Laboratory Biosafety

Laboratory biosafety allows for safe handling of biological material and living organisms including genetically modified organisms (GMOs) in contained laboratory settings to minimize risks to human health and the environment. Authoritative sources on laboratory biosafety including academic and research institutions and internationally recognized organizations such as the World Health Organization (WHO),Organization of Economic Cooperation and Development (OECD), US Centers for Disease Control and Prevention (CDC) and National Institute of Health (NIH) provide excellent guidelines for safe laboratory practices.

The objective of any biosafety program in a laboratory is to ensure containment of potentially harmful biological agents. The term “containment” is used to describe safe methods, facilities and equipment for managing materials in the laboratory where they are handled. The safety measures are meant to protect the laboratory staff, other workers, the experiment, humans and the environment. Greenhouse experiments are also generally carried out and reviewed as contained experiments.

When working on transformation experiments, any hazard will depend on the characteristics of the gene being manipulated and the organism into which it will be introduced. Important characteristics that represent a safety concern are the following:

1. Pathogenicity
   The pathogenicity of an organism indicates whether an organism – for instance a bacterium, a virus, fungus or a parasite – is able to cause a disease in a plant, animal or human. Factors like infectious dose, virulence and the production of toxins by the pathogen play a role in the extent to which the organism is able to cause disease. Note that while these concerns may pertain toplant pathogens, they do not pertain to plants.
2. Toxicity
   Toxicity refers to poisoning. Most substances are not poisonous when they are used under normal circumstances. When the toxicity of living organisms (especially bacteria) is considered, toxicity often coincides with pathogenicity.
3. Allergenicity
   Allergenicity is a non-toxic, immune system mediated, undesired reaction of the body to a substance or agent. An allergic reaction may lead to sneezing, skin irritation, asthma attacks, chronic lung disorders, and sometimes even to a life threatening shock. In order to prevent laboratory staff from contracting lung and skin disorders, the possibility of an allergic reaction should be taken into account.

Working with DNA is not inherently hazardous. Genes are made of DNA. DNA is everywhere, it is contained in anything that is or was alive. The necessary laboratory safety procedures to be employed depend on the nature of what the specific DNA (genes) code for. Risk assessment is necessary in situations where the product of the specific inserted gene has known biologically or pharmacologically active properties such as those described above (e.g., virulence or pathogenicity factors, toxicity, allergenicity) or other biological activities (hormones, cytokines, oncogenes, antibiotic resistance)

• mode and ease of dissemination;
• invasiveness;
• whether a noxious weed or capable of interbreeding with noxious weeds;
• potential for out-crossing between recipient organisms and nearby related species;
• potential for detrimental impact on natural or managed ecosystems.

Although many modifications do not involve genes whose products are inherently harmful, adverse effects may arise as a result of alteration of existing non-pathogenic or pathogenic traits. Modification of normal genes may also alter pathogenicity. To identify these potential hazards, points to be taken into account, include:

• Is there an increase in infectivity or pathogenicity?
• Does the foreign gene encode a pathogenicity determinant from another organism?
• If the foreign DNA does include a pathogenicity determinant, is it foreseeable that this gene could contribute to the pathogenicity of the GMO?

To ensure biosafety in plant transformation experiments, safety measures are instituted in the laboratory or greenhouse depending on the containment level required for the kind of agents being handled. Containment is achieved through a combination of physical containment measures and safe working practices. The containment levels used to categorize laboratories are designated in ascending order, by degree of protection provided to personnel, the environment, and the community. Below are brief descriptions of laboratory biosafety containment levels 1-4. Note that virtually all plant transformation experiments are performed at biosafety level 1 or 2. Levels 3 and 4 apply to laboratories handling human or animal pathogenic agents.

1. Biosafety Level (BS) 1 – BS 1 is suitable for work involving well characterized agents not known to consistently cause disease in healthy adult humans, and of minimal potential hazard to laboratory personnel and the environment.
2. Biosafety Level (BS) 2 – BS 2 is suitable for work involving pathogenic agents that may cause a person (or animal) to be affected by a disease but whose propagation through the community is pretty improbable. The agents in this case pose low risk to personnel and environment.
3. Biosafety Level (BS) 3 – BS 3 is applicable to clinical, diagnostic, teaching, research or production facilities working with GMOs or pathogen agents that may induce a serious disease to humans or animals but for which an efficient treatment is available. The agents in this case pose medium risk to personnel and environment.
4. Biosafety Level (BS) 4 – BS 4 is required for work with GMOs or pathogen agents that induce serious diseases in humans or animals. They are often propagated via air routes and no efficient vaccines nor therapy are available. The agents in this case pose high risk to personnel and environment.

