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The next steps

A big focus of our human practices outreach was to determine if and how our project could be implemented in the real world. By discussing with experts, we were able to understand the laws on GMOs and how they affected the feasibility of our project being used at a larger level. In Switzerland, the use of GMOs in agriculture is currently forbidden, making the phage part of our project impossible to implement here at the moment. However, this isn’t the case for the European Union, the market usually targeted by Swiss phytosanitary product production companies.

The tailocins and Antifreeze protein (AFP) solutions we designed only contain proteins, so applying them to plants is legal in our country. Furthermore, as discussed with one of our interviewees, apricots are a very profitable crop for farmers, the intended users of our product. It is therefore likely that they would invest in the protection of this crop, making the production of our protein-based solution at a large scale financially sound.

We tested our AFP solution on Arabidopsis thaliana to measure the difference in damage at -5 °C with and without treatment. As seen in the AFP portion of our proof of concept page, the AFP solutions successfully reduce damage due to the freezing temperature. We tested our solutions at 178 µM for Ffibp and 50 µM for riAFP. There was statistically no significant difference in the observed protective effect to the plants provided by both AFP solutions at the above-mentioned concentrations. Knowing this, we should test which dosage of AFPs would prevent the most damage caused by freezing (dose-response), and implement a spray at which the concentration is minimal but the protection optimal. We would also test the duration of the protection afforded by the AFPs by freezing Arabidopsis thaliana treated with different doses for various lengths of time. This type of experiment would allow us to determine an appropriate frequency of treatment, especially for the long lasting freezes. We would finally need to monitor plants on which we have applied our sample over a duration of 1-2 years to see how AFPs affect them. As discussed with our expert, Dr. Danilo Christen, the head of a research group on fruit crops in the Alpine region working at Agroscope, the Swiss centre of excellence for agricultural research, we also need to test the toxicity of our product on insects and auxiliary fauna, as well as its toxicity in water. Finally, we would need to check the toxicity of our product on humans.

After further confirming that our AFP protection system is functional when covering plant tissues and not toxic to the environment, we would pursue testing on apricot trees. We would need to start by testing our solution on apricot trees covering a small area for 2 years before allowing our product to be used at a larger scale. This way, we could track how our products affect the trees long-term. For a large-scale implementation, we would use tractors equipped with spray mechanisms to deliver the solution. This would be the best suited mechanism, as the apricot trees in our region grow on mountain sides, making the set up of other types of irrigation systems impossible.

We tested the tailocin solution using our cooling machine, FROZONE, and demonstrated that tailocins inhibit Pseudomonas’ ice nucleation activity through the lysis of the cells (see results) in vitro. The next step would be to use the tailocin solution on apricot plant leaves bearing their natural microbial communities (including pathogens), that would be harvested from the apricot tree fields. We could then test the plants for their ability to withstand a freeze artificially generated in the laboratory. We would also assess different concentrations and the duration/frequency of treatment on the plants with our product. Moreover, we would need to perform more in vitro assays testing the target specificity of the tailocins we used. We would also need to test the products on a wide range of Pseudomonas strains and natural cultivable strains present in apricot tree fields and its toxicity on insects, water, auxiliary fauna, and humans. This would allow us to estimate the potential ecological consequences of our products and its influence on the apricot tree microbiota. If we applied our tailocin solution widely and regularly, it might be necessary to track the potential emergence of resistant bacterial strains.
Our next step would then be to make production cheaper by making tailocin production work in Escherichia coli before being able to commercialize the product (see design page of tailocins). Like for the AFP part, we will test the tailocins first in a small field and evaluate the impact of our product on the field and environment before applying it on a large scale.

Ultimately, we could test if the AFP and tailocin solutions can be sprayed in conjunction to see if the freezing-related issues can be further prevented.

To be able to launch our products, we would first and foremost seek the support of our university’s entrepreneurship hub (HUB) that provides coaching and guidance to projects in life sciences. We would also apply to the university’s funding (InnoTREK) to be able to create a start-up. With this grant, we should be able to evaluate our products before deciding the feasibility of bringing them to market.

Figure 1 | The mechanism with which a tractor could spray our solution on crops.

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