Team:Toulouse/Practices
Practices
Ethics
Genetically Modified Organisms (GMOs) are at the heart of the current debate focusing on their use. Synthetic biology, an emerging science that only now reaches the public, is a field that sets off strong reactions. Used to questions, comments such as “researchers play God by modifying living beings”, and the prevailing societal distrust, we naturally developed a reflexion about the limits to give to our project.
General Considerations
General Public:
Formerly, the field of biology matched its greek definition: bios meaning life and logos, study, biology was indeed the study of life and living beings. This has changed in the recent years since biology today does not describe life, but rather rebuilds it.
This evolution induced a certain fear in societies marked by an historical creationism springing from monotheistic religions. In these societies, living beings are held to be untouchable, it is thus necessary to comprehensively explain what synthetic biology is, what our project is, as well as what experiments we are conducting in the lab in order to give people who have no background in this field the key elements to start a discussion. Indeed, since most people have a vague or no idea of what synthetic biology covers, they tend to be negatively prejudiced towards the unknown. As scientists, we have to take these fears into consideration and address them as best we can.
But this must be accompanied by a questioning of Science itself. Indeed, Science cannot assume that the regulation of its researches is the lot of scientists only, rather it is its duty to communicate the results of the researches and to have transparency on the experiments conducted. Scientists should be able to acknowledge that they don’t know something rather than lying to the public. Indeed it is often easier to trust someone when the doubts are clearly stated than when everything is allegedly under control but it cannot be verified.
Life:
Before we can have any kind of ethical reflexion concerning our project, we asked ourselves how to best define our “work material”, meaning living beings. Indeed, to be able to estimate the possible consequences of our actions, we first have to characterize what we are modifying.
A living being can be defined as a thermodynamic open system that receives and passes information, and is in this way a sort of “anomaly” of nature since it does not tend towards entropy. In fact it seems that normality is illustrated by non living things, completely subjected to the laws of physics and chemistry, and divisible in smaller parts. If they abided by these laws, living beings should thus evolve towards more entropy and only degrade complex molecules in simpler ones. However, anabolism enables organisms to synthesize complex molecules from simpler ones.
As was said previously, living beings are sacred in our society because they still hold some part of mystery. Christianity having been predominant historically in Western Europe, it still has an influence on people’s minds and a notion that could be assimilated to sin still clings to the modification of organisms.
We can then ask ourselves the following questions: is it possible to have control over organisms, and if yes, to which extent?
It actually seems that we do not have the power to control them entirely, we can only act on a few metabolic pathways, a few functions, at the same time.
iGEM:
When discussing the different concepts and approaches of synthetic biology, we have to take into account that ethical reflexions depend on the core values of a society and thus differ from one to the other.
First of all, since we are participating in iGEM, we reflected on its acronym: international Genetically Engineered Machine. Using the word “machine” appears to us as a mechanistic reductionism. A machine is the assembly of known and controllable components, and this term seems to indicate that we assimilate a living being to a completely characterized system responding to physical and chemical laws. But a machine such as a plane can be built from start to finish whereas a living being like a bacterium cannot. Thus it appears that to use the word machine is somewhat misleading, because biology is much more complicated than that.
For the competition, we have to build genetic constructs called Biobricks. The name Biobrick calls to mind the Lego game and this parallel further asks the question of synthetic biology being conceived as an assembly game.
Hence we asked ourselves the extent to which we could consider our experiments commonplace or ordinary.
Human:
Our reflexion on these ethical issues led us to realize that the reluctance towards the modification of living organisms could be linked to an anthropocentric vision of the world. Indeed, as long as human actions do not have an impact on man himself, they do not cause much reaction.
However, it has been shown that bacteria are extremely numerous in nature and that they are even a part of human beings (digestive system…). The relationship between humans and environmental bacteria is close, and even more so with the bacteria in our body. Indeed a symbiosis between man and specific bacteria is essential to its survival, and we can wonder what the impact of the modification of bacteria interacting (closely or not) with human beings will be.
Anthropocentrism, which places human beings above everything and particularly above nature, is the utilitarian vision. Naturalists on the contrary believe that nature takes precedence over humans and must be protected from their utilization. For them, humans are not above nature, they are at a level with non-human elements.
Law:
Ethics is in close relationship with law, especially when it comes to modifying living organisms.
If we focus on the human being, according to the law, his body does not belong to him, he only has a right of use. This means that he cannot sell, rent or modify his body.
On a closer scale, mammals, and for instance domestic animals like cats, are less obvious but still recognized legal entities. They are granted sensibility, have a status of slave, their master is held responsible for their actions, and the law protects them to a certain extent.
