Team:UANL Mty-Mexico/Safety/r

PROJECT: REPROGRAMMATOR
Our project arise safety issues normally associated with working with typical cloning strains of the bacteria of the genus E. coli, Pseudomonas, Bacillus, Serratia and Clavibacter. Thus, in regard to researcher safety, our project pose risks that are contemplated in a normal microbiology and molecular training for a BL-1 and BL-2 laboratories, for example spills, contamination or accidental ingestion. These bacteria have very low competitive advantages against wild-type microorganisms in the case of an accidental release. Appropriate handling measures will be also applied for genetically modified bacteria and materials contaminated with bacteria. For the use of chemical reagents and laboratory equipment we will follow the biosecurity rules imposed by Microbiological and Biomedical Laboratories (HHS 2009) and the WHO.
We are the first line of defense for protecting ourselves, others in the lab, and the public from exposure to hazardous agents. Protection depends on good microbiological practices and the correct use of safety equipment i.e.; always using personal protective equipment in addition to safety glasses, lab coat, gloves, and the safety equipment such as sealed rotors to provide a high degree of protection for us from exposure to microbial aerosols and droplets.
As we know the bacteria of the genus Pseudomonas, Serratia and Clavibacter are classified in level 2 of biosafety level, but only Serratia and Pseudomonas are classified as human pathogens, and these are considered opportunistic pathogens, which can rule out direct infection. So the good laboratory practices will ensure our personal to stay safe and to avoid accidents of biocontainment.
Only the bacteria of the genus Clavibacter is known as infectious to plants, which in the event of a crash would compromise certain species. Through good laboratory procedures, none of the genetically modified bacteria should have a chance of being introduced into the environment. Regarding environmental release, the main concern might be possible horizontal gene transference and possible infections to human and some plants, however any of the strains used can be consider as pathogen for humans or animals or plants. So, the chances for malicious mis-use by other person/group are minimal or null.
At the moment our system does not have any in-built, designed feature meant to alleviate a contingency. However, we have been careful to take measures to avoid any accidental release. Actually, we are working in a system for the specific infection/programmation exclusive for the bacteria strains used in our project.
The project contemplate the use of bacteriophages in order to change the transgenes in bacteria and substitute the resident genetic program for new one with the possibility to change again and over and over as a new program can be generated and insert by phage infection. This project advantage could be potentially risked only if the bacteria/phage specificity can be overturned. Finally, the original program in the receptor bacteria must have specific sequences in order to be deleted by the new incoming program. This make very difficult to widely spread in the environment, as the natural genetic plasmids or genomes, lacks the specific sequence.
The main idea of our project is to develop a specific phage to programme some specific bacteria strains (with unique characteristics) and in this way the interaction of virus-bacteria is able and safe to program and re-programmate the bacteria.

Workplace and Biocontainment
The use of sharp tools or needles that may cause accidental injuries, should be avoided whenever possible. Other practices, such as pipetting, vortexing, and centrifugation, that generate aerosols, thus increasing the probability of exposure and the risk associated to the activity, should be physically contained by using sealed rotors, capped tubes, or biosafety cabinet.
The examination of biological risks related to the use of bacteriophages and the measures taken to reuce these biological risks allow the determination of an adequate containment level to optimally protect human health and the environment against the identified risks. Phages handled in laboratories that are engineered to present no biological risks to human health can be handled in BSL-1. However, the containment level required should be adapted depending on the bacterial strains used to propagate the bacteriophages.
Indeed, bacteriophages represent an important gene reservoir that can be transferred among bacteria, and dissemination of recombinant phages into the environment could have important impacts on the surrounding susceptible bacterial population. The main precaution that has to be taken to avoid such unwanted release consists of properly inactivating all biological wastes generated by the activity that may contain bacteriophages. Bacteriophage inactivation is also important to avoid their dissemination throughout the laboratory (Verheust C., et al 2010).
Cleaning efforts should be as thorough and systematic as possible (work surfaces and all equipment used) to remove any residual phages, and data indicating which disinfectant and inactivation treatment are effective should be readily available for all workers in the form of written standard operating procedures.
Furthermore, since phages can exist for a longer time in wet environments, spills should be immediately cleaned up and any humid area should be eliminated. Good sampling handling and storage techniques and waste management practices are also essential.

• Respect good microbiological practices.
• Use appropriate disinfectant before autoclaving non-disposable equipment.
• Clean working surfaces regularly with adequate disinfecting solution.
• Clean up any spill immediately with a validated disinfectant and disinfection method.
• Avoid procedures generating aerosols.
• Do not open flasks during cultivation unless absolutely necessary.
• Avoid moisture. Dry all glass and plastic materials carefully.

However, even though the probability of dissemination has been evaluated to be negligible, good microbiological practices including appropriate disinfection should be followed. The main biosafety measures recommended to prevent any toxicity or allergenicity towards the worker consist of wearing a lab coat and gloves. Sharp instruments should be avoided if possible.


Hazardous Substances