Team:Calgary/Implementation



OVERVIEW


From the lab to the real world

A technology like Oviita would be meaningless without the means and plans to make it effective in reality. Dr. Warren Wilson cautioned us that humanitarian projects often fail due to a lack of understanding of whatis needed and what will be accepted by the end users. Therefore, we have consulted with stakeholders and community members to find out how Oviita could best be put into action some day. This includes the varying materials, resources, and skills available to build the Field Adapted Bioreactors and cultivate the yeast, and ensuring that it is both safe and acceptable to consumers as a food ingredient.


COMMUNITY BIOREACTOR

Creating an inexpensive and simple bioreactor for the community

In order to provide a sustainable, community-based solution, we plan to genetically modify Rhodosporidium toruloides, an oleaginous yeast that naturally produces beta-carotene and lipids, to be more robust and resource-efficient. By modifying the yeast to produce cellulase, it can then use common agricultural waste products as an energy source for synthesizing its oil. It can then be ingested as a vitamin A supplement. The yeast strain, while naturally safe and non-pathogenic, will also be genetically modified to include a kill switch for bio-containment, and optimized for oil production.


FOOD SAFETY AND COOKBOOK

Food Safety

Yarrowia lipolytica has been evaluated and approved for safe consumption as an edible nutritional yeast by a variety of sources. It was approved by the European Food Safety Authority in 2019 for consumption of whole-cell biomass after being heat-killed. Their findings concluded that there were no nutritional or toxicological concerns for human dietary use, and granted permission to produce Yarrowia cell mass as a safe edible supplement.  Additionally, Y. lipolytica is already a common component of dairy and meat products, particularly sausages. It is part of the desirable flora of common cheeses including Gouda, Picante, Apulian, Camembert, blue-veined, Cheddar, Brie, and Feta (Zinjarde, 2013), where these yeast contribute to the overall flavour. 

A study by Groenewald et. al (2014) of potential pathogenicity of Y lipolytica concluded that the yeast could be considered on an equivalent level of non-pathogenicity as S. cerevisiae, with only a few rare examples of opportunistic infection in severely immunocompromised individuals. It is generally regarded as non-pathogenic (Zinjarde, 2013), is common and ubiquitous in the environment (EPA, 2019), and has been granted Generally Recognized As Safe (GRAS) status by the U.S. Food and Drug Administration (USDA) (Zieniuk 2019).

Additionally, studies on recombinant beta-carotene produced by Y. lipolytica found that it did not produce effects any different from commercial beta-carotene, and shared a similar safety profile regardless of the different source. (Grenfell-Lee et al., 2013)

Finally, thymol is also regarded as a safe food ingredient. It is classified by the FDA as a food for human consumption as well as a food additive and flavouring ingredient, and possesses GRAS designation (EPA, 1993). Naturally occurring in aromatic plants especially thyme, it is widely consumed by humans. A review of its status by the EPA in 2009 concluded that thymol is a normal part of the diet and has minimal toxic effects in mammals, and is therefore an acceptable food additive.

Incorporation into local diets

In order for our project to successfully deliver its Vitamin A and anthelmintics to individuals, it must be integrated into existing food practices by people who are willing to adopt it. An important aspect of that will be taste. Although we cannot know exactly what Oviita will taste like, we can make educated guesses based on other related species of nutritional yeast and fungi. It will most likely have a savoury, cheesy, or nutty flavour, rich in umami. The thymol will additionally provide an aromatic, thyme-like taste. As such, we predict Oviita yeast will impart a pleasant flavour when added to savoury dishes such as rice or curries. Provided below are just a few examples of how Oviita might be incorporated into local meals in South Asia and Sub-Saharan Africa. Roti is included from the suggestion of our HP contact Dharamwati, who mentioned it is a dietary staple in her area.

