Crops are facing multiple stressors

Agriculture is one of the most important economic sectors, essential for sustaining human life as it contributes to approximately 90% of food calories^1–3. As population increases, crops are becoming more and more essential, and their production needs to be as efficient as possible to guarantee food safety. However, multiple stressors, both biotic and abiotic, affect productivity and crop quality (Figure 1) . Abiotic stressors include drought, salinity, and heat, while examples of biotic stressors are parasitic plants, fungal pathogens and insects⁴. These stressors affect crops from the seed germination stage: abiotic stress is a major challenge, and different chemicals produced by biotic stressors can inhibit seed germination^5,6.

What can we do to help tackle these problems? We believe the answer lies in seed coatings.

Seed coatings are exogenous materials applied on top of the seed’s natural seed coat and can increase crop yields by 20-50%⁸. They help agriculture by modifying the physical properties of the seed, through the standardisation of its weight and size, and by precision delivery of active ingredients, such as pesticides and herbicides⁹. This technology has already been used for years in developed and developing countries¹⁰, but is still under improvement. For instance, current seed coatings lack modularity, as it is difficult to adapt one type to different needs. They use multiple materials for different purposes, and still rely largely on synthetic polymers, which contribute to microplastic pollution. Our project aims to use synthetic biology to create a fully biodegradable seed coating with customisable properties. In this way, we can save future crops by improving seed quality and germination, while preventing biotic stressors through the embedding and controlled release of active compounds.

Current seed coatings can help but have limitations

Although the application of commercially available seed coatings can already bring an advantage to agriculture, current seed coatings still have many limitations. One major drawback is the lack of modularity: no modular seed coating is available on the market and different materials are used for specific purposes. Most seed coating formulations also contain synthetic polymers, as they can be easily modified to obtain the desired properties¹¹. However, they contribute to microplastic pollution during production, through their unintentional release, and during use, through the release of microplastic dust¹². This poses a major risk to the environment due to their poor degradability and release of toxic additives, which can also cause adverse effects on human health¹¹. Soon synthetic polymers will no longer be an option, as the EU chemical legislation REACH established that, by 2028, deliberately added microplastics will be restricted in agricultural products, including seed treatments. Furthermore, by 2031 the restriction will extend to all crop protection products and treated seeds. Although more sustainable seed coating options are available, in addition to their advantages, they also present major drawbacks, as highlighted in Table 1.

To tackle the many crop stressors and ensure food safety, improved seed coatings are needed. The ideal seed coating should be fully biodegradable and modular, with properties customised to specific needs, allowing it to efficiently embed and release active compounds. Through extensive research and stakeholder engagement, we came to the conclusion that bacterial cellulose (BC) is the perfect material to serve this purpose.

Why bacterial cellulose?

Cellulose is the most abundant natural polymer on earth, and can be produced by diverse life forms, including bacteria¹⁴. BC, like other cellulose counterparts, is a network of linear homopolysaccharides made of β -1,4 D glucopyranose units^14,15, produced by many bacteria belonging to the Gluconacetobacter and Komagataeibacter genera. These species create a BC shield to protect themselves from stressors such as desiccation and UV damage¹⁶. BC has many properties that make it an intriguing material for seed coatings (Figure 2) . It is completely biodegradable, biocompatible, with high hydrophilicity and water holding capacity (WHC)¹⁷. Compared to other forms of cellulose, it also has higher purity (due to the absence of lignin, pectin and hemicellulose), mechanical strength and modifiability¹⁸. BC has many reactive hydroxyl (-OH) groups, which allow for side-chain modifications and this can be exploited using chemical means. However, we decided to focus on a more sustainable approach by implementing enzymatic treatment instead of using traditional chemistry. In this way we could change properties and even increase WHC, beneficial for fighting drought stress.

All these properties are excellent and desirable for a seed coating, and we believe they can be customised using synthetic biology. As a result, we can obtain tailored seed coatings, that will help specific applications.

