Human Practices

iGEM Guelph is from Ontario, and it is the home of 14.8 million people, but 2.3 million live with food insecurity. Leamington is a small Ontario municipality with 28,000 permanent residents (Government of Ontario, 2022; Westoll, 2022; Municipality of Leamington, 2020). Leamington is the greenhouse capital of Canada and as such, a centre for agriculture innovation (Municipality of Leamington, 2020). It is also a major producer of tomatoes and approximately 3 hours away by car from Guelph. Being so close to Leamington and at a university with a strong agricultural background, iGEM Guelph was inspired to work on an agriculture based project, now called Project Ceres. Project Ceres is a biopesticide designed to manage fungus gnats in greenhouses and thus contribute to growers producing a higher yield. Ours is a project built around a responsible human-focus at its core, as we try to create an innovative and sustainable model that can be used to address food insecurity.

Stakeholder Analysis

Here at iGEM Guelph we first wanted to make sure that we had strong foundations on the values guiding our project. We knew we wanted to have a focus on a project that is sustainable and has a low environmental impact, is curiosity-driven, based on elevating our current scientific understanding, and accessible. When initially meeting to select the project based on these values, both our PIs and grad advisors also reminded us to make sure that we had a project that most importantly is achievable within the competition time-frame. This meant that we were at first determined to achieve project success but narrowed down our focus from crops in greenhouses affected by pests and diseases to focus on one disease that affects one crop grown in greenhouses. Through an initial literature review we found that Ontario is Canada’s biggest greenhouse producer, and that our most common greenhouse crop is tomatoes (Agriculture and Agrifood Canada, 2021). There are many insect pests and fungal diseases which can occur in greenhouse tomatoes, many of which are not specific to tomatoes, but rather to the greenhouse environment in general (Parker et al., 2008). In particular, we chose to focus on fungus gnats as our target pest because they are a common vector for fungal disease in greenhouse tomatoes and other greenhouse crops (Calpas, 2022). Control of fungus gnats is a widely accepted method for prevention of fungal disease in greenhouses (Agriculture and Agrifood Canada, 2006). Fungus gnats also fall into the order Diptera which they share with drosophila, and this means we can use a model organism in place of fungus gnats for our toxicity assays. It was through these literature searches and consultation with our advisors that we started to get a clearer picture of what our project could look like. While ultimately our project, a biopesticide targeting fungus gnats for tomato growers was investigated, the road to get there required a lot of community and scientific feedback.

As our project took shape and lab work began, the business team started market analysis and determining key stakeholders. We wanted to know who our project would impact, both directly and indirectly. Through consultation with the J.F. Wood Centre for Business and Student Entreprise and the iGEM Entrepreneurship Business bootcamp, we determined that our key stakeholders were: greenhouse growers, growing associations, and experts.

Growers

Our target market is large-scale greenhouse growers in Ontario, although we hope to expand our market nationally and then across North America. To get a sense of the growing season that greenhouse tomato growers experience we reached out to growers in 2 ways: by phone and by a survey sent out by email. We were unable to reach the majority of growers by phone, so we switched tactics and sent out an email containing a survey. From those that responded to the survey, we learned that fungus gnats, mites, aphids, and white flies are a major problem for greenhouses. They also indicated that the ideal application method for our biopesticide would be compatible with an irrigation system.

Given that we wanted to balance focusing on one pest and crop but having a project that could be applied on a global scale, these responses made us curious about the problems faced by other types of greenhouses. Ultimately we also conducted community consultations with members of flower greenhouses in Ontario. We learned that aphids and thrips are a major problem. From these consultations and the consultations with greenhouse associations, which is discussed later, it appears that most of greenhouse pests have a root zone factor, and as such our project may also be applicable to flower and cannabis root zone diseases. One flower greenhouse said that they have a pesticide routine that works and they’ve stuck with it for over a decade so they’re not keen on changing it. However, most other growers we consulted appear to welcome change, granted that the product works well for them. They also seem to prefer a powder that can be incorporated into the soil or a water soluble biopesticide that can be used in a drip system. This feedback gave us a lot of optimism because while different crops have different biopesticide requirements, a key learning here is that some of the pests are shared between crops and an ideal delivery system would naturally be universal. This helps our proof of concept address global next steps, as if we develop a good delivery system, then all that needs to be changed are the genes depending on the crop and target pest!

