Animal Biosafety Levels
As a general principle, the biosafety level recommended for working with infectious agents in vivo and in vitro are comparable. However, the activities of the animals themselves can present unique hazards not found in standard microbiological laboratories. For example, Listeria monocytogenes is a BSL-2/ABSL-2 agent but poses a risk to a fetus and pregnant women should be advised of that risk. Brucella melitensis and Coxiella burnetti are BSL-2 organisms for nonpropagative work but are at ABSL-3 for animal research.
Plant Biosafety Levels
Plant research generally, but not always, does not pose a human health hazard; therefore biosafety principles are designed to protect the natural and agricultural environment. Recombinant DNA practices, under the National Institutes of Health (NIH) Guidelines, drive most of the plant biosafety issues. Field work with genetically engineered plants requires permits from USDA/APHIS before work can begin.
Plant biosafety levels are designated with a “P” after the containment level. These agents do not usually pose a threat to human health; however they may pose a threat to plants and the environment. Plants pathogens can be spread by, direct contact between plants, arthropods, soil borne nematodes, plant damage, and pollinators. Plants can be grown in the greenhouse, laboratory, growth chamber, and or field. NIH Guidelines define a greenhouse as a structure with walls, a roof, and a floor designed and used principally for growing plants in a controlled and protected environment.
The principal purpose of plant containment is to avoid the unintentional transmission of a recombinant or synthetic nucleic acid molecule-containing plant genome, including nuclear or organelle hereditary material or release of recombinant or synthetic nucleic acid molecule-derived organisms associated with plants.
Plant Biosafety Level 1 (BSL-1P)
BSL-P1 is recommended for all experiments with transgenic plants and associated agents that have no or limited threat potential. For example: transgenic plants that are not noxious weeds or agents that have no recognized potential for rapid dissemination. Examples of agents worked with at BSL-1P include Agrobacterium tumefaciens and Rhizobium spp.
Requirements at BSL-1P include:
- Access to the laboratory and greenhouse is limited or restricted when experiments are in progress.
- Prior to entering the greenhouse, personnel are required to read and follow instructions on BSL-1P greenhouse practices and procedures.
- All procedures must be performed in accordance with accepted greenhouse practices appropriate to the experimental organism.
- Records will be kept of experiments currently in progress in the greenhouse facility.
- Render experimental organisms biologically inactive by appropriate methods before disposal.
- A program shall be implemented to control undesired species (e.g., weed, rodent, or arthropod pests and pathogens) by methods appropriate to the organisms and in accordance with applicable state and federal laws.
- House arthropods and other motile macroorganisms in appropriate cages. If macroorganisms (e.g., flying arthropods or nematodes) are released within the greenhouse, precautions must be taken to minimize escape from the greenhouse facility.
- The greenhouse floor may be composed of gravel or other porous material. Impervious (e.g., concrete) walkways are recommended.
- Windows and other openings in the walls and roof of the laboratory and greenhouse facility may be open for ventilation as needed for proper operation and do not require any special barrier to contain or exclude pollen, microorganisms, or small flying animals (e.g., arthropods and birds). Screens are recommended.
- Laboratories and greenhouses must be locked when unoccupied. All agents must be secured against accidental exposure, unauthorized use, and theft. All recombinant nucleic acids must be stored in locked containers.
Plant Biosafety Level 2 (BSL-2P)
Recommended for transgenic plants that are noxious weeds, plants in which the introduced DNA represents the complete genome of a non-exotic infectious agent, plants associated with transgenic non-exotic microbe that has a recognized potential for serious detrimental impact on managed or natural ecosystems, or plant pathogens that have a recognized potential for serious detrimental impact on managed or natural ecosystems. Examples of agents worked with at BSL-2P include: Meliodogyne incognita (root-knot nematode), Pepino mosaic virus (PepMV), Pectinophora gossypiella (Pink bollworm), and Pseudomonas syringae.
The following are required when working at BSL-2P:
- A program to control undesired species (e.g., weed, rodent, or arthropod pests and pathogens) by methods appropriate to the organisms and in accordance with applicable state and Federal laws.
- A greenhouse floor composed of an impervious material. Concrete is recommended, but gravel or other porous material under benches is acceptable unless propagates of experimental organisms are readily disseminated through soil. Soil beds are acceptable unless propagates of experimental organisms are readily disseminated through soil.
- Materials containing experimental microorganisms must be transferred in a closed , leak proof container.
- An autoclave must be available for the treatment of contaminated plant material including soil.
- If intake fans are used, measures shall be taken to minimize the ingress of arthropods. Louvers or fans shall be constructed such that they can only be opened when the fan is in operation.
- Greenhouse containment requirements may be satisfied by using a growth chamber or growth room within a building provided that the external physical structure limits access and escape of microorganisms and macroorganisms in a manner that satisfies the intent of the foregoing clauses
- Laboratories and greenhouses must be locked when unoccupied.
- All agents must be secured against accidental exposure, unauthorized use, and theft.
- All recombinant nucleic acids and BSL-2P agents must be stored in locked containers.
- All material in the open bay or common use areas must be secured when not in use.
Standard Operating Procedures
Standard operating procedures (SOPs) are required for all field work involving transgenic plants and all work at BSL-2P. The SOP’s must be easily accessible in the laboratory or field and contain:
- Location of field site/facility/greenhouse.
- Physical containment standards.
- Devitalization procedures.
- How persistence in the environment is controlled.
- Volunteer management procedures.
- Acknowledgement pages.
Research Laboratory & Safety Services has provided templates for creating the required SOPs:
Field Work Involving Transgenic Plants
The U.S. Department of Agriculture - Animal and Plant Health Inspection Service (USDA/APHIS) regulates plants, plant pests, and plant products. APHIS also regulates the movement, importation, and field release of genetically engineered plants and arthropods. Field work with genetically engineered plants requires permits from APHIS before work can begin. If you are not sure about the required permits, contact Research Laboratory & Safety Services before beginning your project.
The following records must be maintained:
- Log for cleaning equipment after use in the field
- Isolation distance
- Log of volunteer management
- Planting reports and final reports
- Harvesting & destruction reports