Safety

Overview

Safety in a laboratory setting is critical. This includes the safety of our product as well as safe practices and protocols in our laboratory which are outlined in the general lab safety. There are specific adjustments made during the progression of Cellucoat along with certain guidelines which are followed in our in-project designs. In the midst of the iGEM competition, COVID-19 still plays a significant role and respective safety measures need to be acknowledged.

In Project Design

The safety of all materials used for packaging foods is controlled under Division 23 of the Canadian Food and Drugs Act and Regulations (1). Specifically, section B.23.001 of which prohibits the sale of foods in packages that may impart any substance to the contents which may be harmful to the consumer of the food (1).

Cellucoat is made from lab-grade bacteria. The E. coli BL21 strains used are non-pathogenic, and K. xylinus is used to ferment kombucha and individuals have been known to eat the SCOBY (symbiotic culture of bacteria and yeast) for its probiotic benefits. However, having bacteria in contact with food products when there is a risk of causing harm to individuals must be addressed during the post-processing stages of creating Cellucoat. There are three ways that Cellucoat’s manufacturing process is designed to ensure that the material is not contaminated with microorganisms and it meets the requirements outlined in Division 23 of the Canadian Food and Drugs Act and Regulations:

Figure 1. Outline of how safety is practiced in iGEM Calgary's Lab

On the note of improving the safety of Cellucoat as a food packaging material, another future direction would be to genetically engineer a strain of E. coli that does not produce endotoxins. Since a part of the post-processing stages of preparing BC for commercial use is sterilizing through putting the material in the autoclave to lyse the E. coli cells and release the antimicrobial peptide, endotoxins will also be released. Since endotoxins are thermostable, even at 121 degrees in the autoclave, it can still pose dangers if consumed (2).

The solution is to use E. coli strains that are able to grow and express recombinant proteins with the endotoxin precursor lipid IVA as the only type of endotoxin (2). Since lipid IVA does not trigger an endotoxin response in humans, it will not pose an issue to the safety of the material when the cells are lysed (2).

General Lab Safety

The iGEM Lab at the University of Calgary is qualified as a biosafety level 2 lab. During the time of our project we worked with bacteria from groups 1 such as K. xylinus, B. subtilis, as well as E. coli BL21. These bacteria were dealt on lab benches disinfected with 70% ethanol and handled aseptically under a bunsen burner flame. This aseptic method is used as a substitute for biosafety cabinets. Furthermore, all plates not in current use were parafilmed, wrapped, and placed into the fridge to minimize contamination and unnecessary handling to to promote the health and well-being of lab members.

A portion of the project was involved in working with spore-forming fungi. However, our lab is not equipped to work with spore-forming fungi and to minimize the amount of spores present in the lab, the lab benches were disinfected with 1% bleach. The University of Alberta was another local team who’s lab was equipped to experiment with these strains of fungi. There we had a partnership with them where they would determine the activity of nisin against fungi and inturn we would help them with docking. The fungi and a sample of the antimicrobial peptide nisin was transported to the University of Alberta iGEM team by following the iGEM Safety & Security Committee recommended guidelines. The fungi plates were parafilmed and put into a Styrofoam box which was placed in another enclosed box. This was placed in the back of the car and transported to Edmonton, where members of the University of Alberta iGEM team received our samples and began experimentation.

Lastly, all members of the team went through lab safety training courses. This included both dry lab and wet lab members. These lab safety courses were required prior to members having lab access, so these training courses were completed throughout April and May. A lab dress code was implemented to ensure that all members in the lab wore the proper PPE such as lab coats, closed toed shoes, gloves, long hair tied back, and ankle-length pants.

Covid-19 Safety

COVID-19 remains a global threat, and it is the collective responsibility of individuals to remain vigilant and reduce the spread. This means masks were a mandatory measure in our classroom and laboratory areas. There were some members of the team who indeed did test positive for COVID-19 and they responsibly isolated until tested negative. Outside of the lab, numerous meetings were done through zoom to protect ourselves and others. During the fall semester, our lab hours were 8am to 4pm with only those of us coming in as necessary for lab experiments only.

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References

1. Health Canada. Food Packaging Regulations [Internet]. Packaging Materials. Government of Canada; 2020 [cited 2022 Oct 6]. Available from: https://www.canada.ca/en/health-canada/services/food-nutrition/food-safety/packaging-materials.html
2. Mamat U, Wilke K, Bramhill D, Schromm AB, Lindner B, Kohl TA, et al. Detoxifying escherichia coli for endotoxin-free production of recombinant proteins. Microbial Cell Factories. 2015;14(1).