Team:SDU-Denmark/safety-b

From 2010.igem.org

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(Risk-assessment for Individual Parts)
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<p style="text-align: justify;">In electing a project there were numerous ideas to be considered. They were divided into the following categories: the environment, foods, health & disease, physics/chemistry/biochemistry and finally other which included some more artistic ideas along with a few humorous ones (like the jeopardy bacteria – knows all the right questions).  Especially while thinking about medical ideas for implementing in the body we knew there were serious risk issues that had to be considered because of the many ways bacteria may interact with the human body.</p>
<p style="text-align: justify;">In electing a project there were numerous ideas to be considered. They were divided into the following categories: the environment, foods, health & disease, physics/chemistry/biochemistry and finally other which included some more artistic ideas along with a few humorous ones (like the jeopardy bacteria – knows all the right questions).  Especially while thinking about medical ideas for implementing in the body we knew there were serious risk issues that had to be considered because of the many ways bacteria may interact with the human body.</p>
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<p style="text-align: justify;">During our closer investigations of project ideas we also considered which bacteria it was possible to use. Our focus was to try and find a project that was possible to carry out by using E. coli. The reasons here for being that (cultivated strains of) E. coli are very well adapted to the laboratory environment since they are easy to keep alive, they can be fairly easy modified and unlike some wild strains of E. coli they no longer have the ability to thrive inside the intestines. Despite of these considerations we started out working on a project we called mE.chanic (because we wanted to make bacteria do mechanical work), and the idea was to have a culture of bacteria contract and relax, thereby making a pump-like movement creating mechanical work.</p>
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<p style="text-align: justify;">During our closer investigations of project ideas, we also considered which bacteria it was possible to use. Our focus was to try and find a project that was possible to carry out by using ''E. coli''. The reasons are that (cultivated strains of) ''E. coli'' are very well adapted to the laboratory environment since they are easy to keep alive, they can be fairly easy modified and unlike some wild strains of ''E. coli'' they no longer have the ability to thrive inside the intestines. Despite of these considerations we started out working on a project we called mE.chanic (because we wanted to make bacteria do mechanical work), and the idea was to have a culture of bacteria contract and relax, thereby making a pump-like movement creating mechanical work.</p>
[[Image:SDU-Denmark-2010-pili.png|600px|thumb|left|Our first project idea involved pili, but we decided it was unsafe, because of it's potential pathogenicity.]]
[[Image:SDU-Denmark-2010-pili.png|600px|thumb|left|Our first project idea involved pili, but we decided it was unsafe, because of it's potential pathogenicity.]]
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<p style="text-align: justify;">Now, we found out that we could use pili as some sort of ‘grappling hooks’ to make a connection between the bacteria. Pili from E.coli had been measured to have a pulling force of about 100 pN (enough to work a nano-machine), and so it seemed that we would have a good chance of making usable mechanical work if we continued this idea. We just needed to find out how to control the formation and retraction of the pili.</p>
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<p style="text-align: justify;">Now, we found out that we could use pili as some sort of ‘grappling hooks’ to make a connection between the bacteria. Pili from ''E.coli'' had been measured to have a pulling force of about 100 pN (enough to work a nano-machine), and so it seemed that we would have a good chance of making usable mechanical work if we continued this idea. We just needed to find out how to control the formation and retraction of the pili.</p>
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<p style="text-align: justify;">The project turned out to be most likely to succeed if we used the pathogenic bacteria Pseudomonas Aeruginosa, because it is very good at making the type VI pili we wanted. But, that is unfortunately due to its pathogenicity since the pili are an important part of this, as they are used for the bacteria to stay put and not get washed away. So using P. Aeruginosa was immediately out of the question because the risk of personal health (fx. getting a cystitis infection) for the researchers was too high, since they are students and not yet fully educated scientists. Also it would be a potential danger for the surrounding community if it spread. So instead we started to investigate the possibilities of using E. coli, but then found that [bacterial name] was a non-pathogenic bacteria with close resemblance to P. Aeruginosa, and also with resemblance to E.coli. So we started working with it. Eventually though, we had to discard this project idea on a safety basis: We had serious doubts that we could succeed with P. Aeruginosa, given that we basically didn’t believe the bacteria would function with all the genes we needed to provide it with (pili are virulence factors, and working with a non-pathogenic bacteria it would be less likely to make enough pili / keep its pili).</p>