INGREDIENTS

 

  • 4-5 cups mixed vegetables, cut into chunky slivers about 1 1/2 inches long (I used a sweet potato, two large gold potatoes, two carrots, and a cup of green beans)
  • 1/2 tsp turmeric
  • 1 13.5 oz can coconut milk (not the light kind)
  • 1 tsp cumin seeds
  • 1 tsp bengal gram dal (can sub with 1 tsp raw rice)
  • 2 green chili peppers like jalapeno (deseed and use more or less depending on how hot the pepper is)
  • 1-2 tbsp coconut oil (more oil means more flavor)
  • 2 sprigs (about 25-30) curry leaves
  • 1 tsp black or brown mustard seeds
  • 1-inch knob ginger (cut into thin slivers)
  • 1/4 cup yogurt
  • 1 tsp compressed fresh Oviita nutritional yeast
  • Salt to taste
  • Coriander and fresh coconut, if you have some, for garnish

 

INSTRUCTIONS

 

Place all the cut veggies into a large pot with 1 cup of water and turmeric. 

Place the coconut milk in the blender with the cumin, green chili peppers, nutritional yeast, and bengal gram dal or rice. Blend into a paste. Set aside.

When the veggies are cooked -- it should take no more than 15-20 minutes -- add the coconut paste to the veggies and stir well to mix. Let the curry come to a gentle boil over medium-low heat.

Turn off the heat and add the yogurt or lime juice. Stir to mix but be careful not to mash up the veggies because they'll be very tender at this stage.

Heat the coconut oil. Add mustard seeds and when they sputter, add the curry leaves and the ginger. Saute for a minute or two until the fragrance permeates the house.

Pour the tempering over the avial and stir to mix.  Garnish, if you wish, with coriander and some fresh coconut.

Serve hot with rice.

 

Recipe by Vaishali,  taken from Holy Cow Vegan

INGREDIENTS

 

  • 2 Tbsp vegetable oil (or Ethiopian niter kibbeh)
  • 1½ cups medium yellow onion, finely chopped (1/2 medium-sized onion)
  • 2 garlic cloves, finely chopped
  • 1½ Tbsp berbere (A mix of garlic, ginger, chili, and other spices important in Ethiopian cuisine)
  • ½  tsp dried Oviita yeast powder
  • 2 Tbsp water
  • 2 Roma tomatoes, diced
  • 1 lb beef (eye of round roast), cut into ½ inch cubes
  • 1 tsp iodine salt
  • 2 jalapeños, deseeded and sliced
  • 3 sprigs fresh rosemary

 

INSTRUCTIONS

 

In a nonstick skillet, heat oil over high heat. When the oil easily slides across the pan, add the onions and garlic. Cook for about 4 minutes until the onions become translucent. Stir often so the garlic doesn’t burn.

Add tomatoes and cook for another 3 minutes, stirring occasionally.

Sprinkle with berbere and 2 tablespoons of water reduce the heat to medium-high and cook for 5 more minutes, stirring often.

Add beef and mix well until the meat cubes are well coated with gravy. Reduce the heat to medium and cook for 15-20 minutes or until the meat is well cooked.If the gravy is too thick you can add another tablespoon of water.

Add salt, jalapenos, and rosemary. Give it a last stir and let it cook for 3 more minutes. Taste and adjust seasoning. If the gravy isn't spicy enough you can add more berbere.

Serve beef tibs on injera bread with a side of Misir Wot (Ethiopian lentils) and gomen (Ethiopian collard greens)

 

Recipe by Hanna and Aurel Pop, taken from Gourmet Cubicle

INGREDIENTS 



  • 6 large tomatoes
  • 4 large onions
  • 6 cloves of pressed garlic
  • 2 chillies (optional)
  • 2 Tbsp tomato paste
  • 2 tsp fresh wet Oviita yeast
  • vegetable oil
  • 500 g of beef, chicken or lamb (alt: mixed vegetables)
  • 800 g long grain rice
  • 1.5 litres of water or stock (± 1 stock cube, depending on how much meat is used)
  • 1 teaspoon each of ground white and black pepper

 

INSTRUCTIONS

 

Put tomatoes into the blender and set aside. Then, in a non-stick pot, fry pre-cooked meat in oil. When these have browned, remove and set aside.