Finally, some organisms are qualified as “res nullus”. This includes for instance bees, bacteria and all other micro-organisms. Nevertheless the loss of a bacterial colony does not affect us emotionally in the same way the loss of a swarm does. Invisible organisms hold less emotional importance in our eyes.
Connection to nature:
The link we keep with nature is complex and ambivalent. On the one hand, we consider nature as fragile and in need of protection whereas on the other hand we see it as strong enough to provide all the resources we need to survive.
In the previous decades or centuries, our vision of nature was more pragmatic and focused on the fulfilling of our needs. Today we tend to “mummify” it in order to preserve it. This change in attitude brought along a defiance towards everything that aims at modifying nature.
Modifying living organisms:
Numerous products synthesized through the modification of microorganisms (reprogrammed bacteria…) are currently of great significance. For example they can be used to synthesize drugs, or produce biofuels…
Modifications can appear on organisms without the intervention of human beings. Microorganisms actually possess diverse means enabling them to evolve in order to adapt to new environments. Among these, we can name horizontal transfer, a phenomenon that happens between bacteria from different lines, from the same species or not. This way, the bacteria that receive the genes develop new characteristics such as resistance to an antibiotic.
These mutations appearing without the intervention of man are accepted by society because they are considered as natural. Synthetic biology reproduces this mechanism while reducing the time normally required for it to appear.
Can we put a patent on life?
Another ethical issue that was raised by the concept of modifying living organisms was that of intellectual property. If patenting an organism were possible, it would lead us back to the perception of life as a machine, or a chemical factory, with no sense of wonder or admiration. GMOs are new creations in that a genetic modification gives a new strain, but this strain has been obtained from organisms coming from nature. Thus it does not seem moral to reduce life to a trading object.
However, patents on living beings do not patent the organism in itself, rather a specific technique or modification. Therefore the patent does not grant a property right on the biological material.
Does having the power to act on DNA and thus to modify the characteristics of a microorganism change our relationship to nature? Working with living organisms induces all kinds of emotions, be it wonder, curiosity or even fear. Contrary to what one could think, feeling fear as a scientist is essential and should not be overlooked since this is what makes us be more cautious and take a step back to study the implications of our work.
Ethical considerations about our project:
Bees have an essential role as pollinators, that’s why their preservation is a major social and environmental concern.
One might ask himself if the importance we give to protecting bees does not have its roots in anthropocentrism since bees produce honey, wax and propolis, products we use. Other pollinators that do not supply us with interesting goods, such as wasps, are less considered.
Furthermore bees are the first pollinating species in terms of fruits and vegetables produced thanks to them thus the advantages of protecting them are obvious for global trade.
Why is our solution better than what already exists?
Being aware of the worrying decrease in bee population and having developed an ethical reflexion since the beginning of this project, we endeavoured to learn more about the causes of this decrease.
It is due primarily to the overuse of pesticides that have proven to act as neurotoxins to bees. However we decided not to act against this threat. Indeed using synthetic biology to solve a problem attributed to a human behaviour did not seem relevant to us. Measures to educate populations to a rational and responsible use of those pesticides would be more appropriate.
Another of the major causes of death for the bees is the parasitic mite Varroa Destructor. The infestation of European bee colonies by this parasite was caused by their contact (wanted or not) with infested asian bees through globalization. Though it is true that the problem first originated with men, today we cannot solve it by changing our behaviour. In this case, synthetic biology could prove to be the solution.
Furthermore we tried to design a system that would be as respectful of the bee’s life cycle and products as possible, hence the circadian regulation and natural molecules.
Finally, we do not aim at completely eradicating the parasite from the environment. In fact, Asian honeybees are capable of identifying the varroa’s presence on themselves and of getting rid of it. Some colonies in Europe seem to start developing behaviours of the same kind. Thus we only wish to slow down the infestation by varroa with our system, to grant honeybees enough time to adapt to the menace and reach a balance with Varroa.
We talked above about the problem of controlling our modified microorganisms, and we put a lot of thought in the biocontainment issue: how could we prevent the dissemination of our E. Coli modified strain? Indeed bacteria are not machines and we cannot control them entirely. It is obvious that we cannot let uncontrolled modified organisms disseminate in the environment. Thus we need to take on a responsible position and accept to not use our system if total biocontainment is not demonstrated, even if positive results are obtained.
Bacteria have a high growth rate and can spread and mutate quickly. This aspect is all the more frightening as bacteria are microscopic and thus the risk is invisible. When the invisible can have an impact on the visible, fear might arise.
