Difference between revisions of "Team:Bordeaux/Practices"

Popularization

This year, IGEM Bordeaux took a particular interest in talking with people about synthetic biology and Genetically modified organisms. You'll find in this section a preview of what we did this year

Cap Sciences

Cap sciences is an animations center in Bordeaux open to all which try to help people discover and understand scientific phenomenons, new technologies and their industrial applications. They try to make science accessible to everyone and to engage a large variety of people in scientific activities (most commonly people which are interested by sciences in general). They were glad to hear about our project and we have been in contact with them all throughout the year, presenting what we've done on popularization.

During the first meeting on the 17th of December, we created the group ImagineLife composed by iGEM team members and other people interested by sciences.

We talked about how we can explain synthetic biology to non-scientific people. After a long discussion, we were agreed to do a survey about synthetic biology to find out:

✵ What people already know / don't know about this field

✵ What what scares them

✵ What do they believe that synthetic biology can do for them

With the results of this survey, we hoped to have a better overview of the situation and be able to explain synthetic biology in simple terms to the general public.

So we worked on a survey, and interviewed people in the streets. Emilie, Charlotte, Hiba,Caroline, Gilles and Diego spent a few afternoons in the streets of Bordeaux trying to get in contact with walkers. These conversations thought us that in Bordeaux the main concern about GMO's is that people are unaware about what they really are and the possibilities that synthetic biology offer us. Furthermore it made us start to realize the importance of scientific communication and for the need to think about our project from an ethical and safety point of view. For more information on the questions we asked ourselves please see the Ethics page! Then we have presented our work during the next meeting, on the 20 of February. Below is a link to the presentation of our interviews and the pdf of the questions we asked. We apologize in advance that the video is in french but you can find the subtitles in a PDF format below the link.

Survey questions
Watch our interview! (In French)

After this short presentation, they gave us a lot of advice to improve our survey which we did. We presented a new survey on the 17th of April and obtained around 20 answers during this meeting and 53 answers online following this link: (PDF à mettre) We also discussed during this meeting about the different way we can use to explain Synthetic biology because we still thinking that it was relatively easy to explain it. So, the idea was to do some video, game, and why not an exposition in the Museum Cap Sciences. As, we were already work on a broad game about biology, we decide to try this way. During the next meeting, the 18 of June, we have presented a first version of our board game: Lab’Life. They give us a lot of advice on the working of the game, on the questions and allow us to better balance the game.

Lab Life

After a few years of thinking about it, our game is finally going to be created physically. It took some determination and long hours playing to optimize it but we're proud to present our 2015 Edition!

Presentation of Synthetic Biology in Lycée Montesquieu

IGEM Bordeaux also wanted to raise teenager's awareness of synthetic biology. On 2015, May, 21st, Jean, Savandara and Lily made a presentation of synthetic biology at the school "Lycée Montesquieu" in Bordeaux, showing its interest by giving some interesting applications.

Ethical aspects of synthetic biology

Before analyzing the ethical aspects of synthetic biology, it seems important to clearly define this new area of biology.[1]

Legal framework in France and in Europe

Creating guidelines and a legal framework is essential for safety reasons and to reassure the general public. In Europe, there is no law specifically concerning synthetic biology but synthetic biology is framed by common biology laws. In France, the first laws concerning bioethics were put in place in 1994. [2] Before, general laws existed but did not deal with the particular ethical aspects which come from biology and using living organisms. On the 1st of July, the law n°94-548 presents guidelines for data processing in medical research. This updated the law n°78-17 (1978) relative to informatic and big data processing in all fields of research. Quickly after, on the 29th of July 1994, two laws were put in place: law n° 94-653 and 94-654 which deal with respecting the human body and products which come from the human body. These laws cover the protection of patients which was then introduced in the Civil code, organizational rules in medical sectors and guidelines to be put in place to protect people involved in medical research programs. In march 2002, patient rights are put in place and certain end of life guidelines are discussed. Futile medical care is banned. Ethic committees set up by the European Union also exist to complement these laws for example, the European Commission, the European Group on Ethics in Science and New Technologies, the Inter-service Group on Ethics and EU policies, and the Executive Management for Research and Innovation

Specifically concerning synthetic biology, a European project called SYNBIOSAFE approved by the European Commission has focused on ethics and safety of synthetic biology. In 2012, it published an article concerning the bioethical and biosafety attentions in iGEM competitions. These two types of frameworks help the researcher to answer to many ethical questions in the clearest way possible. Therefore, not only law may limit synthetic biology. Aspects concerning Politics, Economy, and Philosophy could influence the bioethics questions.

