For over thirty years, the biological degradation of plastics by microorganisms has been promoted through microbiological publications as an eco-friendly alternative for landfilling and incinerating – a clarion call for change – yet today these are still the most economically viable ways of discarding plastics. What's the price of money? Therefore, this master thesis has set out to investigate whether mycelium composites could valourise plastic waste and inject the bioremediation process of plastics into an economically viable material for an industry itself in dire need of sustainable material innovation: the building industry.

Lennert Van Rompaey (2019-2020)

Supervisors: Elsacker, E., De Laet, L.

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

This master thesis attempts to re-join two innovative fields of architecture: additive manufacturing and sustainable living construction materials such as mycelium-based composites. Our main hypothesis is that hollow 3D-printed geometries can provide an efficient oxygen distribution for the fungal growth, maximise the surface of fungal skin and thus improve the mechanical properties of the sample. The objective is to investigate the impact of various infill patterns and infill density on the growth of mycelium composites. Two different experimental methods were performed. The first consisting of 3D-printing sample geometries with a printhead placed on a KUKA 6-axis robot, was difficult to implement due to poor extrudability of the substrate. The second method produced typical hollow geometries with an infill pattern and an outer shell by using a special mould designed and made with a plastic 3D-printer. The results suggest that hollow geometries can provide an efficient oxygen distribution to facilitate fungal growth. However, they did not maximise the surface of fungal skin as we expected but rather led to the formation of a pure foam of mycelium within the cavities.

Noam Kalai (2019-2020)

Supervisors: Elsacker, E., De Laet, L.

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

Mycelium composites are promising bio-composites in research and are labelled as biodegradable. However, no research publication that tests this material characteristic is found. Hence this master’s thesis attempts to find a method to degrade mycelium composite samples and interpret first results. For the methodology, soil burial tests under predefined laboratory conditions are applied in which samples are buried in soil and retrieved after chosen burial intervals. The weight of the sample before and after burial is compared and a weight loss percentage is derived. Additionally, pictures of the samples before and after burial are compared. In general, it can be stated that mycelium which binds everything as a whole tends to loosen first. A first time indication of the degradation rate of mycelium is challenging as this research is explorative and as it depends on the used substrate, fungal strain, additives, production method and external environmental properties. When determining the average of inert samples in the first series of the final tests, a weight loss percentage of approximately 44% is achieved after 16 weeks. If this is extrapolated in a linear trend, complete weight loss of the composite samples would be achieved in about 36 weeks.

Li Li Yap (2019-2020)

Supervisors: Elsacker, E., Van Wylick, A., De Laet, L.,

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

Architectural tensile membrane structures are temporary or limited-lifetime structures that are mostly fossil-fuel based and recycling is limited. In this thesis bacterial cellulose, a sheet material grown at the surface of a culture liquid by a bacteria, is explored and assessed for the first time in the light of an application as structural membrane. The aim is to define whether bacterial cellulose could replace or complement today’s common tensile membrane structures. This is done through a wide exploration of alterations of the plain material with a focus on post-processing such as soaking, coating, heat pressing, creating composites and mixing. Also the creation of connections between sheets is explored. All experiments are subjected to mechanical tensile tests. Also water absorbency and appearance are described. The approach is quite unusual, as academic literature was combined with discussion in an online DIY forum. The results of the thesis are satisfying. Three alterations of bacterial cellulose improved its strength to a competing level with today’s common membrane materials. Furthermore a self-assembled seam connection implying only the drying together of sheets was discovered, a concept to deploy a membrane structure based on this observation was developed.

Bastien Damsin (2018-2029)

Supervisors: Elsacker, E., De Laet, L.

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

By living in an era where creativity and environmental awareness is rising, alternative materials are upcoming. An example of such new materials are mycelium composites. By being fully biodegradable due to the natural components, applications in the construction sector are investigated. Mycelium is mainly known in small projects. However, the knowledge about fabrication on large scale is lacking. Therefore, this master’s thesis aims to develop a protocol to manufacture these biological materials on large scale. After having an overview of the background of mycelium and already printed results, the first testes are launched. The investigation is done by means of hands-on experiments. Different possible compositions and manufacturing methods of the viscous substrates, containing the living organism, are discussed. Subsequently, the manufacturing is adapted to the use of a KUKA robot by developing a protocol. Eventually, conclusions and reflections on the obtained substrates and printing results are given.

