Difference between revisions of "GMO Containment in the Field"

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Genetically modified organisms ([https://en.wikipedia.org/wiki/Genetically_modified_organism GMO]) are regulated in each country. Therefore, each country must resolve the issue of taking out GMO bioreporters in the field.
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Genetically modified organisms ([https://en.wikipedia.org/wiki/Genetically_modified_organism GMO]) are regulated in each country. Therefore, '''each country''' must resolve the issue of taking out GMO bioreporters in the field.
==Containment==
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<br><br>
 
==Model of Bioreporters in the Field==
 
==Model of Bioreporters in the Field==
In [https://en.wikipedia.org/wiki/Regulation_of_genetically_modified_organisms_in_Switzerland Switzerland], we have obtained [http://biodesign.cc/2014/08/29/field-testing-approved/ permission] A120851-07 from the [http://www.bafu.admin.ch/?lang=en Federal Office of the Environment] to use specific vials with caps with a self-sealing septum to take the bacteria in the field. The following diagram is our current model of working with bioreporters to analyse water arsenic levels in the field.
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In [https://en.wikipedia.org/wiki/Regulation_of_genetically_modified_organisms_in_Switzerland Switzerland], we have [http://biodesign.cc/2014/08/29/field-testing-approved/ obtained permission] A120851-07 from the [http://www.bafu.admin.ch/?lang=en Federal Office of the Environment] to use specific vials with caps with a self-sealing silicone septum to take the bacteria in the field. The following diagram is our current model of working with bioreporters to analyse water arsenic levels in the field.
 
[[File:Model-of-Bioreporter-GMO-in-the-field.png|400px|right|Current model in Switzerland]]
 
[[File:Model-of-Bioreporter-GMO-in-the-field.png|400px|right|Current model in Switzerland]]
 
We worked with EPFL Biosafety Office to present [http://biodesign.cc/2014/05/11/biodesign-trip-to-bern/ our project in Bern] to obtain the GMO license to take the bioreporter in the field.<br><br>
 
We worked with EPFL Biosafety Office to present [http://biodesign.cc/2014/05/11/biodesign-trip-to-bern/ our project in Bern] to obtain the GMO license to take the bioreporter in the field.<br><br>
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* Analysis in the field, or back in the community
 
* Analysis in the field, or back in the community
 
* All GMO wastes are properly disposed of (autoclaved) in the licensed institution
 
* All GMO wastes are properly disposed of (autoclaved) in the licensed institution
The approval was facilitated by the fact that [http://www.ufz.de/arsolux/index.php?en=20706 ArsoLux] in Germany had been approved by the German Federal Office of Consumer Protection and Food Safety (BVL) and the State Ministry of the Environment and Agriculture of Saxony (SMUL). During an inspection by Department 55 (Bio- and Genetic Engineering, Chemicals (SMUL)), the ARSOlux team was able to ensure that the field test follows national standards and regulations. The conducted tests for environmental viability and others by ArsoLux are documented in this [http://www.ufz.de/export/data/411/49462_ARSOlux_Risikobewertung_EN.pdf pdf] from the ArsoLux site.
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The approval was facilitated by the fact that [http://www.ufz.de/arsolux/index.php?en=20706 ArsoLux]'s bioluminescent arsenic bioreporter had been approved by the German Federal Office of Consumer Protection and Food Safety (BVL) and the State Ministry of the Environment and Agriculture of Saxony (SMUL). During an inspection by Department 55 (Bio- and Genetic Engineering, Chemicals (SMUL)), the ARSOlux team was able to ensure that the field test follows national standards and regulations. The conducted tests for environmental viability and others by ArsoLux are documented in this [http://www.ufz.de/export/data/411/49462_ARSOlux_Risikobewertung_EN.pdf pdf] from the ArsoLux site.
 
