Team:Paris Bettencourt/Biosafety
Biosafety and Biosecurity
Biosafety
When creating a product destined to the world using synthetic biology, a big concern is biosafety. Which is why we aim to contain Genetically Modified Organisms in a hardware device to prevent them from escaping at all costs.
The product is produced by minicells, non-living organisms which are not considered to be GMOs. Moreover, minicells lyse after a short period of time and they lack the ability to grow/divide/proliferate So, the enzyme produced at the end of the process -even in presence of minicells- will be usable as a non-biological product, free from whole minicells.
The process is automated in a machine to minimize the risks linked to personnel. Minicells are the only biological structure the user can encounter - with the final product- thus limiting the risk of GMO escape.
The process requires separation of the E. coli from the minicells. We developed a novel technique combining phages and filtering for efficient and low-risk separation. The redundancy created by using two different techniques considerably reduces the risk of GMO escape.
Why is biosafety important ?
We designed our system to be versatile, developing it to extend its use to many applications, such as the production of Taq polymerase or other enzymes. But, by expanding the range of uses, we need to take into consideration that toxins or pathogenic microorganisms could now potentially be involved in our device. Thus, by placing biosafety at the center of our project, we aim to better identify and anticipate the problems caused by future uses.
Additionally, the prospective aim of implementing our project to local scales raises another issue. Access to high security and safety labs could indeed be very complicated for communities leading to disparities in terms of accessibility and equity. Therefore, the biosafety of our project is essential to ensure equal accessibility even with limited resources.
Risk Assessment
A risk assessment is necessary to protect the health of staff and users, the environment and the product. It is performed in 4 steps:
Situation definition and risk identification
Risk characterization
Risk hierarchization
Risk mitigation
All equipment and disposables should be ISO certified. The machine will be certified and follow regulations. The risks linked to the machine malfunction are to be assessed separately with a fully functional prototype. We identified no risks linked to defective disposables.
The following risk assessment is performed for:
All the potentially hazardous chemicals and live organisms used during a cycle
All steps involving personnel intervention
It should be reevaluated each time a change is made in the process: an incident which could have had major consequences occurs or when a small incident occurs multiple times. It should be reviewed once a year otherwise.
* The terms risk, risk source, event, consequence(s), control and likelihood are defined according to the norm NF ISO 31000about Risk Management.
** The norm NF ISO 31000 defines risk as the effect of uncertainty on objectives.
*** The risk assessment was performed following norms NF ISO and the recommendations from INRS (National Institute for Research and Security, France) on bioreactors in document ed6258.
**** Chemical risk was assessed through the comparison of at least two Safety Data Sheets.
The consequences linked to malicious intent are limited thanks to the use of unharmful substances (chemical and biological) and the small scale of production. This was confirmed by both Professor Escande and Safety Engineer Lacome (interview section). However, to limit the risk, CCTVs and an alarm system are installed within the laboratory and close to the machine.
Bio-Risk management culture
After performing our risk assessment, we realized that many risks are linked to personnel and therefore, to human behavior which cannot be assessed for and controlled easily. Bio-Risk Management Culture (BRMC) is a concept which tries to answer this problem. BRMC is defined as an assembly of beliefs, attitudes, and patterns of behavior of individuals and organizations that can support, complement or enhance operating procedures, rules, and practices as well as professional standards and ethics designed to prevent the loss, theft, misuse, and diversion of biological agents, related materials, technology or equipment, and the unintentional or intentional exposure to (or release from biocontainment of) biological agents (1) . In the future, we aim to follow the guidelines of BRMC to increase biosecurity and limit the risks linked to human behaviour.
Feedbacks from professionals
We sought professionals with strong expertise in risk assessment to give us their external perspective.