Why do we need Ethics commitees? Interview with Mr FAUCHER

On the 29th of July, Savandara and Charlotte had the possibility to meet with Mr Jacques Faucher, doctor, priest and founder of the regional Ethics commitee "l'Espace Bioéthique Aquitain". Mr FAUCHER generously accepted to come to our lab at the European Institute of Chemistry and Biology (IECB) in order to talk to us about the importance of bioethics in the field of synthetic Biology.

When Mr Potter, American oncologist and surgeon, invents the word ethics in the 1970's and writes his book Bioethics: a bridge to the future he pioneers the questions to come asking himsef questions which go beyond medecine such as: does life have a future on this planet? Questions which have lead to the conclusion that not everything that can be done should be done. Since the modification of a plant's genome or of a unicellular organism's genome is tolerated, what about the human genome? In other words, are there barriers to put on living organisms? Should we have the right to experiment on humans in order to widen our knowledge on the human body? In the case of human clinical trials, ethical aspects are judged on the quality of the scientific protocol. It would not be ethical to use humans as tests if researchers are not certain to have exploitable results. Furthermore, the question of consent also needs to be adressed. While we cannot ask an animal or a plant for their consent, ethical commites insist on having a proof of consent in the European Union. This is particularly important since Anglo-Saxon scientific reviews refuse to publish papers if the have not been approved by an ethics commite. We can ask ourselves if the importance researchers give to Bioethics is truly personal or if they do not regard it as only constraint for publication.

When it comes to bacteria however, what point of view do we have? Humans generally have a vision of the world which is anthropocentric, we consider ourselves superior to other organisms. For example, during the Nuremberg trials against Nazi doctors, Carl Brandt (health minister and part of the German military) was sentenced for having lead experiments on humans in concentration camps believing that he was superior to them. When do these acts become wrong? Recently, in China, certain tests are being done on human embryos. Where should we draw the line for embryonic therapy? In Europe, questions of ethics slows down research on such topics but in Asia, research seams to be without limits.

Ethical considerations about our project

All of these ethical considerations made us think about our project Cur'd Vine.
First, it is important to us that our project was related with transparency, so that uninitiated people can apprehend our scientific approach. Our Wiki, in free access on the WEB, allows us to clearly and concisely expose all the thinking steps which governed our Cur’dVine project. It also allows us to share our knowledge and results to as many people as possible. New iGEM teams or other researchers could use our results to continue our project. Furthermore, our implication in annex projects such as Cap Sciences, making street interviews about GMO's or high school presentations about synthetic biology, is a sign that we have constantly tried to democratize synthetic biology to the general public and helped us to consider the profits and risks. Without being paranoiac, we have to think of all potential dangers that our molecule may create.
Will this molecule be able to create new diseases? Can it break the flora equilibrium for example through effects on bees in the region and creating the proliferation of chinese hornets? We can also ask ourselves questions concerning it's effects on humans. Since curdlan is technically a sugar biopolymer, would it increase the sugar level in wine? These are questions which can be asked by the general public and which might cause them to be frightened of our project. This is why we have taken into consideration these questions and tried to answer them in the best way possible which are resumed in our next section which deals with safety.

Literature Cited:

[1] : 5e avis sur la biologie de synthèse. Comité consultatif commun d'éthique pour la recherche agronomique. INRA - CIRAD.(2014)

[2] : La révision des lois de bio-éthique. Vie Publique(2010)

Safety

To avoid confusion about the terms "biosecurity", "biosafety" and "biohazard", some people gave a definition of each of these terms.

✵ Biosafety describes principles, technologies and containment practices put in place to avoid accidents and unintentional exposure to pathogens and toxins.

✵ Biosecurity describes the protection measures put in place and management of important biological materials in laboratories, in order to prevent them from being accessed without authorization.

✵ Biohazard includes, in this case, both biosafety and biosecurity.

