SAFETY

SAFE LAB WORK

Safety is an essential part of any lab project in science. Proper safety regulations and precautions lower the risk of accidents. They are designed to protect the experimenter, the project, the public and the environment. At ETH-DBSSE it is mandatory for every person to take a Safety Course. The course is held by Mr. Niels Buerckert, who is the Technical Manager and head of the department’s Safety Committee. During the Safety Course we first covered basic safety measurements required for both office and lab members of the department. We were instructed how to handle hazardous situations in case of natural disaster or accident. We got acquainted with locations of fire extinguishers, fire blankets and emergency exits. In the case of a small fire, the fire blanket is used. In the case of a bigger fire, the CO2 fire extinguisher is used. If paper or carton is on fire, foam extinguisher is used instead. We were instructed who to call in the case of fire and how to act and who to call in the case of an injury. After we considered different specific situations in basic safety training, we covered lab safety measurements and regulations.

Lab safety measurements tackle every aspect of lab work. The experimenter has to wear proper clothing which covers as much skin as possible. Shorts and open shoes are not allowed. Additionally the experimenter needs to wear a lab coat, gloves and wear goggles on the top of the head. Food and drinks are not allowed in the lab. Smoking is forbidden. We were instructed how to properly label hazardous chemicals and how to act in the case we encounter unfamiliar or unlabeled chemical. In the case where we encounter unknown chemical in solid form we use a brush to gently remove the chemical and dispose it in hazardous waste. Any unlabeled liquid is considered as a potential hazardous chemical. We got acquainted with the specifics of waste management in this building: the location of clean room, how to properly transport chemicals from one part of the building to another, how to prevent potential spread of chemicals from our lab to other areas of the building. Chemicals need to be transported in the chemical bucket. When transferring chemicals from one floor to another, they are not allowed in the personel elevator. The experimenter must send the chemical separately in the elevator for chemicals, while the experimenter uses the personel elevator. Gloves need to be removed after exiting the lab. If the chemicals require handling in gloves, the experimenter must use one hand without a glove to interact with the surroundings (opening doors etc) and one hand with a glove to carry the chemicals. We were taught how to properly maintain a clean lab space and how to correctly store chemicals in the lab to prevent accidents. Every sink has an eye shower and every floor has emergency showers in case a person gets in contact with a hazardous chemical.

SAFETY IN OUR CURRENT PROJECT DESIGN

The goal of our project in the scope of iGEM until the giant jamboree is to design and show that our system works under controlled experimental conditions in the laboratory which would mimic real life conditions. Thus we have no intention to release bacteria with our current design in the environment or to consume it. In our project we chose to work with several different well known lab strains of E. coli: TOP10, DH5alpha, Keio strains and EcNR1. They are all derivatives of K12 E. coli strain and belong to the biosafety level 1. Biosafety level 1 organisms pose little risk to the researcher and the environment. However, we are working with GMO strains which cannot be released into the environment and low risk does not equal zero risk. For this reason we stick to all safety regulations for biosafety level 1 laboratory, such as wearing and frequently changing gloves, wearing lab coat and disinfecting the working area after the experiment. All waste that has been in contact with the bacteria is autoclaved. To mimic conditions in the gut, we chose to work with DETA/NO (as a source of nitric oxide (NO)), homoserine lactone (AHL) and lactate. We took extra precaution in handling experiments where we used DETA/NO or ganciclovir.

SAFETY FEATURES OF OUR FINAL PROJECT DESIGN

The final goal of our project is to design a bacteria-based detection system to simultaneously detect compounds associated with inflammation and microbiota in the gut of an inflammatory bowel disease (IBD) patient. This requires ingestion of our device by the patient. Our bacteria will travel through the human digestive system in a capsule. The encapsulated bacteria will be collected from the feces and will be analyzed in the lab. The administration and recovery of our bacterial device would be done by trained medical personnel. There are several safety risks which we need to consider in the design of our final device.

RISK OF INFECTING HUMAN GUT

The strains of E. coli we use have a low, but not non-existent, virulence. There is a non-zero probability of our strains mutating to a pathogenic serotype. The device in our final design would consist of bacteria encapsulated in a material (chitosan or alginate) which does not dissolve in the stomach or in the gut, but would allow exchange of small molecule signals. This would isolate the bacteria in the device from the gut microbiota. That would lower the risk of infecting human gut and prevent our bacteria from disrupting the native microbiota population. Because Escherichia coli have pathogenic serotypes, it is not an ideal organism for an in situ study of human gut. Escherichia coli might also cause additional disruption of microbiome balance in the IBD patients. In our final design we would replace E. coli with Lactobacillus sp. We decided to take this safety precaution after discussing with prof. Rogler.

RISK OF HORIZONTAL GENE TRANSFER

Horizontal gene transfer could potentially cause antibiotic resistance of gut microbiota. We would use a capsule to prevent interaction between our designed bacteria and the native microbiota. However, this would not eliminate the risk of horizontal gene transfer, which is why we would have to clone our system into a chromosome to eliminate the need for antibiotic resistance and to maximally reduce the risk of gene transfer. Another option would be to use metabolic complementation instead of antibiotic selection for plasmid maintenance.

RISK OF RELEASE IN THE ENVIRONMENT

The capsule would lower the risk of release in the environment by isolating the bacteria from our device from the environment. Since the capsule can be broken or damaged, we would additionally have to develop a knock-out strain which would rely on a metabolite abundantly present in the gut but not in the environment.

Team:ETH Zurich/Safety