Our project

With BCoated, we set out to produce a completely biodegradable and modular seed coating made of BC produced by Komagataeibacter sucrofermentans. With the use of synthetic biology, we seek to customise the properties of BC so that it can be tailored for a wide range of agricultural applications. Our project is divided into three main phases: functionalisation, production platform and seed coating & application (Figure 3).

Within functionalisation, we worked on obtaining BC with different, customisable properties. In particular, we focused on producing and embedding proteins onto the matrix of BC, and tuning WHC, biodegradability, and porosity. In this part we also developed a part collection with CBD fusion proteins secreted by Saccharomyces cerevisiae. This collection allows for the production and embedding of proteins to happen in situ, while BC is being produced in a consortium.

In the production platform our focus shifted to the yield, from evaluating production conditions to culturing our BC-producing strain, K. sucrofermentans, in a consortium with S. cerevisiae. We also designed a synthetic circuit to maintain the ideal ethanol concentration, that we expect will increase the overall BC yield.

To predict the behaviour of this consortium, we created two models: the first model focused on describing the consortium dynamics, while the second model focused on the simulating the behaviour of the ethanol concentration circuit in S. cerevisiae.

The final part, seed coating & application, is where our produced BC is finally used as a seed coating. To coat the seeds, we performed an in situ assay, in which BC was produced directly around the seed. In addition, we focussed on applications: we developed two use cases, aimed at fighting parasites that are currently damaging crops.

Learn more about our project design

Learn more about our wet lab

Learn more about our dry lab

The role of synthetic biology

Synthetic Biology (SynBio) is a scientific field that applies engineering principles to living organisms and living systems, with the aim to develop useful new biological parts or redesign existing systems found in nature¹⁹. SynBio is at the core of our project as it allowed us to extend seed coating potential through modularity and protein attachment. Design–Build–Test–Learn (DBTL) cycles were the foundation of our workflow and helped us achieve our engineering goals. For instance, we applied this engineering approach in the development of our part collection and in the design of a synthetic ethanol circuit in S. cerevisiae. Furthermore, our BC producing organism, K. sucrofermentans, is very difficult to transform and we developed a successful protocol to do so, further contributing to the SynBio community.

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Creating an impact

BCoated’s journey does not end with the lab: our team also focused on creating an impact by talking to experts and developing an entrepreneurial plan.

We talked to stakeholders from many different backgrounds, including arable farmers, actors in the seed sector, material scientists, BC manufacturers and regulatory experts. Their opinion was crucial for shaping our project: they helped us identify current agricultural issues, choose our BC-producing strain and bioreactor, and in finding a suitable feedstock.

Additionally, entrepreneurship programs and facilities helped us develop an entrepreneurial plan for our project. As a result, we created a business model, taking into account potential risks and benefits. However, this was just the beginning: by conducting a skill gap analysis, we defined each team member’s future role within the company and identified the skills we still needed to acquire.

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Learn more about our entrepreneurship

Conclusion and outlook

We created BCoated with the aim of enhancing the potential of BC and extending its application to seed coatings. We shaped our project based on the insights obtained from experts in different fields, including farmers, material scientists, BC manufacturers and regulatory experts. Their opinions were also crucial in developing our business plan, which allowed us to evaluate the feasibility of our project to become a company.

Our project put a significant step forward in the development of a modular and sustainable seed coating. Through our efforts, we created a part collection that will help the in situ incorporation of beneficial proteins in a BC seed coating. However, these parts can also be useful in other sectors that benefit from the use of BC as we truly want to help future researchers and iGEM teams expand BC's potential.

Our vision is to see BC seed coatings become a sustainable tool to enhance crop yield, and for BC to become a highly customisable material, tailored to different agricultural applications. We forsee our project further expanding its functional capabilities, supporting farmers in achieving healthy crops. Together, we hope to obtain an ideal material for a sustainable future and use tailored coatings to help tomorrow’s crops.