Reference : http://www.ambafrance-uk.org/Biosecurite-dans-les-laboratoires

The team who worked in the lab received instructions of lab safety with safety training. We learned how to act in the case of an accident, where to find emergency showers, fire extinguishers and emergency doors. Some basic safety requirements were discussed such as wearing gloves and a lab coat for most procedures. In some cases, during the use of ethidium bromide, nitrile gloves are mandatory. Other experiments must be conducted with safety glasses. It is also recommended to wear clothes offering a minimum of protection. We also learned about working with chemical products and how to dispose of them. Most products used are classified as non-hazardous (even for the sulfation protocol). Chemical products are just irritants and so need to be handled with gloves. In the lab, there is a specific solvent cabinet for liquid wastes. Moreover, all biological waste will be autoclaved before disposal. All work is done on benches or under open-front hoods.

Since we worked with non-toxic organisms as Escherichia coli (BL21) and Saccharomyces cerevisiae (Invsc1), all waste that has been in contact with bacteria was put to bacterial waste in aseptic conditions but no specific security procedures needed to be put in place.

Why is it safer to use Curdlan rather than copper sulfate ?

1. Copper Sufate

For the Environment Even if copper is essential for life at low doses, it is environmentally toxic at higher doses. Not being biodegradable, it spreads through rain water in soils and in river waters. It accumulates on the soils and vineyards are polluted. Its spread is facilitated by water and wind erosion. This element has been used many times to treat mildew which has resulted in it’s accumulation in the Aquitaine region.
This compound also modifies the biosphere since it is toxic for worms and microorganisms living in vineyard soils. Furthermore, copper sulfate does not distinguish harmful and beneficial fungi. It disrupts, for example, the development of mycorrhiza fungi that live in symbiosis with the plant’s root to help the access to nutrients from the soil to the vegetal. The microfungi that decompose the material to fertilize the soil are also in danger.
These effects are poorly seen on an area not exceeding the current authorized limit in organic farming (4 kg of copper per hectare per year, according to AFSSA (French Agency for Food Safety)). However, in terms of conventional agriculture, there is no limit for this substance. The measure reduces the damage to the biosphere but isn’t sufficient: copper sulfate is tolerated but not banished. It is therefore necessary to take more drastic measures by reducing the use of this substance.

For Humans Through the food chain, copper is an essential element for life at low doses. It is found in insects, fish, mammals and all herbivores, carnivores or omnivores. However, it is harmful if swallowed or inhaled, in cases of skin contact, eye contact (hence the compulsory wearing of gloves masks and glasses during our project) and is very toxic to aquatic organisms. We must not underestimate the long-term effects of chronic copper use.
Also, the copper-related risks are increased if added residues of this product from agricultural cultures, other food sources containing copper as tap water. Living near the wine places, like winemakers, are the most exposed.
Indeed, when it accumulates in the body, it is in too large quantities, it would affect the removal system of the beta-amyloid protein in the brain. This is harmful to the nervous system and is implicated in Alzheimer's disease. The protein, in excess in patients, is a peptide which is mainly aggregates (plaques) characteristics of the disease, and is present in soluble form in healthy patients.
In humans, it is known that copper occurs naturally and in normal amounts. It can protect neurons against oxidation. However, several studies have shown that copper plays a key role in neurodegenerative diseases like Alzheimer's. It is also increasingly referred to as an "environmental illness". Indeed, when it is accumulated in the body in large quantities, it is supposed to affect the removal system of the beta-amyloid protein in the brain. This is harmful to the nervous system and is implicated in Alzheimer's disease. The protein, in excess in patients, is a peptide which is mainly aggregates in the case of disease, and is present in soluble form in healthy patients.

2. Curdlan

Not dangerous for Humans Our project aims to use a polysaccharide which is synthetically produced by our genetically modified organisms. This molecule already exists in nature, it is naturally present in the yeast’s wall. No bad impact on the environment has been identified. Compared to copper sulfate which is a toxic compound, this molecule seems to be more environmentally friendly.
In 2013, the “Savoy ADAbio” tested on vine plants the effect of very low doses of different sugars in order to replace the copper treatment in the fight against pests. For example, they tested the effect of fructose against mildew. After different doses were tested, an encouraging result emerged : the weekly fructose application of 100 g / ha had a similar effect than a treatment of 600 g / ha of copper. According to this testimony, we now know that sugar helps fight against mildew.
Also used in many different applications, Curdlan doesn’t seems to have bad effects (see Other Effects Curdlan). However, we also collaborated with the Oxford iGEM team to test the toxicity of this molecule on the bacteria that already live in nature. This experience is useful to know the impact of Curdlan on other organisms. Another experience would have been interesting : a toxicity test on plant cells or animal cells. Unfortunately, we didn’t have the capacity to carry this test out in our laboratory. For now, we are still waiting for the results.
In food industry, the additive E424 (Curdlan) is used as a thickener, stabilizer, gelling and firming agent. This is reassuring because it is a common product in the food chain, and not reported as toxic.