The risk assessment procedure used to determine the level of containment while working with GMOs consists of three steps:

1. Assess characteristics of the host, vector and donor sequences that are potentially hazardous. Considerations include possible pathogenicity, toxicity, or uncontrolled spreading of the organism if it were to escape containment. This leads to a preliminary identification of the risk level.
2. Determine circumstances under which the organisms can be handled safely, taking into account:
   • the characteristics of the environment that could be exposed to the GMOs;
   • the type and scale of the activity;
   • any non-standard activities or actions.
3. Assign a risk class based on the results of the first two steps.

The containment level to be accorded to a laboratory depends upon the kind of plant transformation experiments to be carried out and the potentially hazardous agents to be handled. When determining the biosafety level of the laboratory, the following factors are taken into consideration:

• Source and nature of the introduced DNA:
  • whether the DNA is well characterized;
  • whether from an exotic infectious agent or pathogenic organism;
  • whether a fragment of DNA or complete genome.
• Recipient organism:
  • mode and ease of dissemination;
  • invasiveness;
  • whether a noxious weed or capable of interbreeding with noxious weeds;
  • potential for out-crossing between recipient organisms and nearby related species;
  • potential for detrimental impact on natural or managed ecosystems.
• Nature of expressed protein:
  • whether a vertebrate toxin or potential or known allergen;
  • whether has potential pharmacological activity;
  • whether toxic to other organisms in local environment;
• Local environment:
  • nature and importance of nearby crops;
  • presence of sexually compatible wild or weedy species.
• Experimental procedures: transfer to or from the laboratory and necessary containment measures.

(Note: for relevant greenhouse containment considerations, please click here.)

Biosafety Level (BS) 1

Work is generally conducted on bench tops using standard good laboratory practices (GLPs). The laboratory is not necessarily separated from the general traffic patterns in the building. Special containment equipment or special facility design is neither required nor generally used. Laboratory personnel have specific training in the procedures conducted in the laboratory and are supervised by a scientist with general training in plant transformation.

Biosafety Level (BS) 2

In addition to measures in BS-1:

• Laboratories are clearly labeled with biohazard signs;
• laboratory personnel have specific training in handling pathogenic agents and are directed by competent scientists;
• access to the laboratory is limited when work is being conducted;
• extreme precautions are taken with contaminated sharp items;
• certain procedures are conducted in biological safety cabinets or other physical containment equipment;
• directional airflow to prevent flow of air into other laboratories.
• proper decontamination of waste before disposal.

Biosafety Level (BS) 3

In addition to measures in BS-2:

• laboratory personnel have specific training in handling pathogenic and potentially lethal agents, and are supervised by competent scientists who are experienced in working with these agents;
• all procedures involving the manipulation of infectious materials are conducted within biological safety cabinets or other physical containment device, or by personnel wearing appropriate personnel protective clothing and equipment;
• laboratory fitted with a double entry door;
• laboratory has special engineering and design features;
• a written contingency plan for handling laboratory accidents involving potential hazardous agents is available.

Biosafety Level (BS) 4

In addition to measures in BS-3:

• laboratory staff have specific and thorough training in handling extremely hazardous infectious agents and they understand the primary and secondary containment functions of the standard and special practices, the containment equipment, and the laboratory design characteristics;
• laboratory staff are supervised by competent scientists who are trained and experienced in working with these agents;
• access to the laboratory is strictly controlled by the individual(s) in charge of the laboratory;
• the facility is either in a separate building or in a controlled area within a building, which is completely isolated from all areas of the building;
• a specific facility operations manual is prepared or adopted for working in the laboratory;
• the laboratory has special engineering and design features to prevent agents from being disseminated into the environment.
• within the laboratory appropriate specialized safety cabinets are fitted.