Team:Exeter/Human Practices/lande

Environmental Considerations BEFORE AND DURING the Project

Since the beginning of our iGEM project, we have carefully considered environmental issues. Our frequent discussions with Dr. David Santillo highlighted a number of interventions we persisted with before laboratory work commenced to avoid environmental issues if a release event of our genetically modified E.coli occurred. We have used non-mobilisable plasmids in the pSB series to prevent transfer of genes and of antibiotic resistance by horizontal gene transfer and/or conjugation. Use of laboratory strains One Shot ^® TOP10 E.coli and E.coli BL21(DE3) for cloning and expression of our glycosyltransferase genes respectively will prevent the issue of replication in the environment and host infection, meaning both survivability and potential hazards of our genetically engineered E.coli in the environment would be unlikely. The production of bespoke polysaccharides are unlikely to be a fitness advantage to our genetically engineered E.coli and the expression of glycosyltransferase proteins in such a prokaryotic system will not be detrimental for natural flora and fauna.

David made aware of a potential issue that when E.coli produces our carbohydrate polymers, there is a possibility that foaming may occur. Foaming is an issue because it can alter liquid flow and may slow down or even impede bacterial growth. Recently, a French team were able to produce human milk oligosaccharides in large-scale by metabolically engineered E.coli, which were generated both intracellularly and extracellularly (Priem et al, 2002). The extracellular deposits of oligosaccharide were predominantly found in the foam, and this is beneficial to our project in two ways. Firstly, it would allow us to remove the foam layer and retrieve our bespoke polysaccharides much more easily than other known methods of extraction such as high-cell-density cultivation. Secondly, the removal of foam would not require anti-foaming chemicals such as silicone oils which may contaminate or even reduce bacterial growth. David also brought to our attention that if our polysaccharides were accidently released, pollution to the environment would be inevitable. However, we are developing our technology in a biosafety level 2 laboratory even though the organism and product we are producing are considered biosafety level 1 containment measures, so release of our polysaccharides into the environment is unlikely. Even if pollution was still possible, we are generating less polluting waste products using biological synthesis of polysaccharides than synthetic chemistry methods (Biological vs. Chemical Synthesis, Impact and the Future).

Environmental Considerations BEYOND the Project

Our meeting with Dr. Janet Cotter continued to raise awareness of the potential impact of our bespoke polysaccharides on the environment, reiterating that the biggest impact on ecosystems would be new sources of nutrition. Polysaccharides, or sugars, are frequently degraded to simple sugars or mono and disaccharides, such as glucose or sucrose respectively. These carbon sources are subsequently used for respiratory metabolism and are critical sources of nutrition. If there was an accidental polysaccharide release event, there is a potential for new sources of food and could lead to explosions in food webs. In fact, for certain species with appropriate enzymes to digest these polysaccharides, a new selection pressure will be applied against those that do not possess such appropriate digestion enzymes. This will dramatically affect biodiversity.

As a result of our discussion with Janet, we propose bell-jar experiments in a contained system within the laboratory to determine what the effects the digestibility of our bespoke polysaccharides could be on soil biodiversity. We will introduce common saprophytic fungi (such as members of the Rhizopus and Mucor families) and bacteria (such as actinomycetes) capable of breaking down polysaccharides into soil and then to determine such affects (if any) by tracing possible degradation products of our carbohydrate polymers. After these experiments, we will identify risks associated with an accidental polysaccharide release event and determine if there are any hazards which require a higher level of containment. This will enforce stricter containment measures and decrease the probability of such a release event occurring.

The phenomenon referred to as the “Mucilage of the Northern Adriatic” is an example of what a polysaccharide release event may have on the environment. This phenomenon that takes place in the sea has been occurring over the last three centuries and is undoubtedly caused by a release of polysaccharides. Originally thought to be extruded by phosphorus limiting algae, others are contesting that cyano- and heterobacteria are capable of producing ectohydrolytic enzymes which generate long-lived polysaccharides. This release event causes scum to form on the ocean surface and its extent is so huge that it can be seen by satellite. This process has been extensively studied because of the potential significance this phenomenon has on global carbon cycles, especially as it is now thought the recently frequent events ensuing is due to climate change. Whilst this process occurs in sea water and not on land, accidental polysaccharide release events clearly have a marked effect and we have considered necessary precautions in such an event occurring on land seriously.

What is the impact to people and businesses?

Priem, B. et al. (2002) A new fermentation process allows large-scale production of human milk oligosaccharides by metabolically engineered bacteria, Glycobiology. 12: 235-240.

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