On the other hand it is also crucial to make sure that our molecules do not have a negative impact on the bees.
Early on in the project, we thought about how best to address the biocontainment issue regarding our bacterial system. Thus we have decided to put our bacteria in a small plastic bag. This specific plastic called TPX® is microporous and the size of the pores will not allow the bacteria to pass through. We also chose to place this bag in a solid device, adding yet another barrier between the bacteria and the environment.
Safety
Team safety and training
INSA safety training
During the summer, our lab was in the engineering school INSA wich possesses a safety department with one prevention advisor and several prevention assistants assigned to the laboratories. Their goal is to ensure the well being of the employees regarding safety rules and risk prevention. In our laboratory, the LISBP, safety is supervised by Nathalie Doubrovine who was the one instructing us regarding safety procedures.
As new interns we had to take part in a general training session to learn how to identify the risks and prevent
ourselves and our colleagues from harm. We also followed several other trainings that allowed us to use technical apparatus such as autoclave or Nuclear Magnetic Resonance (NMR).
The laboratory safety training requirements of the LISBP are detailed in the Rules and Procedures of the LISBP.
New employees safety training
Every new LISBP employee has to attend this training session regardless of its status (researcher, PhD student, intern...).
The training is divided into two parts. The first one is a training concerning general prevention in research laboratories. This one is taken individually with the NEO software and the explanations of the prevention assistant. It also informs the employee about the emergency numbers.
The second one is a training about the techniques used during our stay concerning microbiological, chemical and incendiary risks.
We then have a test to make sure that we were understood and aware of the previous presentations.
Autoclave
The whole team went through an autoclave training, detailing the explosive/implosive danger surrounding work with an under pressure apparel.
A lab coat, heat resistant gloves and glasses must be worn when manipulating the autoclave.
Liquid nitrogen
We also had a training explaining the risks faced when manipulating liquid nitrogen, and how to manage incidents. Isothermic gloves, glasses and of course a lab coat must be worn when working with it.
NMR
We used the NMR machine kindly made available to us by the MetaToul platform to analyse our culture supernatant. This is a powerful but very expensive equipment, based on the magnetic resonance of the atoms. Due to this, specific rules have to be respected when working with it.
In order to protect the user, access to the NMR is not allowed for people wearing pacemakers, drug pump systems, staples on soft or hard tissue, and pregnant women.
Secondly, in order to protect the technological material, when entering the room the user should not wear a lab coat, nor have anything in his pockets, especially anything made of metal.
Legislation and French Labor Law
INSA Toulouse is a public school for engineers thus the biosafety guidelines are not specific to our institution; the French national regulations about working conditions and the manipulation of genetically modified organisms are applied.
The regulation on workers' protection against risks resulting from their exposure to pathogenic biological agents (Decree No. 94-352 of 4 May 1994) includes microorganisms, cell cultures and human endoparasites which may cause infections, allergies or toxicity.
This Decree is the French transposition of the Directive 90/679 / EEC and is also transcribed in the Labour Code (Articles L4421-1 R4421-1 to R4427-5.)
The Decree of the 16th July 2007 describes the technical preventive measures that are to be set up in research laboratories (including containment), education, analysis, anatomy and surgical pathology, autopsy rooms, and industrial and agricultural facilities where workers are likely to be exposed to biological pathogens.
The rules of health, safety, and preventive medicine applied in public services in France (and thus in all public facilities working in scientific and technological domains) are set out in the Decree No. 82-453. This decree refers to the Labour Code, Public Health Code and Environmental Code.
The Decree No. 2011-1177 is related to the use of genetically modified organisms.
Safety in the lab
Equipment
- A conventional lab coat, closed with long sleeves
- Closed shoes
- Gloves
- Glasses if needed (UV exposure, hot water or chemical handling)
Waste
Different trash containers are available in the lab: |
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One for biological waste (yellow). |
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One for common waste (green or orange). |
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Storage
We have three cupboards, each one dedicated to a different kind of chemical product we use: |
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Flammable products |
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Acids |
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Bases |
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Rules
Our workspace is divided between two rooms. We have a break room where no biological material is brought. In this room, we are able to eat and drink (a fridge, a kettle and a coffeemaker are available) but it is also the space where we hold our meetings and work on our computers. On the contrary, in the lab, we have to wear protective equipment and respect basic rules:
- No smoking (all rooms)
- No drinks or food
- Obligation to wear a closed cotton lab coat
- Oblligation to wear closed shoes
- Long hair must be tied up
- Oral pipetting of any substance is prohibited in any laboratory
- Regular handwashing
- In some cases (UV light, projection risk), obligation to wear protection glasses
Apparatus
Chemical hood
We used the chemical hood when we had to manipulate dangerous and volatile chemicals. For example, all manipulations dealing with formic and butyric acid were done under such a hood.