Greenhouse Associations

We met with Canadian greenhouse associations to further gain insight into grower pest problems. They shared that aphids, thrips, and white flies were a huge concern for growers. Mainly pests with a soil stage of development were a major problem for growers, which makes our product highly applicable! Pests however were mentioned to be cyclical and could be a major problem one year and then not as big of a concern for another 4 years. Also, they shared information about the pesticide registration process and that registering is difficult and timely, however they also referred us to the pre-submission consultation form for the pesticide registration process with the Pest Management Regulatory Agency (PMRA), where we could get feedback on our pesticide application before we officially apply for pesticide registration through the PMRA.

Experts

When thinking about accessibility, we knew that we had to find an official channel that we could share our biopesticide with others through. The most common response we received when consulting externally, was through creating a provisional patent that we could cheaply licence to others. So this year iGEM Guelph consulted an intellectual property (IP) specialist, David Hobson, to learn the specifics behind creating a successful patent. David informed us that to be granted a patent, the idea must be novel and that patenting can cost $30,000 or more for a 20 year patent which may take 5 years to be approved. We also need to do landscape analysis to see if there are any current patents or products on the market similar to ours, as we may not be able to apply for a patent in that case, since it would be patent infringement. As we are a student run team, applying for a patent is not a current priority as our ideas will still be protected due to the window of time that one has before they are unable to apply for a patent. David suggested we publish a paper about our work, which we hope to do in the near future!

Another specialist we met with was an entomologist, Dr. Ayyanath, for a professional opinion on whether we met the pest management needs of growers through our biopesticide. We discussed how we can better our project to suit the needs of growers and build a successful pesticide. We talked about what growers look for in a pesticide and how we can meet those expectations as scientists. Also, we discussed the proposed application methods for our pesticide and how we can make our biopesticide water-soluble. Quorum sensing is key to making our bacteria effective for water-soluble delivery in an irrigation system. It is the method that bacteria use to communicate with each other in a given area. We could utilise this to determine how much of our bacteria survives in a hydroponic system and utilise the optimal growth environment for our bacteria to ensure it’s survival in our grower’s hydroponic systems, while avoiding overgrowth. This is explained in further detail on the Best Supporting Entrepreneurship page, under the Quorum Sensing and Future Development sections.

Conclusion

Through our HP work, we relied on our core values to help drive our team, and our consultations helped us realise there is a gap that we need to fill for our project to be brought to market. What it will take to fill this gap will further be explored in the Integrated Human Practices section. Many thanks to those who advised us on our project and to those who spoke with us!

Integrated Human Practices

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Through our Human Practices work, we have developed Project Ceres from being a generalist crop biopesticide into a greenhouse biopesticide targeting insect vectors of disease. Our specific target was fungus gnats and we hoped that by targeting this pest, tomato greenhouse growers would produce higher yields and further food security locally in Ontario. We have met with various stakeholders and incorporated their advice into our project to create a project that builds on our values of accessibility, sustainability, and elevated scientific understanding, while also meeting the needs of growers.

Changes Made from Consultations

Moving from Field Crop Farmers to Greenhouse Farmers

Based on our Human Practices work, our consultations helped guide and inform the first major change that we implemented. We started this project looking solely at helping field crop farmers, as we thought it the most globalised form of agriculture with a form of farming that most growers participate in. However, we very quickly realised from talking to our advisors and through literature such as Beckman and Rüdelsheim (2020), that we needed to change our first end-user from field crop farmers to greenhouse growers. This is because pesticides are contained within the greenhouse environment and only utilised once insects pass a threshold level, or more simply as a last resort, therefore, we can minimise greenhouse gases historically associated with pesticide use (Heimpel et al., 2013) and build a more green connection between agriculture and the environment. The change to focusing on greenhouses also helped strengthen a tangible outcome from the multiple values we were approaching our project with such as accessibility, low environmental impact, and achievability.