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<p style="text-align: justify;">The project turned out to be most likely to succeed if we used the pathogenic bacterium ''Pseudomonas aeruginosa'', because it is very good at making the type VI pili we wanted. But, that is unfortunately due to its pathogenicity since the pili are an important part of this, as they are used for the bacteria to stay put and not get washed away. So using ''P. aeruginosa'' was immediately out of the question because the risk of personal health (fx. getting a cystitis infection) for the researchers was too high, since they are students and not yet fully educated scientists. Also it would be a potential danger for the surrounding community if it spread. So instead we started to investigate the possibilities of using ''E. coli'', but then found that ''Pseudomonas putida'' was a non-pathogenic bacteria with close resemblance to ''P. aeruginosa'', and also with resemblance to ''E. coli''. So we started working with ''P. putida''. Eventually though, we had to discard this project idea on a safety basis: We had serious doubts that we could succeed with ''P. putida'', given that we basically didn’t believe the bacteria would function with all the genes we needed to provide it with (pili are virulence factors, and working with a non-pathogenic bacteria it would be less likely to make enough pili / keep its pili).</p>
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<p style="text-align: justify;">Then our thought was to use E. coli and provide it with pili, but this started a long discussion about whether this would make the E. coli potentially pathogenic. The end of the discussion was to give our project a big make-over keeping the concept of wanting to make mechanical work, but changing the idea of the actual mechanical work to be done.</p>
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<p style="text-align: justify;">Then our thought was to use ''E. coli'' and provide it with pili, but this started a long discussion about whether this would make the ''E. coli'' potentially pathogenic. The end of the discussion was to give our project a big make-over, keeping the concept of wanting to make mechanical work but changing the type of the actual mechanical work to be done.</p>
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<p style="text-align: justify;">We finally decided on the project of Bacterial Micro Flow, and the safety issues in relation to this project will be presented in the following. These include issues of researchers’ safety, public safety and environmental safety. We will also look at safety in relation to the specific biobricks we use and make, and will have a chapter on what the safety-staff at our university think of this project. Now, let us start at the lab.</p>
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<p style="text-align: justify;">We finally decided on the Flow-E project, and the safety issues in relation to this project will be presented in the following. These include issues of researchers’ safety, public safety and environmental safety. We will also look at safety in relation to the specific BioBricks we use and make, and will have a chapter on what the safety-staff at our university think of this project. Now, let us start at the lab.</p>
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==Risk-assessment for Individual Parts==
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==Risk-assessment for individual parts==
Here we consider the safety of each of our parts.
Here we consider the safety of each of our parts.
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===Methode===
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===Method===
<p style="text-align: justify;">We have used our [http://2010.igem.org/Team:SDU-Denmark/safety-d#Procedure risk-assessment guidelines] to describe the safety of each of our contributed parts. The genes were [http://blast.ncbi.nlm.nih.gov/Blast.cgi BLASTed] to find their function and known homologs. This function was then reasearched (mainly via [http://www.ncbi.nlm.nih.gov/PubMed PubMed]), described and considered according to the questions asked in the risk-assessment paper.</p>
<p style="text-align: justify;">We have used our [http://2010.igem.org/Team:SDU-Denmark/safety-d#Procedure risk-assessment guidelines] to describe the safety of each of our contributed parts. The genes were [http://blast.ncbi.nlm.nih.gov/Blast.cgi BLASTed] to find their function and known homologs. This function was then reasearched (mainly via [http://www.ncbi.nlm.nih.gov/PubMed PubMed]), described and considered according to the questions asked in the risk-assessment paper.</p>
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*Vitamin A and its derivatives have been implicated as chemopreventive and differentiating agents in a variety of cancers ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 3]). </p>
*Vitamin A and its derivatives have been implicated as chemopreventive and differentiating agents in a variety of cancers ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 3]). </p>
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<p style="text-align: justify;">These effects are not directly associated with the enzyme itself, but have been observed in humans. It is highly unlikely that high enough doses can be reached with this biobrick. Please see references for more information about the diseases. </p>
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<p style="text-align: justify;">These effects are not directly associated with the enzyme itself, but have been observed in humans. It is highly unlikely that high enough doses can be reached with this BioBrick. Please see references for more information about the diseases. </p>