<br><br>
 
<br><br>
 
Our first trip is documented on the [[Lac de Salanfe and Ottans 2015]] wiki page as well as in our [http://biodesign.cc/2015/08/13/water-collection-trip-in-ottans-lake/ blog].<br><br>
 
Our first trip is documented on the [[Lac de Salanfe and Ottans 2015]] wiki page as well as in our [http://biodesign.cc/2015/08/13/water-collection-trip-in-ottans-lake/ blog].<br><br>
  
==FAQ==
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==Containment==
'''Q: What is the accepted amount of arsenic in drinking water?'''<br>
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In order to deal with this issue of confinement, we use a vial having a silicon septum, thus allowing us to inject a water sample inside the vial containing the bioreporter while keeping the system confined. Indeed, the hole resorbs after the needle is retracted, thus leaving no room for contact between the environment and the GMO; this diagram illustrates this:<br><br>
''A: 10 ppb (µg/L)''<br>
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[[File:Previouskit.png|none|500px]]
The World Health Organization has set 10 ppb (μg/L) for most countries, with the exception of 50 ppb for locations with high amounts of arsenic.
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<br>
Arsenic can be found in groundwater as a result of natural weathering and microbial processes, but also due to industrial and mining contamination.  
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Here’s how this vial actually looks like:<br>
<br><br>
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[[File:Vial.jpeg|none|80px]]<br>
 +
This vial was chosen because of its confinement properties and because it has already been used in another similar project called ARSOlux which was approved by the German authorities. We’ll come back on how we used this as an important argument for the validation of our prototype by the Swiss authorities. <br><br>
  
'''Q: Why do we care about arsenic?'''<br>
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Extra care will be taken to avoid contact between the tip of the syringe and the bioreporter to avoid contamination of the syringe. After use, for safety reasons, the syringe is put in a syringe container like this one: <br>
''A: Chronic exposure can cause cancer.''<br>
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[[File:Syringewastebin.jpeg|none|90px]]<br>
The connection with cancer is now being re-visited, with correlation to skin, bladder, kidney, lung and other cancer risks. The 50 ppb standard represents more than 2000 times the risk for cancer in the lifetime compared to other known carcinogens (benzene, tetrachloroethylene, DDT, PCB). Before cancer, there are visible symptoms on the skin, and alteration in liver and kidney functions.
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Acute symptoms that can lead to death are likely due to exposure to high amounts of inorganic arsenic.
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<br><br>
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'''Q: Why detect arsenic with a bioreporter?'''<br>
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After the measurement of arsenic concentration, the cells in the vial are, at first, partially neutralized with alcohol before being brought back to the lab for destruction by autoclaving or inactivation by a hypochlorite solution.<br><br>
''A: Because assays with bioreporters are inexpensive and easy, and can be used in the field. They do not produce toxic metal waste like chemical field test kits for arsenic detection. ''<br>
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Assays with bioreporters produce an easily detectable signal, such as fluorescence, color, electricity or luminescence. Assays do not produce toxic gases or require expensive reagents or machines (such as the atomic absorption spectroscopy) for the detection to work. Bioreporters for arsenic have a very low detection limit, and both laboratory and field experiments have shown that as little as 1 ppb of Arsenic can be detected in an aqueous sample.
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<br><br>
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'''Q: What is a bioreporter?'''<br>
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Also, all manipulations with the vials containing the GMO are done over a retention tank to avoid any contamination of the environment if the vial would fall and break.<br><br>
''A: A living organism that produces a signal when it detects a target chemical.''<br>
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The term bioreporter mostly refers to an intact living microbial cells, but can also refer to higher organisms (zebrafish, plants, etc.), that have been genetically engineered to produce a detectable "reporter" signal when they have been in contact with a specific target compound.
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<br><br>
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'''Q: What is a Genetically Modified Organism (GMO)?'''<br>
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With all the aforementioned precautions taken, we can say that the use of our system is not considered to present a potential hazard to humans, animals and the environment.<br><br><br>
''A: An organism in which one or more genes have been introduced or modified by means of recombinant DNA technology.''<br>
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A genetically modified organism is an organism whose genetic material has been altered using genetic engineering techniques. It involves the deletion, insertion or mutation of genes in an organism.
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<br><br>
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'''Q: Since when do we do genetic modifications?'''<br>
 
''A: When we started breeding animals and plants for our interests.''<br>
 
Biotechnology has been used by humans since the beginning of time. The first cases date from 12’000 BC when plants or animals were selectively bred to keep only the ones with the specialities in which we were interested (we have always been biohackers). Gene technology for making genetic modifications has been used roughly since the 1980's.
 