Namely, when thinking about accessibility of this project to a global scale, one of the top issues was the public acceptance and willingness to use a genetically engineered biopesticide. And while this is a problem that isn’t restricted to our specific project, but an issue trying to be addressed with synthetic biology as a whole; our project’s pitch for global adoption warrants closer consideration as this technology would be primarily used by rural farmers.

Genetically modified organisms (GMOs) remain polarising within wider society as people often are distrustful of it for diverse reasons and concerns. We know that:

• Levels of education and understanding the technology is a major hurdle.
• There is an historical argument about how new this technology is, leaving many in society concerned about the “unnaturalness” of GMOs due to the intentional editing of the genome.
• There is also a concern about these edited organisms escaping into the environment and being able to outcompete or harm natural members of the ecosystem.

While concerns 1 and 2 are related to education about the technology (and tied in with our values about elevating scientific understanding and accessibility), this was something we attempted to tackle, localised to our own campus community first. We hosted events for new students such as our orientation week event and a table at the University of Guelph’s clubs fair. Through these two events we were able to reach an audience of more than 100 people about our work and the intricacies of synthetic biology. To further the outreach of these events, and thanks to popular demand, we will be running seminars on synthetic biology for the student community at the University of Guelph. As these concerns did not impact project design, you can find out more about this in our Education and Communication page. This third concern of organism escape into the environment is linked to our value of a sustainable and low environmentally impactful project. Thus, organism escape was one of the most important hurdles our team identified that the project could directly address.

Here we realised that our values started to overlap and build off each other. We saw that public acceptance was needed to make this project accessible. However, the acceptance of the technology relied first on that technology having a low environmental impact. Greenhouses helped us address this concern inherently because they are controlled environments where biomaterials moved in and out of the greenhouses are closely monitored and checked for pests anyway. This means that the switch to greenhouses helped us focus on the science of the biopesticide, and also built off of another value of the project which is achievability within the competition timeframe. This is because in the early developments of the project, our team had to consider biosecurity through genetic means because we did not want our organisms escaping into the environment through wastewater or aerosolizing sprays. However, the kill-switch mechanisms we were exploring would require weeks to months of design and testing that would take away from testing the biopesticide itself. So moving to a controlled environment where designing and engineering biosecurity measures isn’t a priority because of greenhouse policies created breathing room for our project design leads. Greenhouses also help address acceptance to concern number 3 just because the modified organisms are not being used in open fields where field crop farmers grow their crops. As well, because the nature of greenhouse agriculture in Ontario is generally tech-forward, greenhouse growers tend to be more accepting of synbio technology.

Another safety feature of our project made based on our reflection of literature was the alteration of the use of Cry proteins to Cyt proteins for target specificity. Cry proteins have a broad spectrum of target species including pollinators, which is unacceptable for the Pesticide Registration process, however Cyt proteins are very target specific and our selected Cyt proteins pose no risk to species outside of the target order Diptera (Valtierra-de-Luis et al., 2020). This target specificity paired with our implementation of an IPTG-responsive promoter (Pgrac) means that in the hypothetical situation of our organism “escaping” a greenhouse, it would pose no threat to natural members of the ecosystem. Please read the Safety section under Proposed Implementation and the Safety page on our wiki for more information.

Expanded End Users

Through our HP work and consultations with greenhouse associations, we learned that our project is also applicable to cannabis and flower root zone diseases. Thanks to this information, we expanded our target market from fungus gnat management in tomatoes to soil-based pests including aphids, thrips, whiteflies and spider mites in flowers and cannabis. Specifically, we will market our current biopesticide model to poinsettia growers as it appears that poinsettias are particularly challenged with fungus gnat infections thanks to the feedback from the growing associations. We will need to modify the cytotoxins in our biopesticide to be more pest specific for root zone pests like aphids, that affect flowers and cannabis, due to the root protection that our biopesticide’s biofilm provides growers with. In particular, root aphids are a problem that is fairly unique to Cannabis crops, which our project happens to be particularly suited to address. Cannabis growing is a relatively new field so there are not many pesticides for cannabis growers so we hope to provide them with more options to grow healthy plants. Our Human Practices section can be reviewed for more information.