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<p style="text-align: justify;">The biobrick has many homologs, that have the same function as this biobrick and is highly conserved in bacteria and eukaryotes. The biobrick does not affect the immunesystem in humans.</p>
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<p style="text-align: justify;">The BioBrick has many homologs that have the same function as this BioBrick and is highly conserved in bacteria and eukaryotes. The BioBrick does not affect the immune system in humans.</p>
====Environmental impact====
====Environmental impact====
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<p style="text-align: justify;">To our knowledge retinal should not play a significant role in environmental processes or would disrupt natural occurring symbiosis. </p>
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<p style="text-align: justify;">To our knowledge, retinal should not play a significant role in environmental processes or would disrupt natural occurring symbiosis. </p>
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<p style="text-align: justify;">The biobrick should not increase its host’s ability to spread, survive outside the laboratory, and will most likely decrease its ability to replicate.</p>
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<p style="text-align: justify;">The BioBrick should not increase its host’s ability to spread, survive outside the laboratory, and will most likely decrease its ability to replicate.</p>
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<p style="text-align: justify;">Beta-caroten monooxygenase is found in a wide variety of different bacteria, insects and animals ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 4]). As such we would be cautious as to letting a system containing this BioBrick into the wild, since it's function might conflict with existing systems. On the other hand one might argue that since it's function is already available in nature, the function is widely available. </p>
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<p style="text-align: justify;">Beta-carotene monooxygenase is found in a wide variety of different bacteria, insects and animals ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 4]). As such we would be cautious as to letting a system containing this BioBrick into the wild, since it's function might conflict with existing systems. On the other hand, one might argue that since its function is already available in nature, the function is widely available. </p>
<p style="text-align: justify;">The product of the BioBrick, retinal, also plays an important function in nature and animals. For this reason we fell that the BioBrick could be used in controlled settings, but not in the wild. </p>
<p style="text-align: justify;">The product of the BioBrick, retinal, also plays an important function in nature and animals. For this reason we fell that the BioBrick could be used in controlled settings, but not in the wild. </p>
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====Possible malign use====
====Possible malign use====
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<p style="text-align: justify;">There is not reason to believe this biobrick could be used for malign uses; it does not increase the hosts ability to vaporize, create spores, regulate the immunesystem or should be pathogenic. </p>
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<p style="text-align: justify;">There is not reason to believe this BioBrick could be used for malign uses; it does not increase the hosts ability to vaporize, create spores, regulate the immunesystem or should be pathogenic. </p>
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====General use====
====General use====
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<p style="text-align: justify;">This BioBrick poses no treat to the welfare of people working with it, as long as this is done in at least a level 1 safety lab by trained people and in non-pathogenic hosts such as E. coli TOP10 or MG1655. No special care is needed when working with this BioBrick. </p>
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<p style="text-align: justify;">This BioBrick poses no treat to the welfare of people working with it, as long as this is done in at least a level 1 safety lab by trained people and in non-pathogenic hosts such as ''E. coli'' TOP10 or MG1655. No special care is needed when working with this BioBrick. </p>
====Potential pathogenicity====
====Potential pathogenicity====
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<p style="text-align: justify;">This biobrick increases the potential of its host to move. Increased motility has been associated the bacteria’s ability to invade humans ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 5]); on the other hand it has also been shown that bacteria that loss the function of the FlhDC operon, are considerable better at colonizing the intestine ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 6]), and so an increased expression might decrease a hosts ability to colonize and invade humans. It is impossible to ensure, that this plasmid is not transferred to pathogenic bacteria since the FlhDC operons is used in a wide array of other bacteria that are known to be pathogenic.</p>