Ichelle Nieberding (2018-2019)

Supervisors: Elsacker, E., De Laet, L.

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

This study is one of a kind, in that it links insights into the expression levels of lignin-degrading enzymes to direct industrial applications during the production process of these composite materials. It is also unique in that it is the first study that explores the expression levels of all individual laccase isozymes in Trametes versicolor. Insights are generated on the incorporation of citric waste streams and their effects on the expression of ligninolytic enzymes and the growth on various substrates. It is found that citric peelings induce the expression of three laccase isozymes on a translational level, as well as exert a general induction of laccase isozymes on a transcriptional level and that growth on plant-based waste streams occurs faster in the presence of citric peelings. 

In this study, well-established techniques such as gene expression analysis by quantitative PCR and enzymatic assays are combined with newly developed experimental setups, such as calorimetry and mechanical stress tests, to analyze the production process and obtained mycelium-based materials, which are novel and innovating materials. As such, first steps are taken towards establishing new techniques for the analysis of mycelium materials, thereby providing a steppingstone for future research. The observations concerning the incorporation of citric peelings and the effect on the expression levels of laccase isozymes add to our growing knowledge on the regulation of these genes in white-rot fungi. In addition, it opens the door to capturing a large biological waste stream and thus creating a more circular economy.

Berend Bassleer (2018-2019)

Supervisors: Vandelook, S., Elsacker, E., Peeters, E.

Master thesis in Biomolecular Sciences at Vrije Universiteit Brussel

This report describes the matter of an internship at Vrije Universiteit Brussel (VUB), in the department of Physical Chemistry and Polymer Science (FYSC), together in collaboration with Microbiology research group within the Bioengineering Sciences Department which has become crucial for my practice. FYSC department is currently dealing with research topics such as self-healing materials or recycling of cementitious materials and Microbiology research group targets on creating composite biomaterials using fungal mycelium. My laboratory experiences included, moreover, the practice on analyses such as FTIR spectroscopy, rheological measurements, SEM operating and microbiological samples inoculation, cultivation and follow-up. All necessary analyses and experiments were carried out every 24 hours on fungi-colonized substrates comprising different types of woods or grass, during its growth. The results of these tests are supporting an ongoing research on mycelium-based composite materials that have relevance for architectural purposes. Such composite materials represent environmentally friendly, renewable thermal insulators and are likely to replace unsustainable synthetic materials like polyurethane and polystyrene.

Vendula Ficelov (2019)

Supervisors: Brancart, J., Elsacker, E.,

Internship

This dissertation proposes to research the environmental impact of mycelium composite building materials compared to conventional building materials. Initially, the state-of-the-art focuses on mycelium, mycelium composites and life cycle assessment (LCA). After a short introduction concerning the origin of mycelium, mycelium composites are reviewed. The aim is to generate an initial understanding of the mycelium composite production process, since the production process is still in a development stage and thereby adapting to possible future building applications. The last part of this literature study aims to provide the framework and guidelines for the conduction of a LCA case study, thereby also reviewing the Totem-tool for buildings and building components in Belgium. Following to the state-of-the-art, this dissertation aims to develop the general production process of a mycelium composite building material, based on a literature review and expert’s input (chapter 3). This research forms the basis for chapter 4, in which the environmental performance of a mycelium composite wall insulation panel is compared to a conventional PUR insulation material by means of a LCA case study. This research reveals that the mycelium composite production process is very unique and therefore requires a number of processes that define the very nature of mycelium composites. Another major finding from this research is that the environmental impact related to the production of a mycelium composite wall insulation panel is relatively high compared to the one of a conventional PUR insulation panel.

Nico Seghers (2018-2019)

Supervisors: Elsacker, E., Galle, W., Vandervaeren, C., De Laet, L.,

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

Mycelium composites are a new emerging material with high potential for sustainable and ecological applications. One of these applications is the use as a building material. However, most materials used in construction are already mechanically characterized; this characterization is currently lacking for mycelium-based composite materials. Therefore, this master’s thesis aims to contribute to the current state of the art of these materials by means of a series of mechanical tests in compression, tension and bending. A broad range of samples are thus tested and analysed by varying the constituents and manufacturing processes.