<br><br>
 
 
'''Q: What is this antibiotic resistance?'''<br>
 
''A: When an antibiotic no longer works to kill bacteria or inhibit their growth.''<br>
 
It is a form of drug resistance where a specific group of microorganism, usually bacteria, is able to survive after exposure to a specific antibiotic.<br>
 
Some antibiotics are frequently used in the laboratory to select for the maintenance of DNA fragments introduced by genetic engineering. The arsenic bioreporter bacterium carries a gene for kanamycin (an antibiotic) resistance. This gene is present on the same DNA fragment that contains the genes for the arsenic reporter. By adding kamamycin to the growth medium of the cells, we ensure that the reporter construct is maintained. Cells that have lost it are sensitive to kanamycin and will die.
 
<br><br>
 
 
'''Q: Can this genetic material be transferred to other bacteria?'''<br>
 
''A: In principle yes, but in practice not very efficiently.''<br>
 
Bacteria have a variety of ways by which they can exchange genetic material between different species. Therefore, in principle, the reporter DNA fragment with the kanamycin resistance gene could be transferred from the reporter cells to some other bacterium in the environment. For this to happen, the reporter cells would have to escape, to die and to release their DNA. Another bacterium would then have to take this DNA up and stick it in its own genome. This scenario is not impossible, but the likelihood of it to happen depends on many factors.  <br>
 
The major risk here is that the gene for kanamycin resistance would be taken up by another bacterium, and may spread to pathogenic bacteria. These could become resistant to kanamycin. This antibiotic would then become ineffective in killing pathogenic bacteria in clinical settings. On the other hand, kanamycin is not in clinical use, which reduces the negative impact if, accidentally, this scenario would take place. Still, there is every reason to avoid that the reporter bacteria can escape.
 
<br><br>
 
 
'''Q: What if accidentally, there is an accident with a bioreporter assay and the living bacteria are introduced in the environment?'''<br>
 
''A: They will most likely die.''<br>
 
Most environments are not favorable for growth of the Escherichia coli (E. coli )-based bioreporter bacteria. There are many other bacterial species present, and E. coli will have to compete to survive and will not have an advantage over other bacterial species. Therefore it will not survive very long - depending on the studies, in 1~4 weeks and only 1 in 100,000 bacteria survive. For a detailed description, see: The Risk Assessment of Arsolux in this [http://biodesign.cc/2014/05/11/biodesign-trip-to-bern/ article on biodesign.cc]
 
<br><br>
 
 
'''Q: What if we accidentally touch it, drink it, breath it?'''<br>
 
''A: They will most likely die.''<br>
 
E. coli is a bacteria that is actually present in our gut. The bioreporter is based on a laboratory strain K12 used world-wide, chosen for its absence to cause human disease. In addition, the strain has several genetic defects that minimize its growth outside the laboratory. This strain has been tested for survival in humans and animals and cannot survive more than 1 week. <br>
 
If the bacteria are ingested by accident, it can effectively be eliminated by other antibiotics available in the market, (with the exception of kanamycin but this is severely toxic for humans and not in clinical use).
 
<br><br>
 
 
'''Q: Can we find your notification on the BAFU (Federal Office for the Environment, Switzerland) website?'''<br>
 
''A: No.''<br>
 
The list of all notifications is not publicly listed on the BAFU website. We have made a public post that you can read in our [http://biodesign.cc/2014/08/29/field-testing-approved/ article]
 
<br><br>
 
 
'''Q: Is Arsenic the only thing detectable by the bioreporter?'''<br>
 
''A: No.''<br>
 
Antimony can also activate the same bioreporter, and iron and other compounds can inhibit the bioreporter. So proper controls must be made to understand the bioreporter signal.<br>
 
We also have bioreporters for other heavy metals, such as mercury, cadmium or zinc, and for oil-spill related compounds, such as alkanes, solvents and polycyclic aromatic hydrocarbons.
 