In the near future, we will further research flower and cannabis growing and explore the applicable legislation and growing permits (especially for cannabis) for testing our biopesticide in the growing environment. We plan to further explore the cannabis and flower grower avenue after the Jamboree and will conduct more HP work similar to our work done for tomato growers.

Likewise to after each HP meeting, we will reflect on potential barriers caused by these modifications to Project Ceres and its impact on commercialization. These barriers are discussed on the Best Supporting Entrepreneurship page under the Barriers, and Limitations and Future Research sections.

Project Reflection

As our work term for the iGEM competition is coming to an end, we’d like to reflect on our work and project. Throughout our project we focused on environmental impact, improving food security, and scientific accessibility and education. While we did have to compromise on the inclusion of a kill switch due current kill switches being non-viable options as a result of our biopesticide’s target environment, we sought other safety measures such as the exclusion of the fertility factor to prevent genetic transfers between our modified bacteria and its natural counterpart (refer to the safety section under the Proposed Implementation page for more information) and thus, we ended up being extremely proud and optimistic about our project progress this year. We designed our project to be as minimally impactful on the environment through promoter choice to greenhouse use, and will continue to improve our team’s goal of environmental health through the pesticide registration process and exploring methods for growers to safely dispose of our biopesticide after use. Also, we have improved scientific understanding of genetically modified organisms in our community at the University of Guelph through various outreach and hope to improve further understanding of genetically modified organisms through offering synbio seminars (which we have started); and also attending research and greenhouse conferences, which we were unfortunately unable to do this year. We hope by attending these conferences we can further improve our goal of global food security starting in the Province of Ontario and through local greenhouse production.

To all those interested in collaborating with us in our goal of improving food security, please feel free to reach out to us!

Works Cited

Beeckman, D. S. A., & Rudelsheim, P. (2020). Biosafety and Biosecurity in Containment: A Regulatory Overview. Frontiers in Bioengineering and Biotechnology, 8, 650–650.

Heimpel, G. E., Yang, Y., Hill, J. D., & Ragsdale, D. W. (2013). Environmental consequences of invasive species: greenhouse gas emissions of insecticide use and the role of biological control in reducing emissions. PloS One, 8(8).

Valtierra-de-Luis, D., Villanueva, M., Berry, C., & Caballero, P. (2020). Potential for Bacillus thuringiensis and Other Bacterial Toxins as Biological Control Agents to Combat Dipteran Pests of Medical and Agronomic Importance. Toxins.

Agriculture and Agri-food Canada. (2006). Crop profile for greenhouse tomato in Canada. Government of Canada.

Agriculture and Agri-food Canada. (2021). Statistical overview of the Canadian greenhouse vegetable industry, 2020. Government of Canada.

Calpas, J. (2022). Pest and disease management in commercial greenhouse tomato production. Alberta.

Food and Agriculture Organization. (2021). The state of food security and nutrition in 2021: The world is at a critical juncture. United Nations.

Food and Agriculture Organization. (2022). Sustainable development goals. United Nations.

Government of Ontario. (2022). Ontario population projections. Ontario.

Masudi, F. (2014). Greenhouses key to water and food security in UAE, expert says. Gulf News.

Monks, K. (2019). Can desert greenhouses diffuse food security time bomb. CNN.

Parker, B. L., Skinner, M., Brownbridge, M., & Doubleday, T. (2008). Greenhouse manager’s guide to integrated pest management in Northern New England. Entomology Research Laboratory, University of Vermont.

United Nations. (N.D.). Causes and effects of climate change. United Nations.

University of Windsor Daily News. (2022). Role for greenhouse growers in national food security subject of discussion. University of Windsor.

Westoll, N. (2022). This is a very serious problem: New report shows food insecurity persists in Ontario. City News.

Municipality of Leamington. (2020). Community profile.

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