The overall background of mycelium composites is handled by focusing on the components and the already performed research. Based on the acquired information, the contribution of the thesis is discussed and the first experiments launched. After a thorough investigation on the possible manufacturing and testing methods, the performed experimental campaigns are discussed. A clear influence of the fibre type, pre-compression and humidity is witnessed on the mechanical properties but the current state of the material however underperforms in the case of structural applications.

Finally, reflections on the master’s thesis evolution and performed work are given and conclusions are drawn.

Aurélie Van Wylick (2017-2018)

Supervisors: Elsacker, E., De Laet, L.,

Master thesis in Architectural Engineering at Vrije Universiteit Brussel

Mycelium based biomaterials are a promising alternative for creating biodegradable packaging, construction and architectural materials. Molecular determination- and phylogenetic tools were used to characterisze genetic variation of different Trametes versicolor strains and to isolate a wild strain. Investigation of growth variations and ligninolytic enzyme production was conducted to collect insights on phenotypic strain variation. ABTS oxidation was used as means of measuring ligninolytic enzyme levels in absence and presence of ligninolytic enzyme inducers. Veratryl alcohol and citrus fruit peelings (mandarin & orange) showed induced overexpression of ligninolytic enzymes at translational- and transcriptional level. Gene expression of Laccase 4 and 5 genes were analysed with qPCR in presence of veratryl alcohol and mandarin peeling extract. Effects of inducers on growth and colonization rates of malt- and hemp agar was investigated. Ground orange peelings supported a high growth and development of mycelium. Waste streams of citrus fruit peelings could be valorised by growing Trametes versicolor for mycelium based biomaterials and thereby creating an economically viable application. Another potential application is the production of Laccases and other valuable biotechnological enzymes. The combination of both applications could result in the bioremediation of pesticides and pollutants present on fruit peelings used in agriculture.

Simon Vandelook (2017-2018)

Supervisors: Elsacker, E., Peeters, E.

Master thesis in Molecular & Cellular Life Sciences at Vrije Universiteit Brussel

Since the industrial revolution, the common way to produce materials for the construction sector has been by extracting finite valuable resources from nature, without taking their end of life and environmental impact into account. Plastic, concrete and synthetic composites are examples of such materials. Cement, for example, is the basis for the construction of any type of structure in reinforced concrete, whether tunnels, tracks, bridges, buildings. To obtain these elements it is necessary to extract from quarries (entire mountains), raw materials and mix them with water to ensure optimum grade. The scarcity of some of these resources has marked in the past the disappearance of complete empires and now threatens the present society. The environmental consequences are increasing, and the dependence of society on many of these resources marks the necessity for more studies of other types of materials and other ways to build. It is time to develop new options to convert the current consumer society into a self-sustainable society. One interesting way to stop this damage is through the employment of materials that can grow biologically, without generating polluting or toxic waste and that are totally biodegradable. A lot of construction applications can be developed, whether in matters of acoustic or thermal insulation, space division, light construction among others. For this, several species of bacteria colonies present in the vegetative part of fungi, mixed with natural fibers, have been recently studied. It was discovered that these composites form a material with a matrix based entirely on natural low-cost constituents, have many advantages. Although, it is still a very new topic which can be optimized. Therefore, examinations into the mechanical and physical properties of mycelium (vegetative part of a fungus made of a mass of branching, thread like-hyphae) based materials are useful.

Paola Pantoja Arboleda (2017-2018)

Supervisors: Elsacker, E., De Laet, L.

Master thesis in Civil Engineering at Vrije Universiteit Brussel

In a world where environmental problems can no longer be ignored, the demand for biodegradable materials rises. This thesis shows the design and realization of a multi material 3D-printhead for robotic additive manufacturing of biodegradable composite materials. This print head was placed on a KUKA robot, a commonly used industrial robot, because of its degrees of freedom and maneuverability. Furthermore, this robot arm has been equipped with an electro pneumatic installation to operate the print heads. The design of the three extruders was made in such a way that different materials can be combined while heterogeneity of the printed structures can be achieved. The biodegradable composite material which was used in the experiments consists of a viscous substance containing natural fibers. A strong network is formed between the fibers by adding mycelium (the root structure of mushrooms). Possible applications are temporary architectural structures.

Brecht Steenhouwer and Daan Van Den Cruyce (2016-2017)

Supervisors: Elsacker, E., De Laet, L., De Troyer, T., Standaert, L.

Master thesis in Industrial Sciences Elektromechanica [Mechatronica] at Vrije Universiteit Brussel