Water-bathes
The water-bathes were used extensively (transformation, digestion, etc.). However, they can be dangerous because of the exposition to hot or even boiling water. To prevent the burning risk associated with heat, steam and projections, we used special gloves. We made sure to always check that the water-bathes were turned off at the end of the day.
Biological safety cabinet
To manipulate into a sterile area and thus avoid external contamination by unwanted microorganisms we used a Biological safety cabinet (FASTER – Ultrasafe). The bench of the BSC was cleaned with ethanol before and after each manipulation. We also cleaned the BSC completely every two weeks.
Ethidium Bromide
Dark room
We have a dark room dedicated to the use of EtBr and UV. This room is key-closed and everyone entering the room has to wear gloves, glasses and a lab coat. Everything in direct contact with something in this room has to stay there.
Waste
Two specific trash cans are dedicated to the gloves or paper and the contaminated agarose gels.
Safety of our project
We have described above the relevant security measures taken all through the summer to minimize the risks of incident in the lab. But when working in the field of synthetic biology, one of the main concerns is the dissemination of our engineered strain, which could be a threat to the general public and the environment.
First of all, we decided to work with the E. Coli chassis, since its genome is very well characterized and it was well adapted to the molecules we wanted to produce. Of course the strain we chose to work with was non-pathogenic. Moreover the used parts are also non-pathogenic (the acids doses produced are harmless for humans).
The main threat is thus the dissemination of our bacteria in the external environment where its DNA or RNA would be in contact with microorganisms present in the external environment.
To minimize this risk, all our waste is autoclaved before being specifically disposed of and the French regulation does not allow any genetically modified microorganisms to be taken outside the lab. For example, if we would like to test our device on an actual hive, it would be by using the chemicals alone, and not the strain that produces them.
But since regulations are different around the world, and to have a complete overview of our project, we decided to reflect on the biosafety issue, if our device were to be put in place on hives.
We first thought about addressing the containment issue through physical means. In fact, the trap we designed is in itself a first barrier, preventing the engineered bacteria from being in contact with their environment (and particularly the bees). But that was not enough, and the second barrier was developed relying on Groeningen’s team work in 2012.
In fact, we reproduced their idea of packing our bacteria in a small bag made of a special plastic called TPX®, by Mitsui Chemicals. This special material is actually microporous, which means it will let gases such as oxygen or formic and butyric acid pass through while the bacteria stay inside the bag. You can see here the sterility tests we realized. You can learn more about this in the video below.
Apart from this physical containment, we were also thinking of adding to our system an auxotrophy to a specific nutriment not found in the natural environment. In this case, bacteria directly died if they are in contact with the external environment.
Finally, one issue specific to our project is the fact that the small bag containing bacteria would probably have to be manipulated by beekeepers. Indeed our system would be optimized to last at most 2 to 3 weeks, after which the bag would have to be replaced. That would designing a specific training for the beekeepers, and a safe way to get rid of the used bags.
IP rights
Our project took place in a research context. We based the three parts (attract, eradicate and regulation) on published scientific references. At the beginning of our brainstorming about the impact of Varroa destructor on honeybees, we found the patent US 8647615 B1 dealing with methods for attracting Varroa destructor. We decided to send an email to the assignee, The United States Of America, As Represented By The Secretary Of Agriculture via their website. We have not received an answer yet.
The iGEM competition is not all about wet lab and finding sponsors. For our project to really be alive, public contact is paramount. When we first started to reach out to the public we realised that the issue of the decline in bee population was a well-known issue. Still in spite of this, most of the people interviewed weren't aware of the role played by Varroa destructor. This is why we focused a big part of our energy on communicating with the public, in order to raise awareness about a worrying situation.
Discussion with beekeepers
At the very beginning of the project, we were not at all specialists of the bees. That’s why we met with a beekeeper (who was also a microbiologist) to discuss our project and the material restrictions we should submit to.
Our project was followed by the regional newspapers (20 minutes, Métronews, La Dépêche), but also by some national magazines such as Capital or Industrie et Technologies.
This increased our visibility, and thanks to an article we were contacted by the Director of the “Laboratoire Français d’Apidologie” (French Laboratory of Apidology). He offered us his collaboration, and made available his infrastructures, and his teams.
