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From 2011.igem.org

Safety

Safety Questions

Question 1:

Would the materials used in your project and/or your final product pose:

a) risks to the safety and health of team members or others in the lab?

Biosafety Level 2 Organisms

It was decided at an early stage of our project that we would work with biofilms. This meant examining a number of organisms which are able to form biofilms, could be made competent and are safe to use. We quickly decided E. coli Nissle 1917 were the safest biofilm-forming organisms to work with. During background research we found that whilst the immediately available lab-strains of E. coli (Top 10 and DS941) were non-pathogenic, they had lost the ability to form biofilms.

We also had access to three strains of the bacterial class Pseudomonas: (P. aeruginosa, P. putida and P. fluorescens). Whilst possessing a well-documented ability to form biofilms, P. aeruginosa is also classed as a level 2 biosafety organism, thus precautions must be taken to prevent percutaneous, ingestion and mucous membrane exposure to clinical materials - as per BSL2 recommendations and Occupational Safety and Health Administration requirements.

This meant that we had to carefully consider the safety of all researchers before we began working with Pseudomonas strains. We consulted a number of guidelines as well as the senior technician for the department regarding working with BSL2 organisms. We also had a meeting with a professor from our Microbiology Institute who advised us on working with Pseudomonas aeruginosa and gave us a brief safety talk. In line with these guidelines, all work with Pseudomonas was carried out:

o after all participants had been shown the proper way of handling and disposing of the organisms and contaminated material
o only under supervision of experienced personnel
o on a part of the building that was not accessible to the public
o wearing spectacles rather than contact lenses (where required).

This was in addition to standard laboratory safety measures such as wearing lab coats and gloves and full decontamination of work surfaces.

To further reduce contact with Pseudomonas aeruginosa, P. putida or E. coli Nissle 1917 (biofilm-forming strain) were used wherever possible.

Latherin

One of the novel biobricks created encodes the protein latherin. The latherin gene originates from horses which have the potential to cause allergic reactions in humans; see question 2 for further discussion.

b) risks to the safety and health of the general public if released by design or accident?

It is important to note that we have been working in a lab with restricted access meaning it cannot be entered by unauthorised personnel. Nevertheless, we carefully considered the possible safety risks to the public in the event of accidental or deliberate release of any aspect of our project.

The organisms we have been working with are lab strains which are unlikely to be able to survive outside of the laboratory environment. We have also been working with E. coli Nissle 1917 cells which are available in probiotic formulation, Mutaflor, in several countries and are therefore not deemed a serious risk.

We do not expect that any of our biobricks would cause harm to the general public if released. This is considered in more detail on our BioBrick Safety page.

c) risks to environmental quality if released by design or accident?

We do not expect that any of our novel biobricks would pose a risk to the environment.

This is considered in more detail on our BioBrick Safety page.

d) risks to security through malicious misuse by individuals, groups or states?

Given the nature of our project, we do not anticipate any way it could be used maliciously.

Question 2:

Please explain your responses (whether yes or no) to these questions. Specifically, are any parts or devices in your project associated with (or known to cause):

- pathogenicity, infectivity, or toxicity?

Yes - as mentioned above, latherin can cause allergic reactions in humans and therefore should be labelled appropriately.

- threats to environmental quality?

No.

- security concerns?

No.

If your response to any of the questions above is yes:

a. explain how you addressed these issues in project design and while conducting laboratory work.

Before working with latherin we asked all participants if they had a known allergy to horses. We then read a report from the Department of Human Services Allergy Working Party to the Minister for Health on Allergens and Anaphylaxis from 2009 and kept contact with the substance to a minimum throughout.

b. describe and document safety, security, health and/or environmental issues as you submit your parts to the Registry.

Safety documentation can be found on our BioBrick Safety page. This contains safety information, DNA sequences and relevant testing data.

Question 3:

Under what biosafety provisions will / do you operate?

a) Does your institution have its own biosafety rules and if so what are they? Provide a link to them online if possible.

The University of Glasgow has its own biosafety guidelines which have been set out by the Safety & Environmental Protection Services.

b) Does your institution have an Institutional Biosafety Committee or equivalent group? If yes, have you discussed your project with them? Describe any concerns or changes that were made based on this review.

We consulted with the university’s Genetically Modified Organism safety officer with regards to our project. The University of Glasgow has a clearly defined policy regarding work with GMOs.

c) Will / did you receive any biosafety and/or lab training before beginning your project? If so, describe this training.

Each of us attended a three day training course prior to the start of our time in the lab designed to introduce us to the essential techniques we would need to work safely in the weeks to come. We were also given handy practical tips, such as safe handling and storage of bacteria, care when using reagents such as SybrSafe and how to work aseptically.

On our first day in the lab we were given an in-depth safety demonstration on how to use the equipment by David Somerville, head of the undergraduate Microbiology Teaching Lab. After setting up the equipment in our lab we had a meeting with our building’s safety officer who discussed our project and the rules of the building we would be working in. We then went on a tour of the building to ensure that everyone knew the location of fire exits, eye wash baths, first aid boxes and where we could find a first aider if so required. We spent the rest of our first day reading and signing the relevant COSHH forms. These were then compiled into a folder which was kept in an accessible place along with a copy of the instructions of all the equipment we would be using, and the data sheets for chemicals.

d) Does your country have national biosafety regulations or guidelines? If so, provide a link to them online if possible.

Yes, the UK has a number of national biosafety regulations and guidelines.

The Genetically Modified Organisms (Contained Use) Regulations (2000) .

Biosafety level 1 & 2 - A table of practical guidelines for working with organisms of biosafety level 1 & 2

WHO Laboratory Biosafety Manual, pp9-19 - A comprehensive list of instructions for working with biosafety level 1 & 2 organisms.

Question 4:

Do you have other ideas on how to deal with safety or security issues that could be useful for future iGEM competitions? How could parts, devices and systems be made even safer through biosafety engineering?

We propose that teams working on light-induced promoters should include the additional safety measure of using a chemically-induced promoter. This would mean that, in the event of the microbe escaping the lab, the likelihood of it meeting the chemical (in the correct concentration and volume) that induces the promoter is extremely low. It would therefore become less likely to be activated unintentionally.

To further the idea of a chemically-activated promoter we have been working with a the pBAD promoter. Not only is this only activated by the sugar arabinose, it is also inactivated by the sugar glucose - a substance which is found abundantly in humans. This has improved the safety of our BioBricks as if we did happen to accidentally ingest any bacteria transformed with our constructs they would be unable to trigger their arabinose-induced gene responses.