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<p style="text-align: justify;">This BioBrick increases the potential of its host to move. Increased motility has been associated to the bacteria’s ability to invade humans ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 5]); on the other hand it has also been shown that bacteria that lose the function of the ''FlhDC'' operon, are considerably better at colonizing the intestine ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 6]), and so an increased expression might decrease a host's ability to colonize and invade humans. It is impossible to ensure, that this plasmid is not transferred to pathogenic bacteria since the ''FlhDC'' operons is used in a wide array of other bacteria that are known to be pathogenic.</p>
<p style="text-align: justify;">A number of effector cells in the human immune system react specifically to bacteria’s flagella, and so a hyperflagellated host will most likely induce a stronger immune response. </p>
<p style="text-align: justify;">A number of effector cells in the human immune system react specifically to bacteria’s flagella, and so a hyperflagellated host will most likely induce a stronger immune response. </p>
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====Environmental impact====
====Environmental impact====
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<p style="text-align: justify;">The biobrick might increase its host ability in finding foods, but we do not think it will be able to outmatch naturally occurring bacteria. It will not increase the hosts ability to replicate, but will increase its ability to spread, which might increase its ability to survive. </p>
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<p style="text-align: justify;">The BioBrick might increase its host's ability to find foods, but we do not think it will be able to outmatch naturally occurring bacteria. It will not increase the host's ability to replicate, but will increase its ability to spread, which might increase its ability to survive. </p>
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<p style="text-align: justify;">The biobrick itself will most likely not make an environmental impact, since it only regulates internal systems of the bacteria.</p>
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<p style="text-align: justify;">The BioBrick itself will most likely not make an environmental impact, since it only regulates internal systems of the bacteria.</p>
====Possible malign use====
====Possible malign use====
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====Potential pathogenicity====
====Potential pathogenicity====
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<p style="text-align: justify;">This BioBrick consists of three different parts: The first 224 amino acid residues come from the NpSopII gene from halobacteria, encoding a blue-light photon receptor with 15 residues removed at the C-terminal. The following 9 amino acids are a linker. The last part is HtrII fused with Tsr from E. Coli. The complex' first 125 amino acid residues come from HtrII and the remaining 279 from Tsr ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 7]). NpHtrII is thought to function in signal transduction and activation of microbial signalling cascades ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 8]).  </p>
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<p style="text-align: justify;">This BioBrick consists of three different parts: The first 224 amino acid residues come from the ''NpSopII'' gene from ''Natronomonas pharaonis'', encoding a blue-light photon receptor with 15 residues removed at the C-terminal. The following 9 amino acids are a linker. The last part is ''HtrII'' fused with ''Tar'' from ''E. coli''. The complex' first 125 amino acid residues come from ''HtrII'' and the remaining 279 from ''Tar'' ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 7]). ''NpHtrII'' is thought to function in signal transduction and activation of microbial signalling cascades ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 8]).  </p>
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<p style="text-align: justify;">A single article has been written about haloarchaea in humans indicating that these played a role in patients with inflammatory bowl disease ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 9]), but there is no evidence that the genes or near homologs, this BioBrick is made from, are involved in any disease process, toxic products or invasion properties. They do not regulate the immune system in any way.</p>
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<p style="text-align: justify;">A single article has been written about haloarchaea in humans indicating that these played a role in patients with inflammatory bowel disease ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 9]), but there is no evidence that the genes this BioBrick is made from or any near homologs are involved in any disease processes, toxic products or invasion properties. They do not regulate the immune system in any way.</p>
====Environmental impact====
====Environmental impact====
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====Possible malign use====
====Possible malign use====
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<p style="text-align: justify;">This BioBrick will not increase it’s hosts ability to survive in storage conditions, to be arosoled, to be vaporized or create spores. None of its proteins regulate or affect the immune system or are pathogenic towards humans and animals.</p>
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<p style="text-align: justify;">This BioBrick will not increase its hosts ability to survive in storage conditions, to be aerosoled, to be vaporized or create spores. None of its proteins regulate or affect the immune system or are pathogenic towards humans and animals.</p>

Revision as of 21:12, 26 October 2010