We are not using the entire cell Another important theme is the spread of new organisms in nature. Indeed, this could alter biodiversity. If we want to directly use our synthetic organism on the plant, we must integrate death genes in their genomes to create organisms that are not be able to live in natural conditions. In the case of our project, we are not planning on directly applying our genetically engineered bacteria on the plant but only our purified molecule. In other words, we chose to spray on the vine only the cell product: Curdlan. In our project, the cell can’t evolve and in any manner alter the environment.

Effect on the plant

Economy

A bit of French Touch

The production of Bordeaux is 6.4 million hectoliters with only 14% for white wines. More than 850 million bottles are sold every year and worth almost € 3.7 billion. Today, the red wines represent 86% of total volumes produced by the vineyards of Bordeaux, 89% of exports in volume and 92% in value. The Bordeaux vineyard now covers 123,000 hectares for 270,000 hectares of farmland. The climate and the terroir is particulary favourable for a exceptionnal vineyard. Indeed, soils and Girondins basements are rather shallow. The Gironde, the Garonne and the Dordogne make the climate cool and provide the water needed by the vineyard. On the left side of the Gironde and Garonne, there is gravelly, sandy, clay soil made by the erosion of the Pyrenees. On the other side, soils are likely limestone wich is suitable for vineyard. The warm ocean current of the Gulf Stream accentuates the temperate climate and the Landes forest forms a protective shield against the winds of the ocean.

One of the caracteristics of the bordeaux vineyard is the great diversity of the grape variety, we can find lots of grapes :
Reds: Merlot, Cabernet Sauvignon, Cabernet Franc, Malbec, Petit Verdot.
Whites: Sauvignon, Semillon, Muscadelle, Ugni Blanc, Colombard.

Some mildew key figures

15 is , on average, the number of treatments that had to make a winemaker in 2012 to fight against mildew.
11 ° C is the minimum temperature required for initiation of primary infections in the spring.
€ 246 per ha is the average investment of a winemaker in France in 2012 for his fight against mildew.
4 days this is the optimal time necessary to complete mildew its entire cycle from contamination by spore until the following sporulation.
1878 is the year when the mildew , Plasmopara viticola , was observed for the first time in France.

Treatments against mildew have a cost that varies between € 50,000 and € 164,000 per year.Over the last 20 years, no less than 3 mildew epidemics were recorded in France . These outbreaks have a direct impact on the production of wine. As shown in the graph below, represents the number of hL (millions) produced over time , production in 1998 , 2001 and 2006 dropped by over 50 % compared to previous years. You can find on the vinopôle website the evolution of mildiou risk every week !

In 2010, each hectar or grapevine had received around 16 chemical treatments (this number was 15 in 2006) and varies greatly between regions. Amongst the biological threats on grapevine, mushroom parasites are far ahead of insects and fungi (12 fungicides are used against 2 insecticides and 2 weedkillers). Over 95% of the fongicide treatments in 2010 were due to mildew and oidium, sicknesses that are favorized by rain, humidity and heat. They bring the global quality of the wine down by reducing it's composition in phenols and sugars. The risc of contamination on the leaves begin during the period where the floral buds and futur grapes are growing and lasts until the moment when the grapes touch each other. The sensibility period between oidium and mildew have common momments and vinyards don't heasitate to mix anti-mildew and anti-oidium solutions making the concentrations of these chemicals twice more concentrated in the grounds.

Downy mildew requires optimum conditions to reproduce and infect. A warm, moist, and humid environment is required. Studies in Sicily have shown optimum time for oospore germination is between the end of February and the middle of March. With this understanding, if fungicides are used just before these conditions occur, and have proven to be efficient measures. Other methods include proper watering, and a good location where the plant can receive continual sunlight.

Reference : www.oenologie.fr / www.vins-bordeaux-negoce.com / www.vignevin.com / www.vinopôle.com