 

In a nonstick skillet, heat oil over high heat. Add onions and fry until soft before adding garlic and blended tomatoes. Add meat stock and/or stock cube(s), tomato paste, ground white and black pepper and stir. Remember to season strongly because rice will later be added to this sauce.

 

Cook for 10 minutes on medium heat before adding rice and nutritional yeast. Stir and mix well before covering. Cook on low heat for apprroximately 20 minutes. Add vegetables and mix well. Add about 1 cup more of water and continue to cook on low heat until rice is done.

 

Recipe by Cynthia Prah, taken from African Food Map

INGREDIENTS 

 

  • 2 cups whole wheat flour (Atta)
  • 1 tsp dried Oviita yeast
  • 1 cup boiling water
  • pinch of salt 
  • melted butter for brushing the rotis

 

INSTRUCTIONS

 

Mix dried nutritional yeast into flour. Pour the cup of boiling water into the flour, and mix in using a fork or spoon. You will have a wet, lumpy mixture.

The mixture will have cooled down a little. Knead into a soft dough. The dough will feel a little sticky, but as you knead the dough will become silky smooth.

Heat a non-stick pan on the stovetop on medium heat. 

Divide dough into 8 pieces. Roll each piece into a neat round ball. Flatten the dough with the palm of your hands, and roll on a lightly floured surface. You can add more flour as you are rolling to prevent the dough from sticking. Roll out into a circle, about 20cm in diameter

Place the roti on the pan. Cook until it starts to bubble a little, then flip it over. Cook until the other side bubbles. Brush with butter and turn over again.Brush the other side with butter and turn over. You will be flipping your roti over 3 times. It takes about a minute or less to cook the roti.

To keep rotis soft, place them in a container with paper towel at the bottom and more on top of the rotis.

 

Recipe by Lorraine, taken from Tamarind Thyme

FUTURE DIRECTIONS


Preparation


Unfortunately, we will need money to get started. Now that we have officially registered as a non-profit, we can begin applying for as many grants as possible. Once we become a registered charity, we can develop and utilize our crowdfunding website to start fundraising for operations. Before anything can happen on the ground, though, we need to collect data. This means sifting through survey programs, contacting government agencies and contacting other organizations. If these data for an area are not available, we can provide community health workers with our Randle Cell Testing Device to gather the necessary data. These days, data is sometimes worth more than gold, and this data will determine where Oviita should go and all of the factors that could affect our product’s success. From the data, we as an organization will sit down and develop a carefully crafted concept note that we can use to attract donors.

With data in hand, and as suggested by Robert Gough from Western Heads East, a similar organization that now serves probiotic yogurt to 160 000 people, it would be immensely beneficial first to contact a local university in a flagship location. From there, we could then create a network of community partners. Given all three of our major partners, CAWST, WHE, and West Nile Youth Empowerment Center (a local community partner), have operations in Uganda, and Uganda has a high vitamin A deficiency rate, Uganda would be a rational choice.

The appeal of partnering with a university is the fact that they are usually fully developed information centers that can provide essential resources like equipment for culturing and storing our yeast or conducting collaborative research. Considering how much our project has to gain from research, such as bioreactor designs, social enterprise development, community impact, or even economic impact, we intend to make it a big part of our organization. To ensure that we are conducting this research ethically, we need to conduct this research with locals and not remotely.

While we are forging relationships with the local community, this would be the time to start reaching out to regulatory bodies. That means contacting the local Ministry of Health, The National Bureau Of Standards, University boards, ethics committees and at the same time, going through the same process here in Canada. We will need to prove our product is safe, and as Robert Gough mentioned, public perception will be a major obstacle.

First Move


At this point, we’ve established a relationship with a handful of community partners (like West Nile Youth Empowerment Center), community health workers and a university research team. We’ve made our way through the regulatory bodies, we have constructed a new concept note, now we start developing a pilot program.

