While putting BeeT into use during the competition was out of the question (as per iGEM’s Do Not Release requirements), the fact remains that BeeT was designed as a proof-of-concept for a commercial product. As such, the organism itself needs to be safe to handle and apply by layman beekeepers as well as trained lab workers. In addition, there are major ethical and legal concerns regarding the possibility of any GMO - even one as simple as BeeT - escaping into the natural environment. As it is better to be safe than sorry, we included a dual kill switch in the organism’s design as a biocontainment measure.
BeeT is designed to minimize the risks in both its creation and application. The machine’s chassis is Escherichia coli, a very well-understood level 1 organism. The same risk level applies to the majority of organisms used during development. In the rare occasion that a gene from a Level 2 organism was required - a Serratia gene coding for chitinase, for instance - we opted instead to create it from sequence, rather than using the whole organism. An integral part of BeeT’s premise is a presenting a lower danger/nuisance to bees and humans as compared to more conventional Varroa-exterminating methods. This is the reason we opted for extremely specific toxins.
BeeT’s twin redundant kill switches are its primary biocontainment measures, designed to trigger when the organism leaves its designated environment. In order to carry out its function, BeeT only needs to be active inside the beehive; its presence anywhere else is undesirable. The first kill switch, the Optogenetic TA system, keeps the organism alive as long as it remains within the dark interior of the hive. Being exposed to sunlight over a period of time - which can be inferred as an escape from the hive - will activate the system, killing the bacteria. The second switch, the Auxotrophy-Cas9 system, relies on the inclusion of a synthetic amino acid when BeeT is introduced to the hive. In absence of the amino acid - outside the hive, or when the supply is spent - the system will trigger. As the activated Cas9 targets BeeT’s own DNA, this also helps address the issue of horizontal gene transfer. This design makes BeeT both self-containing and self-disposing.
For more information regarding safety in our project, see our Safety Form.
Maintaining a healthy and contamination-free work environment is a major priority in the field of synthetic biology. Before starting, each member of Team Wageningen UR involved in wet lab work received an introductory tour by the laboratory technician in charge of safety. The tour and subsequent training covered proper clothing, waste disposal, equipment sterility and so forth. The majority of BeeT development was done under Biosafety Level 1 (BSL-1) conditions, with a small amount of work done at BSL-2. For the latter case, specialized training, as well as up-to-date vaccinations, were required.
Wageningen UR’s general safety regulations can be found here.
Though no risk from microorganisms, modified or otherwise, was involved, our work at the Wageningen beehives also required some safety precautions. Working on a project like BeeT becomes much harder when you are covered in bee stings.
Team members working on the beehives always wore at least the top half of a beekeeper’s suit, complete with gloves. Team members were never allowed to go to the beehives alone. Additionally, smoke from a bee smoker was used in order to keep the colony calm when disturbing the hive. (Bees are calmed by exposure to smoke, becoming less easily aggravated.) Even so, getting stung - through the fabric of snug-fitting pants, for instance - still occurred. We were provided with easy access to an epinephrine pen in case of severe allergic reactions, along with clear instructions for its use. For future work, we recommend the use of a full beekeeper’s suit, so as to reduce the amount of bee stings received.