We first had an interview with Jean Escande, a professor from the University of Technology of Compiègne who specialized in risk management. He gracefully accepted to guide us through the different steps of the procedure to ensure we did not forget or overlook any important information. Here are the main key points and conclusions from this interview:
If the consequences of a risk are negligible, there is no need to pay much attention to it
If the gravity of an event is important, even if the risk is unlikely to happen, special attention should be paid to it
Because the hardware is not destined to be commercialized, has a very small-scale production and is based on an unharmful process, there are no particular precautions to take for its implementation and in terms of biosecurity
Even if the product follows regulations, difficulties could arise from the fact that GMOs are an extremely sensitive subject in society
After this interview, we decided to review the risk linked to the potential presence of indoxyl in the last tank. According to Professor Escande, the risk only affects a single staff and the threat will disappear once the compound is oxidized, the risk is confined to a place, the laboratory, and could therefore be downgraded to “severe”. However, there should be emphasis on PPE (Personal Protective Equipment) for it to prevent the worst case scenario from happening. It was decided the staff would have to wear glasses and a mask as well as a blouse and gloves when removing the last tank of the bioreactor.
We realized that to have a more exhaustive understanding of the risks linked to our project, we needed to obtain feedback from people working with similar problematics. We contacted Carolina Lacome, health and security engineer at the University of Technology of Compiègne.
She was able to tell us more about how risks are managed in the industry and gave us specific examples or risks, incidents and their management in factories.
Here is what we should remember:
The risks linked to malicious intent exist, therefore we should identify what could be added to our bioreactor to cause an incident. We did not identify any risk.
Sometimes the probability of a risk is so low, nothing can be done. But the question must be asked and answered
Every risk should be tested, all quantities quantified. Then, the risks can be reevaluated and the safety measures improved. We plan on performing such tests in the future
About indoxyl, she suggested adding not one but two air inputs in the culture. This way, if one were to malfunction the other would ensure the oxidation of indoxyl. She also insisted on the importance of maintenance and verification of the prevention equipment
We went over the entire risk assessment in detail together to ensure it follows the state of the art.
Documentation
An important aspect of risk management is documentation. All normes and recommendations insist on the need of detailed documentation of the organisms used, the different steps of the process containing the protocol, the risks linked to it, the safety procedures and equipment to prevent them, the measures to take in case of an incident.
We created a:
REFERENCES
(1) Khripunov, I., Smidovich, N., & Williams, D. M. (2017). Bio-risk Management Culture: Concept, Model, Assessment. Cyber and Chemical, Biological, Radiological, Nuclear, Explosives Challenges, 199–234. https://doi.org/10.1007/978-3-319-62108-1_10
(2) International Biological Threat Reduction, & Sandia National Laboratories. (2019). Laboratory Biosafety and Biosecurity Risk Assessment Technical Guidance Document [Review of Laboratory Biosafety and Biosecurity Risk Assessment Technical Guidance Document]. In CDC (pp. 5–51). CDC. https://www.cdc.gov/labtraining/docs/resources/Laboratory-Biosafety-and-Biosecurity-Risk-Assessment-Technical-Guidance-Document-PDF.pdf
(3) World Health Organization. (2004). Laboratory biosafety manual Third edition [Review of Laboratory biosafety manual Third edition]. https://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf
(4) US EPA. (1997). ATTACHMENT I--FINAL RISK ASSESSMENT OF ESCHERICHIA COLI K-12 DERIVATIVES [Review of ATTACHMENT I--FINAL RISK ASSESSMENT OF ESCHERICHIA COLI K-12 DERIVATIVES]. https://www.epa.gov/sites/default/files/2015-09/documents/fra004.pdf
(5) KARCHER, S. J. (1995). RECOMBINANT DNA CLONING. Molecular Biology, 45–134. https://doi.org/10.1016/b978-012397720-5.50036-0
(6) Hossain, M. J., Thurlow, C. M., Sun, D., Nasrin, S., & Liles, M. R. (2015). Genome modifications and cloning using a conjugally transferable recombineering system. Biotechnology Reports, 8, 24–35. https://doi.org/10.1016/j.btre.2015.08.005
(7) Smillie, C., Garcillan-Barcia, M. P., Francia, M. V., Rocha, E. P. C., & de la Cruz, F. (2010). Mobility of Plasmids. Microbiology and Molecular Biology Reviews, 74(3), 434–452. https://doi.org/10.1128/mmbr.00020-10