Furthermore, several beekeepers from the forum “La Ruche Warré” heard about the project thanks to the same article. In order to discuss our project with people having a first hand experience of the problem, we joined this forum. We had a very interesting exchange with the beekeepers about our project and synthetic biology in general, explaining about its principle, its interest and the practicability of our solution with the beekeepers.
ExpoSciences Midi-Pyrénées
Our team took part in the regional ExpoSciences in Toulouse. This event brings together scientists of all ages and levels who want to share their knowledge. It’s a good exercise in communication and it enabled us to make synthetic biology better known. Given that DNA is our main work tool, we decided to engage the children by letting them extract DNA from bananas, with ethanol and dishwashing soap.
Recipe for a successful banana DNA extraction!
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This simple yet interesting experience helped us attract children to our stand. It was a great starting point
for us to start some indepth conversation with them about what they knew of DNA and what could be done with it.
Some of them weren't aware that banana had DNA at all and some teachers told us that our demonstration was very well designed and easy to understand without distorting the truth.
The stand having drawn everybody’s attention (thanks to the pungent smell of smashed bananas), our intervention was broadcasted on a local TV (TLT[1]), and we were also selected to take part in International ExpoSciences in Brussels.
Communication in junior high school
To carry on with our education mission, we agreed with one of the science teachers from the Leonard de Vinci junior high school of Tournefeuille to meet with the students. The plan? Bees, Varroas, an introduction to synthetic biology and, of course, banana DNA extraction!
Teaching about varroas and bees
We prepared a short presentation about varroa which was followed by a Q and A session where student were able to interact with us and each other about the disappearance of bees, pesticides, varroa and synthetic biology.
With the help of the teacher we were able to explain the basics of synthetic biology, the idea of biobricks
and spark a quick debate about GMO use. Most of the children were not antagonized to the use of a modified bacterium as long as it was confined in our Trapicoli, but we felt that the biological knowledge needed to fully understand synthetic biology was a little foreign to them, so we switched our lecture to focus
more on bees and the colony collapse disorder.
An unexpected advice
During the Q and A, we were asked about our studies and the kind of diploma we had. One question leading to another, without realising it we where already giving a full guidance lecture on the choices these children would have to make in the following years (which school to attend and why, why study science, etc...). Having a first hand account from students who had to go through the same process not so long ago reassured them and gave them some perspective on their future studies.
We hope that our intervention helped create a few scientific vocations (and even maybe a future iGEMer!)
The main aim of this intervention was to teach children about varroa and synthetic biology while trying to spark an interest inscience in general. But to us it was a time that taught us how to adapt our speech to a young audience. Indeed we had to stop taking for granted notions such as genes and even DNA. The experience we acquired there was put to good use later and made us able to switch from a full science explanation to a simpler overview of our project without getting boring or simplistic.
Giant Jamboree training: International Exposciences in Brussels
Our team had the wonderful opportunity to take part in the International ExpoSciences in Brussels.
During this science fair we had the chance to meet people from all around the world and exchange knowledge and ideas. This happened at the end of July and our project had matured a lot thus the ExpoSciences was the occasion for us to test its welcome in the real world by exchanging with scientists.
We also spent a good deal of our time explaining the iGEM competition to visitors and participants alike, bringing the synthetic biology field, still quite unknown, into light.
This communication experience was fully integrated to the Apicoli project, enhancing its exposure and credibility towards the public. This experience also enabled us to meet his Excellency M. Bernard Valero, the ambassador of France in Belgium, and M. Jean-Claude Guiraudon, CIRASTI President and MILSET Honorary President. Both of them wished us luck in our endeavours.
Finally we took the opportunity of being in Belgium to organize a quick meetup with the KU Leuven iGEM team.
Overall this experience in Brussels was quite intense, we met many people interested in science and it was refreshing to be able to get out of the lab and interact with people about our project. We hope to feel the same degree of engagement this September in Boston!
Exciting perspectives!
During the summer vacations and particularly August, we had trouble organizing any education event. But iGEM does not stop on the 28th of September, and we have everything planned to keep our education project alive after the competition:
- Student's week: between the 9th and the 18th of October in Toulouse. The "Student's week" occuring in several towns in the Midi-Pyrénées region facilitates the integration of new students in Toulouse by allowing them to participate in free events organized by diverse associations. We will be one of the first ever to present a scientific stand!
- Scientilivre: on the 17th and 18th of October. Taking place in Toulouse, Scientilivre ("Science book") is very rightly named and is there to present both books and science to the public. We plan to introduce our project Apicoli through posters and some playful experimentations for families.