<br><br>
 
 
'''Q : In specific conditions, could the bacterium accidentally start dividing and growing its population out of control ?'''<br>
 
''A: Most likely not.''<br>
 
See the answer to “What if accidentally, this bacteria escapes?”. The only reason why escaped bacteria would grow out of control is when they find abundant food sources that are not used by other bacteria. Fortunately, in and on our bodies, and in the environment, this is not the case. Bacteria exist everywhere and, as a consequence, food is limited in all ecosystems. The other bacteria in the microbial community keep invaders in check. If, however, we change conditions in the environment so that we would kill or inhibit all other bacteria, then invaders may get a chance to proliferate. This is thus a good reason not to overuse antibiotics or release them in the environment, as this could create selective conditions for strains that are resistant to such compounds.
 
<br><br>
 
  
'''Q : If your technique proves being so efficient, it will inspire many other similar devices. How to make sure that they’ll all be as safe and carefully prepared as yours?'''<br>
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=====Materials=====
''A: Each design will have to be approved for safety.''<br>
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Here’s a list of the elements you’ll need:<br>
This device is approved with a very specific vial known for containment. If we change any part of the design, the design will have to be approved in Switzerland. This will be the same in other countries. The bacterial strains used to make the bioreporters are standardized strains. Bioreporters can be other organisms, not only bacteria. Most importantly, one of the main ideas of our project is to build a collaboration between the citizen scientist and a local university or hospital.
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* PK100 screw top clear vial 4ml Sigma Aldrich/Fluka: Vials containing the bio reporter in which we will transfer the water sample into.
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::Where to get it: [http://www.sigmaaldrich.com/catalog/product/supelco/cb_000572?lang=en&region Sigma Aldrich product link]
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::Price: CHF 26.88 for a 100 vials
 +
 +
* Assembled screw cap with hole and PTFE/silicone septum: Caps which allows the injection of the water sample into the vial while keeping the content of the vial confined.
 +
::Where to get it: [http://www.sigmaaldrich.com/catalog/product/supelco/27018?lang=en&region Sigma Aldrich product link]
 +
::Price: CHF 92.4 for a 100 caps
 +
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* Safe Syringe: So we can safely inject the water sample into the vial.
 +
::Where to get it: [http://www.bd.com/hypodermic/products/safetyinjection/ BD product link]
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::Price: Varies depending on the model chosen; from 30 $ to 80 $ for a 100 syringes.
 +
 +
* Syringe Container; E-safe 0.5 l Universal A: To dispose of used needles.
 +
::Where to find it: You can find several online. We’d like to thank Dr. Sabrina Leuenberger for supplying one for free.
 +
::Price: ~ CHF 15
 +
 +
* Retention Tank: Over which we do the manipulations.
 +
::Where to get it: It’s just a plastic box, look around you.
 +
::Price: Not much
 +
 +
_______________________________________<br>
 +
::Total price: ~ CHF 180 for a 100 samples<br><br>
 +
Let’s note that this amount can be reduced by choosing cheaper elements. Their might also be a way to reuse some of these elements if they are properly cleaned from all GMO after usage.
 
<br><br>
 
<br><br>
 +
==More Information==
 +
=====From our project=====
 +
* We are looking for ways to remove the syringe in this process - look at our efforts in the [[Double Cap Design]]
 +
[[Category:Ongoing Projects]]

Latest revision as of 22:09, 27 April 2016

Genetically modified organisms (GMO) are regulated in each country. Therefore, each country must resolve the issue of taking out GMO bioreporters in the field.