This pilot program will act as the momentum, so it’s important it’s done properly. Our partner, Kelly James from CAWST, really stressed the importance of developing an effective community education strategy. Fortunately, CAWST has an abundance of training materials for both organizations and local people. Capitalizing on those resources, we develop a handful of training programs while also looking at other training programs in the community that we might be able to collaborate with, such as those put on by community health workers or even our partners. Our programs might include general health and nutrition, how to use the bioreactor or how to run a microenterprise. This would be a good way to get feedback on our public perception as well.

We can really kick things up a notch on our crowdfunding platform at this point as well. We will need to fund those small things like storage containers or office supplies. These are things that our platform works best with. In addition, we will also be able to launch social media campaigns the increase our traffic.

At this point, we will more than likely be in contact with local women’s rights organizations and will reach out to them for assistance in determining who will run our pilot program. If possible, we would like to reach out to Yogurt Mamas, an extremely successful women’s group of self-sustaining female enterprises, so that they could act as mentors for Oviita. Because our mission is to encourage and support others, we will hire a local director of operations to ensure things run smoothly. While our pilot program is running, we will be conduct studies with our university partners so that we can start making plans for the next stage.

Full Implementation


Upon a successful pilot program, we should now have evidence to help influence our public perception in new locations as well as a good amount of data on a successful approach. Moving to a new location will follow a similar sequence of steps as the first, but with a little more speed. However, the biggest difference is that now that we are a little more established, we can start curating programs and resources from all our partners to specifically fit that community’s needs. If a vitamin A deficient-community is in an isolated area and doesn’t have access to electricity, we can collaborate with partners to provides solar panels. If a community doesn’t have access to clean drinking water, we can collaborate with a partner like CAWST to allow for access to clean water. We hope that we can ensure a successful community influence by taking as many factors into account as possible. Any plans past this point would be pure speculation, but our mission will remain the same: to provide local communities with support in their vision of implementation of Oviita.


REFERENCES

Environmental Protection Agency (EPA). (1993, September). US EPA - Pesticides - Fact Sheet for Thymol. United States of America EPA. https://www3.epa.gov/pesticides/chem_search/reg_actions/reregistration/fs_PC-080402_1-Sep-93.pdf

Environmental Protection Agency (EPA). (2009). Thymol; Exemption from the Requirement of a Tolerance (74 FR 12613, pp. 12613-12617, Rep.). United States of America EPA. doi:https://www.federalregister.gov/documents/2009/03/25/E9-6262/thymol-exemption-from-the-requirement-of-a-tolerance

FSA NDA Panel (EFSA Panel on Nutrition, Novel Foods and Food Allergens), Turck, D, Castenmiller, J, de Henauw, S, Hirsch‐Ernst, K‐I, Kearney, J, Maciuk, A, Mangelsdorf, I, McArdle, HJ, Naska, A, Pelaez, C, Pentieva, K, Siani, A, Thies, F, Tsabouri, S, Vinceti, M, Cubadda, F, Engel, K‐H, Frenzel, T, Heinonen, M, Marchelli, R, Neuhäuser‐Berthold, M, Pöting, A, Poulsen, M, Sanz, Y, Schlatter, JR, van Loveren, H, Ackerl, R and Knutsen, HK. (2019). Scientific Opinion on the safety of Yarrowia lipolytica yeast biomass as a novel food pursuant to Regulation (EU) 2015/2283. EFSA Journal 2019;17(2):5594, 12 pp. https://doi.org/10.2903/j.efsa.2019.5594

Grenfell-Lee, D., Zeller, S., Renato, C., & Pucaj, K. (2014). The safety of β-carotene from Yarrowia lipolytica. Food and Chemical Toxicology, 65, 1-11. doi:https://doi.org/10.1016/j.fct.2013.12.010

Zieniuk, B., Fabiszewska, A. Yarrowia lipolytica: a beneficious yeast in biotechnology as a rare opportunistic fungal pathogen: a minireview. World J Microbiol Biotechnol 35, 10 (2019). https://doi.org/10.1007/s11274-018-2583-8

Zinjarde, S. S. (2014). Food-related applications of Yarrowia lipolytica. Food Chemistry, 152, 1-10. doi:https://doi.org/10.1016/j.foodchem.2013.11.117