Model of Bioreporters in the Field

In Switzerland, we have obtained permission A120851-07 from the Federal Office of the Environment to use specific vials with caps with a self-sealing silicone septum to take the bacteria in the field. The following diagram is our current model of working with bioreporters to analyse water arsenic levels in the field.

Current model in Switzerland

We worked with EPFL Biosafety Office to present our project in Bern to obtain the GMO license to take the bioreporter in the field.

Our process was this:

  • Work with a (federally) certified institution (biosafety level 1) to construct the bioreporter
  • Work with the appropriate federal office to obtain a license to work with the biorerporters in the field
  • In the laboratory, prepare the bioreporters in the approved vials
  • Take the vials out in the field, introduce samples into the vials with the GMO bioreporters
  • Analysis in the field, or back in the community
  • All GMO wastes are properly disposed of (autoclaved) in the licensed institution

The approval was facilitated by the fact that ArsoLux's bioluminescent arsenic bioreporter had been approved by the German Federal Office of Consumer Protection and Food Safety (BVL) and the State Ministry of the Environment and Agriculture of Saxony (SMUL). During an inspection by Department 55 (Bio- and Genetic Engineering, Chemicals (SMUL)), the ARSOlux team was able to ensure that the field test follows national standards and regulations. The conducted tests for environmental viability and others by ArsoLux are documented in this pdf from the ArsoLux site.

Our first trip is documented on the Lac de Salanfe and Ottans 2015 wiki page as well as in our blog.

Containment

In order to deal with this issue of confinement, we use a vial having a silicon septum, thus allowing us to inject a water sample inside the vial containing the bioreporter while keeping the system confined. Indeed, the hole resorbs after the needle is retracted, thus leaving no room for contact between the environment and the GMO; this diagram illustrates this:

Previouskit.png


Here’s how this vial actually looks like:

Vial.jpeg

This vial was chosen because of its confinement properties and because it has already been used in another similar project called ARSOlux which was approved by the German authorities. We’ll come back on how we used this as an important argument for the validation of our prototype by the Swiss authorities.

Extra care will be taken to avoid contact between the tip of the syringe and the bioreporter to avoid contamination of the syringe. After use, for safety reasons, the syringe is put in a syringe container like this one:

Syringewastebin.jpeg

After the measurement of arsenic concentration, the cells in the vial are, at first, partially neutralized with alcohol before being brought back to the lab for destruction by autoclaving or inactivation by a hypochlorite solution.

Also, all manipulations with the vials containing the GMO are done over a retention tank to avoid any contamination of the environment if the vial would fall and break.

With all the aforementioned precautions taken, we can say that the use of our system is not considered to present a potential hazard to humans, animals and the environment.



Materials

Here’s a list of the elements you’ll need:


  • PK100 screw top clear vial 4ml Sigma Aldrich/Fluka: Vials containing the bio reporter in which we will transfer the water sample into.
Where to get it: Sigma Aldrich product link
Price: CHF 26.88 for a 100 vials

  • Assembled screw cap with hole and PTFE/silicone septum: Caps which allows the injection of the water sample into the vial while keeping the content of the vial confined.
Where to get it: Sigma Aldrich product link
Price: CHF 92.4 for a 100 caps

  • Safe Syringe: So we can safely inject the water sample into the vial.
Where to get it: BD product link
Price: Varies depending on the model chosen; from 30 $ to 80 $ for a 100 syringes.

  • Syringe Container; E-safe 0.5 l Universal A: To dispose of used needles.
Where to find it: You can find several online. We’d like to thank Dr. Sabrina Leuenberger for supplying one for free.
Price: ~ CHF 15

  • Retention Tank: Over which we do the manipulations.
Where to get it: It’s just a plastic box, look around you.
Price: Not much


 _______________________________________

Total price: ~ CHF 180 for a 100 samples

Let’s note that this amount can be reduced by choosing cheaper elements. Their might also be a way to reuse some of these elements if they are properly cleaned from all GMO after usage.

More Information

From our project
  • We are looking for ways to remove the syringe in this process - look at our efforts in the Double Cap Design